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    Echelon Biosciences pi 3 5 p 2
    ( A ) Representative basal I TPC2 step currents elicited by a voltage step protocol in the whole-endolysosome (EL) configuration. Voltage steps from −140 to 100 mV with a voltage increment (∆V) of 20 mV for 0.5 s were used to elicit I TPC2 in A-D. HP = 0 mV. Unless otherwise indicated, symmetric (bath/cytosol vs pipette/lumen) Na + (150 mM) solutions were used for all whole-endolysosome recordings, and PI(3,5)P 2 (0.3 µM), LyNa-VA1.2 (100 µM), and LyNA1 (300 µM) were bath- applied to induce I TPC2 . ( B–D ) Representative I TPC2 step currents activated by PI(3,5)P 2 ( B ), LyNa-VA1.2 ( C ), and LyNA1 ( D ). ( E ) Representative normalized I-V plots based on the instantaneous currents activated by various agonists. ( F ) Rectification index, calculated as the ratio of the current amplitudes between +80 and −80 mV, of PI(3,5)P 2 -, LyNa-VA1.2-, and LyNA1- activated I TPC2 . ( G ) The inactivation of I TPC2 at −120 mV was quantified as the ratio of current amplitudes at 10 vs. 500 ms, based on step currents in B , ( C ) and D. ( H ) Voltage steps from −120 to 100 mV (∆V = 20 mV) for 1 s were used to elicit tail currents at −120 mV shown in ( I ) and ( J ). ( I ) The tail currents of PI(3,5)P 2 - evoked whole-endolysosome I TPC2 . ( J ) The tail currents of LyNa-VA1.2- activated whole-endolysosome I TPC2 . Arrows in ( I ) and ( J ) indicate where the currents were measured to calculate the channel conductance (G = I/V). ( K ) Normalized G-V curves of PI(3,5)P 2 - and LyNa-VA1.2- activated I TPC2 . LyNa-VA1.2 activated I TPC2 in a voltage dependent manner with a V 1/2 = −20.3 ± 3.5 mV (n = 5 patches). For panels F and G , individual data and Mean ± S.E.M. are presented. ***, p<0.001. Individual data for ( F ) and ( G ) are presented in  .  Figure 2—source data 1. The inactivation of I TPC2 and rectification index of TPC2.
    Pi 3 5 P 2, supplied by Echelon Biosciences, used in various techniques. Bioz Stars score: 94/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    86
    Bio-Rad p 3504 10g
    ( A ) Representative basal I TPC2 step currents elicited by a voltage step protocol in the whole-endolysosome (EL) configuration. Voltage steps from −140 to 100 mV with a voltage increment (∆V) of 20 mV for 0.5 s were used to elicit I TPC2 in A-D. HP = 0 mV. Unless otherwise indicated, symmetric (bath/cytosol vs pipette/lumen) Na + (150 mM) solutions were used for all whole-endolysosome recordings, and PI(3,5)P 2 (0.3 µM), LyNa-VA1.2 (100 µM), and LyNA1 (300 µM) were bath- applied to induce I TPC2 . ( B–D ) Representative I TPC2 step currents activated by PI(3,5)P 2 ( B ), LyNa-VA1.2 ( C ), and LyNA1 ( D ). ( E ) Representative normalized I-V plots based on the instantaneous currents activated by various agonists. ( F ) Rectification index, calculated as the ratio of the current amplitudes between +80 and −80 mV, of PI(3,5)P 2 -, LyNa-VA1.2-, and LyNA1- activated I TPC2 . ( G ) The inactivation of I TPC2 at −120 mV was quantified as the ratio of current amplitudes at 10 vs. 500 ms, based on step currents in B , ( C ) and D. ( H ) Voltage steps from −120 to 100 mV (∆V = 20 mV) for 1 s were used to elicit tail currents at −120 mV shown in ( I ) and ( J ). ( I ) The tail currents of PI(3,5)P 2 - evoked whole-endolysosome I TPC2 . ( J ) The tail currents of LyNa-VA1.2- activated whole-endolysosome I TPC2 . Arrows in ( I ) and ( J ) indicate where the currents were measured to calculate the channel conductance (G = I/V). ( K ) Normalized G-V curves of PI(3,5)P 2 - and LyNa-VA1.2- activated I TPC2 . LyNa-VA1.2 activated I TPC2 in a voltage dependent manner with a V 1/2 = −20.3 ± 3.5 mV (n = 5 patches). For panels F and G , individual data and Mean ± S.E.M. are presented. ***, p<0.001. Individual data for ( F ) and ( G ) are presented in  .  Figure 2—source data 1. The inactivation of I TPC2 and rectification index of TPC2.
    P 3504 10g, supplied by Bio-Rad, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    86
    Millipore p3504
    KEY RESOURCES TABLE
    P3504, supplied by Millipore, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore p3504 10g

    P3504 10g, supplied by Millipore, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore p3504 100g

    P3504 100g, supplied by Millipore, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    ( A ) Representative basal I TPC2 step currents elicited by a voltage step protocol in the whole-endolysosome (EL) configuration. Voltage steps from −140 to 100 mV with a voltage increment (∆V) of 20 mV for 0.5 s were used to elicit I TPC2 in A-D. HP = 0 mV. Unless otherwise indicated, symmetric (bath/cytosol vs pipette/lumen) Na + (150 mM) solutions were used for all whole-endolysosome recordings, and PI(3,5)P 2 (0.3 µM), LyNa-VA1.2 (100 µM), and LyNA1 (300 µM) were bath- applied to induce I TPC2 . ( B–D ) Representative I TPC2 step currents activated by PI(3,5)P 2 ( B ), LyNa-VA1.2 ( C ), and LyNA1 ( D ). ( E ) Representative normalized I-V plots based on the instantaneous currents activated by various agonists. ( F ) Rectification index, calculated as the ratio of the current amplitudes between +80 and −80 mV, of PI(3,5)P 2 -, LyNa-VA1.2-, and LyNA1- activated I TPC2 . ( G ) The inactivation of I TPC2 at −120 mV was quantified as the ratio of current amplitudes at 10 vs. 500 ms, based on step currents in B , ( C ) and D. ( H ) Voltage steps from −120 to 100 mV (∆V = 20 mV) for 1 s were used to elicit tail currents at −120 mV shown in ( I ) and ( J ). ( I ) The tail currents of PI(3,5)P 2 - evoked whole-endolysosome I TPC2 . ( J ) The tail currents of LyNa-VA1.2- activated whole-endolysosome I TPC2 . Arrows in ( I ) and ( J ) indicate where the currents were measured to calculate the channel conductance (G = I/V). ( K ) Normalized G-V curves of PI(3,5)P 2 - and LyNa-VA1.2- activated I TPC2 . LyNa-VA1.2 activated I TPC2 in a voltage dependent manner with a V 1/2 = −20.3 ± 3.5 mV (n = 5 patches). For panels F and G , individual data and Mean ± S.E.M. are presented. ***, p<0.001. Individual data for ( F ) and ( G ) are presented in  .  Figure 2—source data 1. The inactivation of I TPC2 and rectification index of TPC2.

    Journal: eLife

    Article Title: Agonist-specific voltage-dependent gating of lysosomal two-pore Na + channels

    doi: 10.7554/eLife.51423

    Figure Lengend Snippet: ( A ) Representative basal I TPC2 step currents elicited by a voltage step protocol in the whole-endolysosome (EL) configuration. Voltage steps from −140 to 100 mV with a voltage increment (∆V) of 20 mV for 0.5 s were used to elicit I TPC2 in A-D. HP = 0 mV. Unless otherwise indicated, symmetric (bath/cytosol vs pipette/lumen) Na + (150 mM) solutions were used for all whole-endolysosome recordings, and PI(3,5)P 2 (0.3 µM), LyNa-VA1.2 (100 µM), and LyNA1 (300 µM) were bath- applied to induce I TPC2 . ( B–D ) Representative I TPC2 step currents activated by PI(3,5)P 2 ( B ), LyNa-VA1.2 ( C ), and LyNA1 ( D ). ( E ) Representative normalized I-V plots based on the instantaneous currents activated by various agonists. ( F ) Rectification index, calculated as the ratio of the current amplitudes between +80 and −80 mV, of PI(3,5)P 2 -, LyNa-VA1.2-, and LyNA1- activated I TPC2 . ( G ) The inactivation of I TPC2 at −120 mV was quantified as the ratio of current amplitudes at 10 vs. 500 ms, based on step currents in B , ( C ) and D. ( H ) Voltage steps from −120 to 100 mV (∆V = 20 mV) for 1 s were used to elicit tail currents at −120 mV shown in ( I ) and ( J ). ( I ) The tail currents of PI(3,5)P 2 - evoked whole-endolysosome I TPC2 . ( J ) The tail currents of LyNa-VA1.2- activated whole-endolysosome I TPC2 . Arrows in ( I ) and ( J ) indicate where the currents were measured to calculate the channel conductance (G = I/V). ( K ) Normalized G-V curves of PI(3,5)P 2 - and LyNa-VA1.2- activated I TPC2 . LyNa-VA1.2 activated I TPC2 in a voltage dependent manner with a V 1/2 = −20.3 ± 3.5 mV (n = 5 patches). For panels F and G , individual data and Mean ± S.E.M. are presented. ***, p<0.001. Individual data for ( F ) and ( G ) are presented in . Figure 2—source data 1. The inactivation of I TPC2 and rectification index of TPC2.

    Article Snippet: Chemical compound, drug , PI(3,5)P 2 , Echelon Biosciences , Cat. #: P-3508 , .

    Techniques: Transferring

    ( A ) The effects of PI(3,5)P 2 (0.3 µM) and LyNa-VA1.2 (100 µM) on whole-endolysosome I TPC2-K204A in TPC2 K204A -transfected HEK293 cells . ( B ) Comparison effects of LyNa-VA1.2 and PI(3,5)P 2 on WT and PI(3,5)P 2 -insensitive K204A  mutant TPC2 channels (also see  ). ( C ) The synergistic effects of PI(3,5)P 2 (50 nM) and LyNa-VA1.1 on whole-endolysosome I TPC2 and I TPC2-K204A currents. Data are presented as Mean ± S.E.M (n = 3 patches). ( D ) The summary of EC 50 of LyNa-VA1.1 with or without PI(3,5)P 2 for WT and K204A mutant TPC2 channels. ( E, F ) Co-application of PI(3,5)P 2 (0.3 µM) and LyNa-VA1.1 (50 µM) activated whole-endolysome I TPC in WT ( E ) but not TPC1/2 DKO ( F ) HAP1 cells. Note that the endogenous TPCs were more difficult to activate compared to overexpressed TPCs. ( G ) Summary of LyNa-VA1.1 effects on whole-endolysome I TPC in WT and TPC1/2 DKO cells. For panels B, D and F, individual data and Mean ± S.E.M are presented. *, p<0.05; **, p<0.01; ***, p<0.001; N.S., no significance. Individual data for ( B–D ) and ( G ) are presented in  .  Figure 3—source data 1. Synergistic activation of TPC2 channels by TCAs and PI(3,5)P 2 .

    Journal: eLife

    Article Title: Agonist-specific voltage-dependent gating of lysosomal two-pore Na + channels

    doi: 10.7554/eLife.51423

    Figure Lengend Snippet: ( A ) The effects of PI(3,5)P 2 (0.3 µM) and LyNa-VA1.2 (100 µM) on whole-endolysosome I TPC2-K204A in TPC2 K204A -transfected HEK293 cells . ( B ) Comparison effects of LyNa-VA1.2 and PI(3,5)P 2 on WT and PI(3,5)P 2 -insensitive K204A mutant TPC2 channels (also see ). ( C ) The synergistic effects of PI(3,5)P 2 (50 nM) and LyNa-VA1.1 on whole-endolysosome I TPC2 and I TPC2-K204A currents. Data are presented as Mean ± S.E.M (n = 3 patches). ( D ) The summary of EC 50 of LyNa-VA1.1 with or without PI(3,5)P 2 for WT and K204A mutant TPC2 channels. ( E, F ) Co-application of PI(3,5)P 2 (0.3 µM) and LyNa-VA1.1 (50 µM) activated whole-endolysome I TPC in WT ( E ) but not TPC1/2 DKO ( F ) HAP1 cells. Note that the endogenous TPCs were more difficult to activate compared to overexpressed TPCs. ( G ) Summary of LyNa-VA1.1 effects on whole-endolysome I TPC in WT and TPC1/2 DKO cells. For panels B, D and F, individual data and Mean ± S.E.M are presented. *, p<0.05; **, p<0.01; ***, p<0.001; N.S., no significance. Individual data for ( B–D ) and ( G ) are presented in . Figure 3—source data 1. Synergistic activation of TPC2 channels by TCAs and PI(3,5)P 2 .

    Article Snippet: Chemical compound, drug , PI(3,5)P 2 , Echelon Biosciences , Cat. #: P-3508 , .

    Techniques: Transfection, Mutagenesis, Activation Assay

    ( A ) Basal whole-endolysosome I TPC1 (left) or I TPC1-R540I (right) was elicited by voltage steps (−140 to +100 mV with ΔV = 20 mV) in the absence of PI(3,5)P 2 in TPC1-transfected Cos1 cells . Tail currents were elicited at −120 mV. ( B ) The effects of PI(3,5)P 2 (0.3 µM) on I TPC1 (left) and I TPC1-R540I (right) step currents. ( C ) I-V plots of steady-state I TPC1 and I TPC1-R540I step currents shown in B. ( D ) LyNa-VA1.2-induced whole-endolysosome I TPC2-I551R was evoked by a voltage ramp from −120 to +120 mV (duration = 200 ms). ( E ) The basal (right), LyNa-VA1.2 (200 µM)- (middle) and PI(3,5)P 2 (0.3 µM)- (left) activated lysosomal I TPC1-I551R were recorded from the same vacuole. A preconditioning voltage (+80 mV, 0.1 s) was applied before voltage steps starting from −140 to 100 mV (0.5 s, ∆V = 20 mV). HP = 0 mV.

    Journal: eLife

    Article Title: Agonist-specific voltage-dependent gating of lysosomal two-pore Na + channels

    doi: 10.7554/eLife.51423

    Figure Lengend Snippet: ( A ) Basal whole-endolysosome I TPC1 (left) or I TPC1-R540I (right) was elicited by voltage steps (−140 to +100 mV with ΔV = 20 mV) in the absence of PI(3,5)P 2 in TPC1-transfected Cos1 cells . Tail currents were elicited at −120 mV. ( B ) The effects of PI(3,5)P 2 (0.3 µM) on I TPC1 (left) and I TPC1-R540I (right) step currents. ( C ) I-V plots of steady-state I TPC1 and I TPC1-R540I step currents shown in B. ( D ) LyNa-VA1.2-induced whole-endolysosome I TPC2-I551R was evoked by a voltage ramp from −120 to +120 mV (duration = 200 ms). ( E ) The basal (right), LyNa-VA1.2 (200 µM)- (middle) and PI(3,5)P 2 (0.3 µM)- (left) activated lysosomal I TPC1-I551R were recorded from the same vacuole. A preconditioning voltage (+80 mV, 0.1 s) was applied before voltage steps starting from −140 to 100 mV (0.5 s, ∆V = 20 mV). HP = 0 mV.

    Article Snippet: Chemical compound, drug , PI(3,5)P 2 , Echelon Biosciences , Cat. #: P-3508 , .

    Techniques: Transfection

    ( A ) Representative PI(3,5)P 2 -evoked whole-endolysosome I TPC2 elicited by a voltage ramp from −120 to 120 mV. The recordings were performed under a bi-ionic condition with 150 (in mM) Na + in the pipette solution and 150 Na + or 150 K + in the bath solution. PI(3,5)P 2 (0.3 µM) was bath-applied. ( B, C ) Representative PI(3,5)P 2 - evoked I TPC2 elicited with voltage steps (−140 mV to 100 mV with a ∆V = 20 mV, 0.5 s) with 150 mM Na + ( B ) or K + ( C ) in the bath solution. ( D, E ) Representative LyNa-VA1.2-activated I TPC2 step currents. ( F ) Representative I-V plots of LyNa-VA1.2- activated I TPC2 measured from the instantaneous currents in ( D ) and ( E ). Note the reversal potentials (E rev ) of LyNa-VA1.2- activated I TPC2 in the presence of Na + or K + bath solution. ( G ) LyNa-VA1.2- activated I TPC2 under the bi-ionic conditions of bath/cytosolic Na + and pipette/luminal Ca 2+ . 150 Na + solution contained (in mM) 145 NaCl, 5 NaOH, 20 HEPES, (pH 7.2); isotonic (105 mM) Ca 2+ solution contained (in mM) 100 CaCl 2 , 5 Ca(OH) 2 , 20 HEPES (pH 7.2). Right panel zoom-in micrograph shows the E rev of LyNa-VA1.2- activated I TPC2 . ( H–J ) Representative I-V plots of LyNa-VA1.2- evoked I TPC2-N653G ( J ) measured from Na + ( H ) and K + ( I ) bath solution. ( K ) Summary of Na + vs. K + /Ca 2+ selectivity of WT TPC2 and TPC2 N653G channels. Individual data and Mean ± S.E.M are presented (also see  ). N.S., no significance.  Figure 5—source data 1. Ionic selectivity of WT TPC2 and N653G mutant channels.

    Journal: eLife

    Article Title: Agonist-specific voltage-dependent gating of lysosomal two-pore Na + channels

    doi: 10.7554/eLife.51423

    Figure Lengend Snippet: ( A ) Representative PI(3,5)P 2 -evoked whole-endolysosome I TPC2 elicited by a voltage ramp from −120 to 120 mV. The recordings were performed under a bi-ionic condition with 150 (in mM) Na + in the pipette solution and 150 Na + or 150 K + in the bath solution. PI(3,5)P 2 (0.3 µM) was bath-applied. ( B, C ) Representative PI(3,5)P 2 - evoked I TPC2 elicited with voltage steps (−140 mV to 100 mV with a ∆V = 20 mV, 0.5 s) with 150 mM Na + ( B ) or K + ( C ) in the bath solution. ( D, E ) Representative LyNa-VA1.2-activated I TPC2 step currents. ( F ) Representative I-V plots of LyNa-VA1.2- activated I TPC2 measured from the instantaneous currents in ( D ) and ( E ). Note the reversal potentials (E rev ) of LyNa-VA1.2- activated I TPC2 in the presence of Na + or K + bath solution. ( G ) LyNa-VA1.2- activated I TPC2 under the bi-ionic conditions of bath/cytosolic Na + and pipette/luminal Ca 2+ . 150 Na + solution contained (in mM) 145 NaCl, 5 NaOH, 20 HEPES, (pH 7.2); isotonic (105 mM) Ca 2+ solution contained (in mM) 100 CaCl 2 , 5 Ca(OH) 2 , 20 HEPES (pH 7.2). Right panel zoom-in micrograph shows the E rev of LyNa-VA1.2- activated I TPC2 . ( H–J ) Representative I-V plots of LyNa-VA1.2- evoked I TPC2-N653G ( J ) measured from Na + ( H ) and K + ( I ) bath solution. ( K ) Summary of Na + vs. K + /Ca 2+ selectivity of WT TPC2 and TPC2 N653G channels. Individual data and Mean ± S.E.M are presented (also see ). N.S., no significance. Figure 5—source data 1. Ionic selectivity of WT TPC2 and N653G mutant channels.

    Article Snippet: Chemical compound, drug , PI(3,5)P 2 , Echelon Biosciences , Cat. #: P-3508 , .

    Techniques: Transferring, Mutagenesis

    KEY RESOURCES TABLE

    Journal: Cell reports

    Article Title: A Molecular Mechanism for Turning Off IRE1α Signaling during Endoplasmic Reticulum Stress

    doi: 10.1016/j.celrep.2020.108563

    Figure Lengend Snippet: KEY RESOURCES TABLE

    Article Snippet: PonceauS solution , Sigma-Aldrich , P3504.

    Techniques: Magnetic Beads, Protease Inhibitor, Recombinant, Plasmid Preparation, Software

    Journal: STAR Protocols

    Article Title: Protocol to use RNaseH1-based CRISPR to modulate locus-associated R-loops

    doi: 10.1016/j.xpro.2022.101734

    Figure Lengend Snippet:

    Article Snippet: Ponceau S , Sigma-Aldrich/Millipore , Cat# P3504-10G.

    Techniques: Purification, Recombinant, Fluorescence, Protease Inhibitor, Software, Hood, Microscopy, Real-time Polymerase Chain Reaction, Spectrophotometry

    Journal: Cell Reports

    Article Title: Sae2 and Rif2 regulate MRX endonuclease activity at DNA double-strand breaks in opposite manners

    doi: 10.1016/j.celrep.2021.108906

    Figure Lengend Snippet:

    Article Snippet: Ponceau s sodium practical grade , Sigma-Aldrich , P3504-100G.

    Techniques: Subcloning, Recombinant, Electrophoresis, Transfection, Protease Inhibitor, Purification, Plasmid Preparation, Expressing, Software