Structured Review

Epitomics k atpase
Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for <t>NBCe2</t> along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + <t>/ATPase</t> (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.
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Images

1) Product Images from "Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells"

Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells

Journal: PLoS ONE

doi: 10.1371/journal.pone.0189464

Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.
Figure Legend Snippet: Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.

Techniques Used: Western Blot, Marker, Staining

Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).
Figure Legend Snippet: Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).

Techniques Used: Activity Assay

The effect of BI6015 on the expression of Na + ,K + /ATPase (NKA) and the putative anion transporter type 1 (PAT-1). Compared to vehicle control (VEH), neither NKA nor PAT-1 is affected by the addition of the HNF4A inhibitor BI6015. α tubulin was used as a protein loading control.
Figure Legend Snippet: The effect of BI6015 on the expression of Na + ,K + /ATPase (NKA) and the putative anion transporter type 1 (PAT-1). Compared to vehicle control (VEH), neither NKA nor PAT-1 is affected by the addition of the HNF4A inhibitor BI6015. α tubulin was used as a protein loading control.

Techniques Used: Expressing

2) Product Images from "Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells"

Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells

Journal: PLoS ONE

doi: 10.1371/journal.pone.0189464

Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.
Figure Legend Snippet: Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.

Techniques Used: Western Blot, Marker, Staining

Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).
Figure Legend Snippet: Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).

Techniques Used: Activity Assay

3) Product Images from "Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells"

Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells

Journal: PLoS ONE

doi: 10.1371/journal.pone.0189464

Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).
Figure Legend Snippet: Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).

Techniques Used: Activity Assay

Effect of HNF4A inhibitors on basal expression of NHE3, PAT1 and Na + ,K + /ATPase in hRPTCs. A) NHE3 protein expression Basal NHE3 protein expression is greater in HV than WT SLC4A5 hRPTCs (N = 3, *P
Figure Legend Snippet: Effect of HNF4A inhibitors on basal expression of NHE3, PAT1 and Na + ,K + /ATPase in hRPTCs. A) NHE3 protein expression Basal NHE3 protein expression is greater in HV than WT SLC4A5 hRPTCs (N = 3, *P

Techniques Used: Expressing

4) Product Images from "Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells"

Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells

Journal: PLoS ONE

doi: 10.1371/journal.pone.0189464

Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.
Figure Legend Snippet: Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.

Techniques Used: Western Blot, Marker, Staining

Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).
Figure Legend Snippet: Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).

Techniques Used: Activity Assay

5) Product Images from "Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells"

Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells

Journal: PLoS ONE

doi: 10.1371/journal.pone.0189464

Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.
Figure Legend Snippet: Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.

Techniques Used: Western Blot, Marker, Staining

Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).
Figure Legend Snippet: Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).

Techniques Used: Activity Assay

6) Product Images from "Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells"

Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells

Journal: PLoS ONE

doi: 10.1371/journal.pone.0189464

Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.
Figure Legend Snippet: Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.

Techniques Used: Western Blot, Marker, Staining

Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).
Figure Legend Snippet: Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).

Techniques Used: Activity Assay

The effect of BI6015 on the expression of Na + ,K + /ATPase (NKA) and the putative anion transporter type 1 (PAT-1). Compared to vehicle control (VEH), neither NKA nor PAT-1 is affected by the addition of the HNF4A inhibitor BI6015. α tubulin was used as a protein loading control.
Figure Legend Snippet: The effect of BI6015 on the expression of Na + ,K + /ATPase (NKA) and the putative anion transporter type 1 (PAT-1). Compared to vehicle control (VEH), neither NKA nor PAT-1 is affected by the addition of the HNF4A inhibitor BI6015. α tubulin was used as a protein loading control.

Techniques Used: Expressing

Effect of HNF4A inhibitors on basal expression of NHE3, PAT1 and Na + ,K + /ATPase in hRPTCs. A) NHE3 protein expression Basal NHE3 protein expression is greater in HV than WT SLC4A5 hRPTCs (N = 3, *P
Figure Legend Snippet: Effect of HNF4A inhibitors on basal expression of NHE3, PAT1 and Na + ,K + /ATPase in hRPTCs. A) NHE3 protein expression Basal NHE3 protein expression is greater in HV than WT SLC4A5 hRPTCs (N = 3, *P

Techniques Used: Expressing

7) Product Images from "Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells"

Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells

Journal: PLoS ONE

doi: 10.1371/journal.pone.0189464

The effect of BI6015 on the expression of Na + ,K + /ATPase (NKA) and the putative anion transporter type 1 (PAT-1). Compared to vehicle control (VEH), neither NKA nor PAT-1 is affected by the addition of the HNF4A inhibitor BI6015. α tubulin was used as a protein loading control.
Figure Legend Snippet: The effect of BI6015 on the expression of Na + ,K + /ATPase (NKA) and the putative anion transporter type 1 (PAT-1). Compared to vehicle control (VEH), neither NKA nor PAT-1 is affected by the addition of the HNF4A inhibitor BI6015. α tubulin was used as a protein loading control.

Techniques Used: Expressing

8) Product Images from "Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells"

Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells

Journal: PLoS ONE

doi: 10.1371/journal.pone.0189464

Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).
Figure Legend Snippet: Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).

Techniques Used: Activity Assay

Effect of HNF4A inhibitors on basal expression of NHE3, PAT1 and Na + ,K + /ATPase in hRPTCs. A) NHE3 protein expression Basal NHE3 protein expression is greater in HV than WT SLC4A5 hRPTCs (N = 3, *P
Figure Legend Snippet: Effect of HNF4A inhibitors on basal expression of NHE3, PAT1 and Na + ,K + /ATPase in hRPTCs. A) NHE3 protein expression Basal NHE3 protein expression is greater in HV than WT SLC4A5 hRPTCs (N = 3, *P

Techniques Used: Expressing

9) Product Images from "Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells"

Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells

Journal: PLoS ONE

doi: 10.1371/journal.pone.0189464

Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.
Figure Legend Snippet: Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.

Techniques Used: Western Blot, Marker, Staining

Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).
Figure Legend Snippet: Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).

Techniques Used: Activity Assay

The effect of BI6015 on the expression of Na + ,K + /ATPase (NKA) and the putative anion transporter type 1 (PAT-1). Compared to vehicle control (VEH), neither NKA nor PAT-1 is affected by the addition of the HNF4A inhibitor BI6015. α tubulin was used as a protein loading control.
Figure Legend Snippet: The effect of BI6015 on the expression of Na + ,K + /ATPase (NKA) and the putative anion transporter type 1 (PAT-1). Compared to vehicle control (VEH), neither NKA nor PAT-1 is affected by the addition of the HNF4A inhibitor BI6015. α tubulin was used as a protein loading control.

Techniques Used: Expressing

10) Product Images from "Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells"

Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells

Journal: PLoS ONE

doi: 10.1371/journal.pone.0189464

The effect of BI6015 on the expression of Na + ,K + /ATPase (NKA) and the putative anion transporter type 1 (PAT-1). Compared to vehicle control (VEH), neither NKA nor PAT-1 is affected by the addition of the HNF4A inhibitor BI6015. α tubulin was used as a protein loading control.
Figure Legend Snippet: The effect of BI6015 on the expression of Na + ,K + /ATPase (NKA) and the putative anion transporter type 1 (PAT-1). Compared to vehicle control (VEH), neither NKA nor PAT-1 is affected by the addition of the HNF4A inhibitor BI6015. α tubulin was used as a protein loading control.

Techniques Used: Expressing

11) Product Images from "Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells"

Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells

Journal: PLoS ONE

doi: 10.1371/journal.pone.0189464

Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.
Figure Legend Snippet: Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.

Techniques Used: Western Blot, Marker, Staining

Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).
Figure Legend Snippet: Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).

Techniques Used: Activity Assay

12) Product Images from "Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells"

Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells

Journal: PLoS ONE

doi: 10.1371/journal.pone.0189464

Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.
Figure Legend Snippet: Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.

Techniques Used: Western Blot, Marker, Staining

Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).
Figure Legend Snippet: Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).

Techniques Used: Activity Assay

13) Product Images from "Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells"

Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells

Journal: PLoS ONE

doi: 10.1371/journal.pone.0189464

Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.
Figure Legend Snippet: Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.

Techniques Used: Western Blot, Marker, Staining

Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).
Figure Legend Snippet: Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).

Techniques Used: Activity Assay

The effect of BI6015 on the expression of Na + ,K + /ATPase (NKA) and the putative anion transporter type 1 (PAT-1). Compared to vehicle control (VEH), neither NKA nor PAT-1 is affected by the addition of the HNF4A inhibitor BI6015. α tubulin was used as a protein loading control.
Figure Legend Snippet: The effect of BI6015 on the expression of Na + ,K + /ATPase (NKA) and the putative anion transporter type 1 (PAT-1). Compared to vehicle control (VEH), neither NKA nor PAT-1 is affected by the addition of the HNF4A inhibitor BI6015. α tubulin was used as a protein loading control.

Techniques Used: Expressing

Effect of HNF4A inhibitors on basal expression of NHE3, PAT1 and Na + ,K + /ATPase in hRPTCs. A) NHE3 protein expression Basal NHE3 protein expression is greater in HV than WT SLC4A5 hRPTCs (N = 3, *P
Figure Legend Snippet: Effect of HNF4A inhibitors on basal expression of NHE3, PAT1 and Na + ,K + /ATPase in hRPTCs. A) NHE3 protein expression Basal NHE3 protein expression is greater in HV than WT SLC4A5 hRPTCs (N = 3, *P

Techniques Used: Expressing

Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).
Figure Legend Snippet: Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).

Techniques Used: Activity Assay

14) Product Images from "Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells"

Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells

Journal: PLoS ONE

doi: 10.1371/journal.pone.0189464

Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.
Figure Legend Snippet: Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.

Techniques Used: Western Blot, Marker, Staining

Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).
Figure Legend Snippet: Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).

Techniques Used: Activity Assay

The effect of BI6015 on the expression of Na + ,K + /ATPase (NKA) and the putative anion transporter type 1 (PAT-1). Compared to vehicle control (VEH), neither NKA nor PAT-1 is affected by the addition of the HNF4A inhibitor BI6015. α tubulin was used as a protein loading control.
Figure Legend Snippet: The effect of BI6015 on the expression of Na + ,K + /ATPase (NKA) and the putative anion transporter type 1 (PAT-1). Compared to vehicle control (VEH), neither NKA nor PAT-1 is affected by the addition of the HNF4A inhibitor BI6015. α tubulin was used as a protein loading control.

Techniques Used: Expressing

15) Product Images from "Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells"

Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells

Journal: PLoS ONE

doi: 10.1371/journal.pone.0189464

Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.
Figure Legend Snippet: Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.

Techniques Used: Western Blot, Marker, Staining

Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).
Figure Legend Snippet: Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).

Techniques Used: Activity Assay

16) Product Images from "Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells"

Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells

Journal: PLoS ONE

doi: 10.1371/journal.pone.0189464

Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.
Figure Legend Snippet: Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.

Techniques Used: Western Blot, Marker, Staining

Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).
Figure Legend Snippet: Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).

Techniques Used: Activity Assay

17) Product Images from "Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells"

Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells

Journal: PLoS ONE

doi: 10.1371/journal.pone.0189464

Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.
Figure Legend Snippet: Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.

Techniques Used: Western Blot, Marker, Staining

Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).
Figure Legend Snippet: Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).

Techniques Used: Activity Assay

18) Product Images from "Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells"

Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells

Journal: PLoS ONE

doi: 10.1371/journal.pone.0189464

Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.
Figure Legend Snippet: Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.

Techniques Used: Western Blot, Marker, Staining

Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).
Figure Legend Snippet: Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).

Techniques Used: Activity Assay

19) Product Images from "Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells"

Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells

Journal: PLoS ONE

doi: 10.1371/journal.pone.0189464

Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.
Figure Legend Snippet: Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.

Techniques Used: Western Blot, Marker, Staining

Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).
Figure Legend Snippet: Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).

Techniques Used: Activity Assay

20) Product Images from "Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells"

Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells

Journal: PLoS ONE

doi: 10.1371/journal.pone.0189464

Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.
Figure Legend Snippet: Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.

Techniques Used: Western Blot, Marker, Staining

Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).
Figure Legend Snippet: Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).

Techniques Used: Activity Assay

The effect of BI6015 on the expression of Na + ,K + /ATPase (NKA) and the putative anion transporter type 1 (PAT-1). Compared to vehicle control (VEH), neither NKA nor PAT-1 is affected by the addition of the HNF4A inhibitor BI6015. α tubulin was used as a protein loading control.
Figure Legend Snippet: The effect of BI6015 on the expression of Na + ,K + /ATPase (NKA) and the putative anion transporter type 1 (PAT-1). Compared to vehicle control (VEH), neither NKA nor PAT-1 is affected by the addition of the HNF4A inhibitor BI6015. α tubulin was used as a protein loading control.

Techniques Used: Expressing

21) Product Images from "Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells"

Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells

Journal: PLoS ONE

doi: 10.1371/journal.pone.0189464

Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.
Figure Legend Snippet: Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.

Techniques Used: Western Blot, Marker, Staining

Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).
Figure Legend Snippet: Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).

Techniques Used: Activity Assay

The effect of BI6015 on the expression of Na + ,K + /ATPase (NKA) and the putative anion transporter type 1 (PAT-1). Compared to vehicle control (VEH), neither NKA nor PAT-1 is affected by the addition of the HNF4A inhibitor BI6015. α tubulin was used as a protein loading control.
Figure Legend Snippet: The effect of BI6015 on the expression of Na + ,K + /ATPase (NKA) and the putative anion transporter type 1 (PAT-1). Compared to vehicle control (VEH), neither NKA nor PAT-1 is affected by the addition of the HNF4A inhibitor BI6015. α tubulin was used as a protein loading control.

Techniques Used: Expressing

Effect of HNF4A inhibitors on basal expression of NHE3, PAT1 and Na + ,K + /ATPase in hRPTCs. A) NHE3 protein expression Basal NHE3 protein expression is greater in HV than WT SLC4A5 hRPTCs (N = 3, *P
Figure Legend Snippet: Effect of HNF4A inhibitors on basal expression of NHE3, PAT1 and Na + ,K + /ATPase in hRPTCs. A) NHE3 protein expression Basal NHE3 protein expression is greater in HV than WT SLC4A5 hRPTCs (N = 3, *P

Techniques Used: Expressing

Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).
Figure Legend Snippet: Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).

Techniques Used: Activity Assay

Related Articles

other:

Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells
Article Snippet: The blot was probed with NBCe2 [ ], CD-13 (APN), Na+ ,K+ /ATPase, PAT-1, and NBCe1 antibodies.

Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells
Article Snippet: Cells were loaded with the sodium sensitive ratiometric dye SBFI and sodium influx was initiated by inhibiting α- Na + /,K + /ATPase with ouabain (100 μmol/L) and measuring the amount of sodium accumulation inside the cells.

Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells
Article Snippet: The dopamine-1 receptor (D1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na+ ,K+ /ATPase (without the red line, for simplicity) activities resulting in reduced reabsorption and increased Na+ excretion.

Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells
Article Snippet: WC homogenates demonstrated the presence of NBCe2, NBCe1, CD-13, and Na+ ,K+ /ATPase, as expected.

Activity Assay:

Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells
Article Snippet: .. Because basolateral exit of sodium was prevented by inhibiting Na+ ,K+ /ATPase with ouabain, this result suggested that sodium entry at the luminal membrane is increased that could be due, in part, to increased NBCe2 activity in HV SLC4A5 hRPTCs. .. Because the activity of NHE3 and NBCe2 may be tightly linked and protein activity is influenced by protein expression, we measured NHE3 expression in WT and HV SLC4A5 hRPTC lines.

Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells
Article Snippet: .. Because there were no differences between HV and WT hRPTCs in basal HNF4A binding by ChIP assay as well as basal expression of NHE3 protein, sodium transport and Cl-/HCO3 exchange activity, we tested the effect of HNF4A inhibitors under basal conditions. αNa+ , K+ /ATPase and PAT1 expressions both with and without the HNF4A inhibitor BIM6015 was measured by western blot ( ). .. The basal protein expression of α- Na + /,K + /ATPase (NKA) or PAT1 expression was not changed by HNF4A inhibition.

Expressing:

Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells
Article Snippet: .. Because there were no differences between HV and WT hRPTCs in basal HNF4A binding by ChIP assay as well as basal expression of NHE3 protein, sodium transport and Cl-/HCO3 exchange activity, we tested the effect of HNF4A inhibitors under basal conditions. αNa+ , K+ /ATPase and PAT1 expressions both with and without the HNF4A inhibitor BIM6015 was measured by western blot ( ). .. The basal protein expression of α- Na + /,K + /ATPase (NKA) or PAT1 expression was not changed by HNF4A inhibition.

Marker:

Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells
Article Snippet: .. CD-13 (APN) was used as an hRPTC microvilli marker and Na+ ,K+ /ATPase and NBCe1 were used as hRPTC basolateral membrane markers. .. Overexpressed (OE): A Lentiviral construct (CCSB-Broad Lentiviral Expression Human SLC4A5 Clone; CloneId: ccsbBroad304_12409) was purchased from Thermo Scientific.

Western Blot:

Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells
Article Snippet: .. Because there were no differences between HV and WT hRPTCs in basal HNF4A binding by ChIP assay as well as basal expression of NHE3 protein, sodium transport and Cl-/HCO3 exchange activity, we tested the effect of HNF4A inhibitors under basal conditions. αNa+ , K+ /ATPase and PAT1 expressions both with and without the HNF4A inhibitor BIM6015 was measured by western blot ( ). .. The basal protein expression of α- Na + /,K + /ATPase (NKA) or PAT1 expression was not changed by HNF4A inhibition.

Binding Assay:

Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells
Article Snippet: .. Because there were no differences between HV and WT hRPTCs in basal HNF4A binding by ChIP assay as well as basal expression of NHE3 protein, sodium transport and Cl-/HCO3 exchange activity, we tested the effect of HNF4A inhibitors under basal conditions. αNa+ , K+ /ATPase and PAT1 expressions both with and without the HNF4A inhibitor BIM6015 was measured by western blot ( ). .. The basal protein expression of α- Na + /,K + /ATPase (NKA) or PAT1 expression was not changed by HNF4A inhibition.

Chromatin Immunoprecipitation:

Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells
Article Snippet: .. Because there were no differences between HV and WT hRPTCs in basal HNF4A binding by ChIP assay as well as basal expression of NHE3 protein, sodium transport and Cl-/HCO3 exchange activity, we tested the effect of HNF4A inhibitors under basal conditions. αNa+ , K+ /ATPase and PAT1 expressions both with and without the HNF4A inhibitor BIM6015 was measured by western blot ( ). .. The basal protein expression of α- Na + /,K + /ATPase (NKA) or PAT1 expression was not changed by HNF4A inhibition.

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    Epitomics k atpase
    Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for <t>NBCe2</t> along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + <t>/ATPase</t> (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.
    K Atpase, supplied by Epitomics, used in various techniques. Bioz Stars score: 92/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/k atpase/product/Epitomics
    Average 92 stars, based on 2 article reviews
    Price from $9.99 to $1999.99
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    Epitomics na k atpase
    Significant increase in K7-positive ductal population in the NOD/ShiLtJ SMG. ( A ) NOD/ShiLtJ and CD-1 mouse SMG tissue sections were subjected to IF for the ductal marker, K7 (green), acinar marker aquaporin 5 (Aqp5) (red), and epithelial marker <t>Na/K-ATPase</t>
    Na K Atpase, supplied by Epitomics, used in various techniques. Bioz Stars score: 90/100, based on 2 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/na k atpase/product/Epitomics
    Average 90 stars, based on 2 article reviews
    Price from $9.99 to $1999.99
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    Image Search Results


    Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.

    Journal: PLoS ONE

    Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells

    doi: 10.1371/journal.pone.0189464

    Figure Lengend Snippet: Western blot of whole cell (WC) homogenates and apical membrane fractions from two immortalized hRPTC lines grown in Transwell membranes. Arrows on the left-hand side of the blots indicate the molecular sizes, while the arrows on the right-hand side indicate the molecular sizes of the bands of the proteins of interest. The blot was probed for NBCe2 along with the following membrane markers: CD-13 (APN microvilli marker), Na + ,K + /ATPase (basolateral membrane marker), and NBCe1 (basolateral membrane marker). The results demonstrate that two isoforms of NBCe2 exist in the apical membrane of polarized RPTCs grown on Transwells. The control western blots demonstrate that αNa + ,K + /ATPase (NKA) and NBCe1 only stain weakly in the apical fraction, probably due to some basolateral contamination inherent in this kind of preparation. WC homogenates demonstrate the presence of NBCe2, NBCe1, CD-13, and Na + ,K + /ATPase, as expected.

    Article Snippet: The blot was probed with NBCe2 [ ], CD-13 (APN), Na+ ,K+ /ATPase, PAT-1, and NBCe1 antibodies.

    Techniques: Western Blot, Marker, Staining

    Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).

    Journal: PLoS ONE

    Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells

    doi: 10.1371/journal.pone.0189464

    Figure Lengend Snippet: Models of ion transport in hRPTCs. This is a model of the hRPTC with the apical (brush border, facing the lumen) (left hand side) and the basolateral side (right hand side). The principal ion transporters and some of the receptors that regulate them are shown. Starting at 11 o’clock in blue is shown the classic pathway for transporting bicarbonate (HCO 3 - ) into the cell. Filtered NaHCO 3 dissociates into Na + and HCO 3 . HCO 3 - in the luminal fluid and H + secreted into the lumen form H 2 CO 3 . Carbonic anhydrase type 4 (CA IV) in the luminal membrane catalyzes the conversion of H 2 CO 3 to H 2 O and CO 2 .CO 2 diffuses inside the hRPTC where intracellular carbonic anhydrase type 2 (CA II) catalyzes the conversion of CO 2 and H 2 O into H 2 CO 3 which then dissociates into HCO 3 - and H + . At 9 o’clock is NHE3 which exchanges one Na + from the lumen with one H + inside the hRPTC. At 7 o’clock HCO 3 - Cl - exchanger (PAT1) is depicted which exchanges luminal Cl - with cytoplasmic HCO 3 - . At 3 o’clock is depicted NBCe1 at the basolateral membrane which electrogenically transports 2–3 Na + and one HCO 3 - into the basolateral space. At 4 o’clock is Na + , K + /ATPase which pumps 3 Na + out of the cell into the blood stream and pumps in 2 K + inside the cell The topic of this manuscript deals with NBCe2, drawn at 8 o’clock. Under a normal sodium load it plays a minor role in Na + and HCO 3 - transport into the hRPTC. There are various plasma membrane receptors that regulate some of these transporters/ exchanger/pumps. The dopamine-1 receptor (D 1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na + , K + /ATPase (without the red line, for simplicity) activities resulting in reduced Na + reabsorption and increased Na + excretion. The AT 1 R (5 o’clock) increases Na + , K + /ATPase activity (green arrow) resulting in increased Na + reabsorption. An increase in intracellular Na + increases Na + , K + /ATPase activity (5 o’clock) that is abetted by AT 1 R (green arrow) resulting in increased Na + transport from inside the cell to the basolateral space. The D 1 R and AT 1 R oppose each other. The D 1 R inhibits the AT 1 R, resulting in reduced Na + transport. We showed that NBCe2 is not affected by stimulation of the D 1 R or AT 1 R. Under basal conditions, NBCe1 is more active than NBCe2 (depicted as relatively larger directional transport arrows).

    Article Snippet: The blot was probed with NBCe2 [ ], CD-13 (APN), Na+ ,K+ /ATPase, PAT-1, and NBCe1 antibodies.

    Techniques: Activity Assay

    The effect of BI6015 on the expression of Na + ,K + /ATPase (NKA) and the putative anion transporter type 1 (PAT-1). Compared to vehicle control (VEH), neither NKA nor PAT-1 is affected by the addition of the HNF4A inhibitor BI6015. α tubulin was used as a protein loading control.

    Journal: PLoS ONE

    Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells

    doi: 10.1371/journal.pone.0189464

    Figure Lengend Snippet: The effect of BI6015 on the expression of Na + ,K + /ATPase (NKA) and the putative anion transporter type 1 (PAT-1). Compared to vehicle control (VEH), neither NKA nor PAT-1 is affected by the addition of the HNF4A inhibitor BI6015. α tubulin was used as a protein loading control.

    Article Snippet: The blot was probed with NBCe2 [ ], CD-13 (APN), Na+ ,K+ /ATPase, PAT-1, and NBCe1 antibodies.

    Techniques: Expressing

    Effect of HNF4A inhibitors on basal expression of NHE3, PAT1 and Na + ,K + /ATPase in hRPTCs. A) NHE3 protein expression Basal NHE3 protein expression is greater in HV than WT SLC4A5 hRPTCs (N = 3, *P

    Journal: PLoS ONE

    Article Title: Sodium bicarbonate cotransporter NBCe2 gene variants increase sodium and bicarbonate transport in human renal proximal tubule cells

    doi: 10.1371/journal.pone.0189464

    Figure Lengend Snippet: Effect of HNF4A inhibitors on basal expression of NHE3, PAT1 and Na + ,K + /ATPase in hRPTCs. A) NHE3 protein expression Basal NHE3 protein expression is greater in HV than WT SLC4A5 hRPTCs (N = 3, *P

    Article Snippet: The dopamine-1 receptor (D1 R) (ten o’clock) when stimulated with dopamine (green box) inhibits (red lines) both NHE3 and Na+ ,K+ /ATPase (without the red line, for simplicity) activities resulting in reduced reabsorption and increased Na+ excretion.

    Techniques: Expressing

    Significant increase in K7-positive ductal population in the NOD/ShiLtJ SMG. ( A ) NOD/ShiLtJ and CD-1 mouse SMG tissue sections were subjected to IF for the ductal marker, K7 (green), acinar marker aquaporin 5 (Aqp5) (red), and epithelial marker Na/K-ATPase

    Journal: Anatomical record (Hoboken, N.J. : 2007)

    Article Title: Changes in the Submandibular Salivary Gland Epithelial Cell Subpopulations During Progression of Sjögren’s Syndrome-Like Disease in the NOD/ShiLtJ Mouse Model

    doi: 10.1002/ar.23190

    Figure Lengend Snippet: Significant increase in K7-positive ductal population in the NOD/ShiLtJ SMG. ( A ) NOD/ShiLtJ and CD-1 mouse SMG tissue sections were subjected to IF for the ductal marker, K7 (green), acinar marker aquaporin 5 (Aqp5) (red), and epithelial marker Na/K-ATPase

    Article Snippet: Both human and mouse TMA slides were subjected to multiplexed immunofluorescence with antibodies to detect epithelial cadherin (ECAD) (BD Biosciences, 610182), smooth muscle α-actin (SM α-actin) (Sigma, C6198), cytokeratin 7 (K7) (Abcam, Cambridge MA, ab9021), cytokeratin 5 (K5) (Covance, Princeton NJ, PRB160P), CD4 (Novus Biologicals, Littleton CO, NBP1-19371, Human Only), CD45R (Millipore, Billerica MA, 557683, Mouse Only), aquaporin 5 (Aqp5) (Alomone, aqp-005), Na+ /K+ -ATPase (Epitomics 2047-1), and nuclei were stained using DAPI (Life Technologies, Carlsbad CA, D1306).

    Techniques: Marker