ps 473 akt Search Results


p akt  (ECM Biosciences)
93
ECM Biosciences p akt
Effects of GHS-R1α activation by ghrelin on the pathways regulating translation initiation and on the translation initiation complex in HEK293 GHS-R1α-EGFP+ cells. a , b Western blotting analysis of time- ( a ) and concentration- ( b ) dependent effects of ghrelin on the activity of <t>AKT/mTOR</t> and ERK signalling cascades, eIF2 phosphorylation and PDCD4 levels. In ( a ), 100 nM ghrelin was used; in ( b ), cells were treated for 30 min. c RPPA analysis of the effects of ghrelin treatment (500 nM, 1 h) on AMPK activity, reported by p-AMPKα (T172) and p-ACC (S79) levels ( N = 3). d Effect of ghrelin on GHS-R1α internalisation and cytosolic Ca 2+ levels: live cell confocal imaging analysis of translocation of GFP-tagged ghrelin receptor to cytosol and changes in Fura Red fluorescence upon ghrelin treatment. Note that a large proportion of GHS-R1α undergoes internalisation and aggregates in non-treated cells due to the constitutive activity of the receptor. Fura Red signals decrease upon Ca 2+ elevation; photo-bleaching of the dye is shown as mock. In all experiments ( N = 3), cells were pre-incubated in DMEM supplemented with 1% FBS for 12–14 h and then treated with 100 nM ghrelin. Fluorescence images are stacks of three (GFP) and five confocal planes (Fura Red) taken with 0.5 µm steps. e RPPA analysis of the effect of ghrelin treatment (500 nM, 1 h) on p-4E-BP1 (S65) levels; phosphorylation blocks the capacity of 4E-BP1 to inhibit initiation. f Composition of cap-bound proteins-regulators of translation initiation; changes in eIF4G and 4E-BP1 protein levels during 2 h treatment with ghrelin are shown. Input protein analysis is used as control; α-tubulin and eIF4G2 are detected only in the input samples, demonstrating specificity of cap binding. Data are shown as individual data points (gradient circles), mean values (horizontal bars) and mean ± SD. Asterisks have not been used because of the complexity of plots; see statistical significance report below. Statistical details: a effect of ghrelin treatment time: (a) <t>p-AKT</t> (4,8) = 39.625, p < 0.001, one-way ANOVA, p = 0.011 for 15 min, p = 0.024 for 30 min, p = 0.005 for 45 min, p = 0.068 for 60 min, Dunnett post hoc test vs mock treatment ( N = 3); (b) p-TSC2 (4,8) = 12.738, p = 0.002, one-way ANOVA, p = 0.046 for 15 min, p = 0.038 for 30 min, p = 0.057 for 45 min, p = 0.198 for 60 min, Dunnett post hoc test vs mock treatment ( N = 3); (c) p-mTOR (4,12) = 14.25, p < 0.001, one-way ANOVA, p < 0.001 for 15 and 30 min, p = 0.008 for 45 min, p = 0.007 for 60 min, Dunnett post hoc test vs mock treatment ( N = 4); (d) p-ERK (4,20) = 134.963, p < 0.001, one-way ANOVA, p < 0.001 for 15, 30 and 45 min, p = 0.001 for 60 min, Dunnett post hoc test vs mock treatment ( N = 6); (e) p-eIF2a (4,8) = 3.162, p = 0.078, one-way ANOVA, p = 0.614 for 15 min, p = 0.164 for 30 min, p = 0.035 for 45 min, p = 0.132 for 60 min, Dunnett post hoc test vs mock treatment ( N = 3). b Effect of ghrelin concentration: (a) p-AKT (4,8) = 4.92, p = 0.019, one-way ANOVA, p = 0.995 for 3 nM, p = 0.421 for 11 nM, p = 0.033 for 33 nM, p = 0.022 for 100 nM, Dunnett post hoc test vs mock treatment ( N = 3); (b) p-TSC2 (4,8) = 2.872, p = 0.08, one-way ANOVA, p = 0.424 for 3 nM, p = 0.26 for 11 nM, p = 0.023 for 33 nM, p = 0.246 for 100 nM, Dunnett post hoc test vs mock treatment ( N = 3); (c) p-mTOR (4,12) = 8.382, p < 0.001, one-way ANOVA, p = 0.214 for 3 nM, p = 0.025 for 11 nM, p < 0.001 for 33 and 100 nM, Dunnett post hoc test vs mock treatment ( N = 4); (d) p-ERK (4,20) = 80.687, p < 0.001, one-way ANOVA, p = 0.311 for 3 nM, p < 0.001 for 11 nM, 33 nM and 100 nM, Dunnett post hoc test vs mock treatment ( N = 6); e) p-eIF2a (4,8) = 0.523, p = 0.722, one-way ANOVA, p = 0.77, 0.989, 1 and 0.993 for 3 nM, 11 nM, 33 nM and 100 nM, accordingly, Dunnett post hoc test vs mock treatment ( N = 3). c RPPA analysis, N = 3, independent samples T test for: a) AMPK (pT172): t(4) = – 2.507, p = 0.066; b) ACC (pS79): t (4) = – 2.053, p = 0.109. d Confocal analysis of Fura Red signal (30 min after ghrelin treatment), independent samples T test, ( N = 3), T(24) = – 26.197, p < 0.001. f Effect of ghrelin treatment on the cap-binding capacity of eIF4G1 and 4E-BP1: (a) eIF4G1 (4,16) = 0.391, p = 0.812, one-way ANOVA for the effect of treatment time, p = 0.863, 0.916, 1 and 0.753 for 15, 30, 60 and 124E-BP1 eIF4G (4,16) = 5.456, p = 0.006, one-way ANOVA for the effect of treatment time, p = 0.271, 0.482, 0.939 and 0.055 for 15, 30, 60 and 120 min, respectively, Dunnett post hoc test vs mock (0 time point) treatment ( N = 5 for 15 and 30 min and N = 4 for 60 and 120 min of ghrelin treatment)
P Akt, supplied by ECM Biosciences, used in various techniques. Bioz Stars score: 93/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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Average 93 stars, based on 1 article reviews
p akt - by Bioz Stars, 2025-04
93/100 stars
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ps 473 akt pakt  (Cell Signaling Technology Inc)
86
Cell Signaling Technology Inc ps 473 akt pakt
Effects of GHS-R1α activation by ghrelin on the pathways regulating translation initiation and on the translation initiation complex in HEK293 GHS-R1α-EGFP+ cells. a , b Western blotting analysis of time- ( a ) and concentration- ( b ) dependent effects of ghrelin on the activity of <t>AKT/mTOR</t> and ERK signalling cascades, eIF2 phosphorylation and PDCD4 levels. In ( a ), 100 nM ghrelin was used; in ( b ), cells were treated for 30 min. c RPPA analysis of the effects of ghrelin treatment (500 nM, 1 h) on AMPK activity, reported by p-AMPKα (T172) and p-ACC (S79) levels ( N = 3). d Effect of ghrelin on GHS-R1α internalisation and cytosolic Ca 2+ levels: live cell confocal imaging analysis of translocation of GFP-tagged ghrelin receptor to cytosol and changes in Fura Red fluorescence upon ghrelin treatment. Note that a large proportion of GHS-R1α undergoes internalisation and aggregates in non-treated cells due to the constitutive activity of the receptor. Fura Red signals decrease upon Ca 2+ elevation; photo-bleaching of the dye is shown as mock. In all experiments ( N = 3), cells were pre-incubated in DMEM supplemented with 1% FBS for 12–14 h and then treated with 100 nM ghrelin. Fluorescence images are stacks of three (GFP) and five confocal planes (Fura Red) taken with 0.5 µm steps. e RPPA analysis of the effect of ghrelin treatment (500 nM, 1 h) on p-4E-BP1 (S65) levels; phosphorylation blocks the capacity of 4E-BP1 to inhibit initiation. f Composition of cap-bound proteins-regulators of translation initiation; changes in eIF4G and 4E-BP1 protein levels during 2 h treatment with ghrelin are shown. Input protein analysis is used as control; α-tubulin and eIF4G2 are detected only in the input samples, demonstrating specificity of cap binding. Data are shown as individual data points (gradient circles), mean values (horizontal bars) and mean ± SD. Asterisks have not been used because of the complexity of plots; see statistical significance report below. Statistical details: a effect of ghrelin treatment time: (a) <t>p-AKT</t> (4,8) = 39.625, p < 0.001, one-way ANOVA, p = 0.011 for 15 min, p = 0.024 for 30 min, p = 0.005 for 45 min, p = 0.068 for 60 min, Dunnett post hoc test vs mock treatment ( N = 3); (b) p-TSC2 (4,8) = 12.738, p = 0.002, one-way ANOVA, p = 0.046 for 15 min, p = 0.038 for 30 min, p = 0.057 for 45 min, p = 0.198 for 60 min, Dunnett post hoc test vs mock treatment ( N = 3); (c) p-mTOR (4,12) = 14.25, p < 0.001, one-way ANOVA, p < 0.001 for 15 and 30 min, p = 0.008 for 45 min, p = 0.007 for 60 min, Dunnett post hoc test vs mock treatment ( N = 4); (d) p-ERK (4,20) = 134.963, p < 0.001, one-way ANOVA, p < 0.001 for 15, 30 and 45 min, p = 0.001 for 60 min, Dunnett post hoc test vs mock treatment ( N = 6); (e) p-eIF2a (4,8) = 3.162, p = 0.078, one-way ANOVA, p = 0.614 for 15 min, p = 0.164 for 30 min, p = 0.035 for 45 min, p = 0.132 for 60 min, Dunnett post hoc test vs mock treatment ( N = 3). b Effect of ghrelin concentration: (a) p-AKT (4,8) = 4.92, p = 0.019, one-way ANOVA, p = 0.995 for 3 nM, p = 0.421 for 11 nM, p = 0.033 for 33 nM, p = 0.022 for 100 nM, Dunnett post hoc test vs mock treatment ( N = 3); (b) p-TSC2 (4,8) = 2.872, p = 0.08, one-way ANOVA, p = 0.424 for 3 nM, p = 0.26 for 11 nM, p = 0.023 for 33 nM, p = 0.246 for 100 nM, Dunnett post hoc test vs mock treatment ( N = 3); (c) p-mTOR (4,12) = 8.382, p < 0.001, one-way ANOVA, p = 0.214 for 3 nM, p = 0.025 for 11 nM, p < 0.001 for 33 and 100 nM, Dunnett post hoc test vs mock treatment ( N = 4); (d) p-ERK (4,20) = 80.687, p < 0.001, one-way ANOVA, p = 0.311 for 3 nM, p < 0.001 for 11 nM, 33 nM and 100 nM, Dunnett post hoc test vs mock treatment ( N = 6); e) p-eIF2a (4,8) = 0.523, p = 0.722, one-way ANOVA, p = 0.77, 0.989, 1 and 0.993 for 3 nM, 11 nM, 33 nM and 100 nM, accordingly, Dunnett post hoc test vs mock treatment ( N = 3). c RPPA analysis, N = 3, independent samples T test for: a) AMPK (pT172): t(4) = – 2.507, p = 0.066; b) ACC (pS79): t (4) = – 2.053, p = 0.109. d Confocal analysis of Fura Red signal (30 min after ghrelin treatment), independent samples T test, ( N = 3), T(24) = – 26.197, p < 0.001. f Effect of ghrelin treatment on the cap-binding capacity of eIF4G1 and 4E-BP1: (a) eIF4G1 (4,16) = 0.391, p = 0.812, one-way ANOVA for the effect of treatment time, p = 0.863, 0.916, 1 and 0.753 for 15, 30, 60 and 124E-BP1 eIF4G (4,16) = 5.456, p = 0.006, one-way ANOVA for the effect of treatment time, p = 0.271, 0.482, 0.939 and 0.055 for 15, 30, 60 and 120 min, respectively, Dunnett post hoc test vs mock (0 time point) treatment ( N = 5 for 15 and 30 min and N = 4 for 60 and 120 min of ghrelin treatment)
Ps 473 Akt Pakt, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/ps 473 akt pakt/product/Cell Signaling Technology Inc
Average 86 stars, based on 1 article reviews
ps 473 akt pakt - by Bioz Stars, 2025-04
86/100 stars
  Buy from Supplier
ps 473 akt pakt  (Thermo Fisher)
86
Thermo Fisher ps 473 akt pakt
Effects of GHS-R1α activation by ghrelin on the pathways regulating translation initiation and on the translation initiation complex in HEK293 GHS-R1α-EGFP+ cells. a , b Western blotting analysis of time- ( a ) and concentration- ( b ) dependent effects of ghrelin on the activity of <t>AKT/mTOR</t> and ERK signalling cascades, eIF2 phosphorylation and PDCD4 levels. In ( a ), 100 nM ghrelin was used; in ( b ), cells were treated for 30 min. c RPPA analysis of the effects of ghrelin treatment (500 nM, 1 h) on AMPK activity, reported by p-AMPKα (T172) and p-ACC (S79) levels ( N = 3). d Effect of ghrelin on GHS-R1α internalisation and cytosolic Ca 2+ levels: live cell confocal imaging analysis of translocation of GFP-tagged ghrelin receptor to cytosol and changes in Fura Red fluorescence upon ghrelin treatment. Note that a large proportion of GHS-R1α undergoes internalisation and aggregates in non-treated cells due to the constitutive activity of the receptor. Fura Red signals decrease upon Ca 2+ elevation; photo-bleaching of the dye is shown as mock. In all experiments ( N = 3), cells were pre-incubated in DMEM supplemented with 1% FBS for 12–14 h and then treated with 100 nM ghrelin. Fluorescence images are stacks of three (GFP) and five confocal planes (Fura Red) taken with 0.5 µm steps. e RPPA analysis of the effect of ghrelin treatment (500 nM, 1 h) on p-4E-BP1 (S65) levels; phosphorylation blocks the capacity of 4E-BP1 to inhibit initiation. f Composition of cap-bound proteins-regulators of translation initiation; changes in eIF4G and 4E-BP1 protein levels during 2 h treatment with ghrelin are shown. Input protein analysis is used as control; α-tubulin and eIF4G2 are detected only in the input samples, demonstrating specificity of cap binding. Data are shown as individual data points (gradient circles), mean values (horizontal bars) and mean ± SD. Asterisks have not been used because of the complexity of plots; see statistical significance report below. Statistical details: a effect of ghrelin treatment time: (a) <t>p-AKT</t> (4,8) = 39.625, p < 0.001, one-way ANOVA, p = 0.011 for 15 min, p = 0.024 for 30 min, p = 0.005 for 45 min, p = 0.068 for 60 min, Dunnett post hoc test vs mock treatment ( N = 3); (b) p-TSC2 (4,8) = 12.738, p = 0.002, one-way ANOVA, p = 0.046 for 15 min, p = 0.038 for 30 min, p = 0.057 for 45 min, p = 0.198 for 60 min, Dunnett post hoc test vs mock treatment ( N = 3); (c) p-mTOR (4,12) = 14.25, p < 0.001, one-way ANOVA, p < 0.001 for 15 and 30 min, p = 0.008 for 45 min, p = 0.007 for 60 min, Dunnett post hoc test vs mock treatment ( N = 4); (d) p-ERK (4,20) = 134.963, p < 0.001, one-way ANOVA, p < 0.001 for 15, 30 and 45 min, p = 0.001 for 60 min, Dunnett post hoc test vs mock treatment ( N = 6); (e) p-eIF2a (4,8) = 3.162, p = 0.078, one-way ANOVA, p = 0.614 for 15 min, p = 0.164 for 30 min, p = 0.035 for 45 min, p = 0.132 for 60 min, Dunnett post hoc test vs mock treatment ( N = 3). b Effect of ghrelin concentration: (a) p-AKT (4,8) = 4.92, p = 0.019, one-way ANOVA, p = 0.995 for 3 nM, p = 0.421 for 11 nM, p = 0.033 for 33 nM, p = 0.022 for 100 nM, Dunnett post hoc test vs mock treatment ( N = 3); (b) p-TSC2 (4,8) = 2.872, p = 0.08, one-way ANOVA, p = 0.424 for 3 nM, p = 0.26 for 11 nM, p = 0.023 for 33 nM, p = 0.246 for 100 nM, Dunnett post hoc test vs mock treatment ( N = 3); (c) p-mTOR (4,12) = 8.382, p < 0.001, one-way ANOVA, p = 0.214 for 3 nM, p = 0.025 for 11 nM, p < 0.001 for 33 and 100 nM, Dunnett post hoc test vs mock treatment ( N = 4); (d) p-ERK (4,20) = 80.687, p < 0.001, one-way ANOVA, p = 0.311 for 3 nM, p < 0.001 for 11 nM, 33 nM and 100 nM, Dunnett post hoc test vs mock treatment ( N = 6); e) p-eIF2a (4,8) = 0.523, p = 0.722, one-way ANOVA, p = 0.77, 0.989, 1 and 0.993 for 3 nM, 11 nM, 33 nM and 100 nM, accordingly, Dunnett post hoc test vs mock treatment ( N = 3). c RPPA analysis, N = 3, independent samples T test for: a) AMPK (pT172): t(4) = – 2.507, p = 0.066; b) ACC (pS79): t (4) = – 2.053, p = 0.109. d Confocal analysis of Fura Red signal (30 min after ghrelin treatment), independent samples T test, ( N = 3), T(24) = – 26.197, p < 0.001. f Effect of ghrelin treatment on the cap-binding capacity of eIF4G1 and 4E-BP1: (a) eIF4G1 (4,16) = 0.391, p = 0.812, one-way ANOVA for the effect of treatment time, p = 0.863, 0.916, 1 and 0.753 for 15, 30, 60 and 124E-BP1 eIF4G (4,16) = 5.456, p = 0.006, one-way ANOVA for the effect of treatment time, p = 0.271, 0.482, 0.939 and 0.055 for 15, 30, 60 and 120 min, respectively, Dunnett post hoc test vs mock (0 time point) treatment ( N = 5 for 15 and 30 min and N = 4 for 60 and 120 min of ghrelin treatment)
Ps 473 Akt Pakt, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/ps 473 akt pakt/product/Thermo Fisher
Average 86 stars, based on 1 article reviews
ps 473 akt pakt - by Bioz Stars, 2025-04
86/100 stars
  Buy from Supplier
ps 473 akt  (Cell Signaling Technology Inc)
86
Cell Signaling Technology Inc ps 473 akt
Effects of GHS-R1α activation by ghrelin on the pathways regulating translation initiation and on the translation initiation complex in HEK293 GHS-R1α-EGFP+ cells. a , b Western blotting analysis of time- ( a ) and concentration- ( b ) dependent effects of ghrelin on the activity of <t>AKT/mTOR</t> and ERK signalling cascades, eIF2 phosphorylation and PDCD4 levels. In ( a ), 100 nM ghrelin was used; in ( b ), cells were treated for 30 min. c RPPA analysis of the effects of ghrelin treatment (500 nM, 1 h) on AMPK activity, reported by p-AMPKα (T172) and p-ACC (S79) levels ( N = 3). d Effect of ghrelin on GHS-R1α internalisation and cytosolic Ca 2+ levels: live cell confocal imaging analysis of translocation of GFP-tagged ghrelin receptor to cytosol and changes in Fura Red fluorescence upon ghrelin treatment. Note that a large proportion of GHS-R1α undergoes internalisation and aggregates in non-treated cells due to the constitutive activity of the receptor. Fura Red signals decrease upon Ca 2+ elevation; photo-bleaching of the dye is shown as mock. In all experiments ( N = 3), cells were pre-incubated in DMEM supplemented with 1% FBS for 12–14 h and then treated with 100 nM ghrelin. Fluorescence images are stacks of three (GFP) and five confocal planes (Fura Red) taken with 0.5 µm steps. e RPPA analysis of the effect of ghrelin treatment (500 nM, 1 h) on p-4E-BP1 (S65) levels; phosphorylation blocks the capacity of 4E-BP1 to inhibit initiation. f Composition of cap-bound proteins-regulators of translation initiation; changes in eIF4G and 4E-BP1 protein levels during 2 h treatment with ghrelin are shown. Input protein analysis is used as control; α-tubulin and eIF4G2 are detected only in the input samples, demonstrating specificity of cap binding. Data are shown as individual data points (gradient circles), mean values (horizontal bars) and mean ± SD. Asterisks have not been used because of the complexity of plots; see statistical significance report below. Statistical details: a effect of ghrelin treatment time: (a) <t>p-AKT</t> (4,8) = 39.625, p < 0.001, one-way ANOVA, p = 0.011 for 15 min, p = 0.024 for 30 min, p = 0.005 for 45 min, p = 0.068 for 60 min, Dunnett post hoc test vs mock treatment ( N = 3); (b) p-TSC2 (4,8) = 12.738, p = 0.002, one-way ANOVA, p = 0.046 for 15 min, p = 0.038 for 30 min, p = 0.057 for 45 min, p = 0.198 for 60 min, Dunnett post hoc test vs mock treatment ( N = 3); (c) p-mTOR (4,12) = 14.25, p < 0.001, one-way ANOVA, p < 0.001 for 15 and 30 min, p = 0.008 for 45 min, p = 0.007 for 60 min, Dunnett post hoc test vs mock treatment ( N = 4); (d) p-ERK (4,20) = 134.963, p < 0.001, one-way ANOVA, p < 0.001 for 15, 30 and 45 min, p = 0.001 for 60 min, Dunnett post hoc test vs mock treatment ( N = 6); (e) p-eIF2a (4,8) = 3.162, p = 0.078, one-way ANOVA, p = 0.614 for 15 min, p = 0.164 for 30 min, p = 0.035 for 45 min, p = 0.132 for 60 min, Dunnett post hoc test vs mock treatment ( N = 3). b Effect of ghrelin concentration: (a) p-AKT (4,8) = 4.92, p = 0.019, one-way ANOVA, p = 0.995 for 3 nM, p = 0.421 for 11 nM, p = 0.033 for 33 nM, p = 0.022 for 100 nM, Dunnett post hoc test vs mock treatment ( N = 3); (b) p-TSC2 (4,8) = 2.872, p = 0.08, one-way ANOVA, p = 0.424 for 3 nM, p = 0.26 for 11 nM, p = 0.023 for 33 nM, p = 0.246 for 100 nM, Dunnett post hoc test vs mock treatment ( N = 3); (c) p-mTOR (4,12) = 8.382, p < 0.001, one-way ANOVA, p = 0.214 for 3 nM, p = 0.025 for 11 nM, p < 0.001 for 33 and 100 nM, Dunnett post hoc test vs mock treatment ( N = 4); (d) p-ERK (4,20) = 80.687, p < 0.001, one-way ANOVA, p = 0.311 for 3 nM, p < 0.001 for 11 nM, 33 nM and 100 nM, Dunnett post hoc test vs mock treatment ( N = 6); e) p-eIF2a (4,8) = 0.523, p = 0.722, one-way ANOVA, p = 0.77, 0.989, 1 and 0.993 for 3 nM, 11 nM, 33 nM and 100 nM, accordingly, Dunnett post hoc test vs mock treatment ( N = 3). c RPPA analysis, N = 3, independent samples T test for: a) AMPK (pT172): t(4) = – 2.507, p = 0.066; b) ACC (pS79): t (4) = – 2.053, p = 0.109. d Confocal analysis of Fura Red signal (30 min after ghrelin treatment), independent samples T test, ( N = 3), T(24) = – 26.197, p < 0.001. f Effect of ghrelin treatment on the cap-binding capacity of eIF4G1 and 4E-BP1: (a) eIF4G1 (4,16) = 0.391, p = 0.812, one-way ANOVA for the effect of treatment time, p = 0.863, 0.916, 1 and 0.753 for 15, 30, 60 and 124E-BP1 eIF4G (4,16) = 5.456, p = 0.006, one-way ANOVA for the effect of treatment time, p = 0.271, 0.482, 0.939 and 0.055 for 15, 30, 60 and 120 min, respectively, Dunnett post hoc test vs mock (0 time point) treatment ( N = 5 for 15 and 30 min and N = 4 for 60 and 120 min of ghrelin treatment)
Ps 473 Akt, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
https://www.bioz.com/result/ps 473 akt/product/Cell Signaling Technology Inc
Average 86 stars, based on 1 article reviews
ps 473 akt - by Bioz Stars, 2025-04
86/100 stars
  Buy from Supplier
antiphosphoserine ps 473 akt  (Upstate Biotechnology Inc)
86
Upstate Biotechnology Inc antiphosphoserine ps 473 akt
Effects of GHS-R1α activation by ghrelin on the pathways regulating translation initiation and on the translation initiation complex in HEK293 GHS-R1α-EGFP+ cells. a , b Western blotting analysis of time- ( a ) and concentration- ( b ) dependent effects of ghrelin on the activity of <t>AKT/mTOR</t> and ERK signalling cascades, eIF2 phosphorylation and PDCD4 levels. In ( a ), 100 nM ghrelin was used; in ( b ), cells were treated for 30 min. c RPPA analysis of the effects of ghrelin treatment (500 nM, 1 h) on AMPK activity, reported by p-AMPKα (T172) and p-ACC (S79) levels ( N = 3). d Effect of ghrelin on GHS-R1α internalisation and cytosolic Ca 2+ levels: live cell confocal imaging analysis of translocation of GFP-tagged ghrelin receptor to cytosol and changes in Fura Red fluorescence upon ghrelin treatment. Note that a large proportion of GHS-R1α undergoes internalisation and aggregates in non-treated cells due to the constitutive activity of the receptor. Fura Red signals decrease upon Ca 2+ elevation; photo-bleaching of the dye is shown as mock. In all experiments ( N = 3), cells were pre-incubated in DMEM supplemented with 1% FBS for 12–14 h and then treated with 100 nM ghrelin. Fluorescence images are stacks of three (GFP) and five confocal planes (Fura Red) taken with 0.5 µm steps. e RPPA analysis of the effect of ghrelin treatment (500 nM, 1 h) on p-4E-BP1 (S65) levels; phosphorylation blocks the capacity of 4E-BP1 to inhibit initiation. f Composition of cap-bound proteins-regulators of translation initiation; changes in eIF4G and 4E-BP1 protein levels during 2 h treatment with ghrelin are shown. Input protein analysis is used as control; α-tubulin and eIF4G2 are detected only in the input samples, demonstrating specificity of cap binding. Data are shown as individual data points (gradient circles), mean values (horizontal bars) and mean ± SD. Asterisks have not been used because of the complexity of plots; see statistical significance report below. Statistical details: a effect of ghrelin treatment time: (a) <t>p-AKT</t> (4,8) = 39.625, p < 0.001, one-way ANOVA, p = 0.011 for 15 min, p = 0.024 for 30 min, p = 0.005 for 45 min, p = 0.068 for 60 min, Dunnett post hoc test vs mock treatment ( N = 3); (b) p-TSC2 (4,8) = 12.738, p = 0.002, one-way ANOVA, p = 0.046 for 15 min, p = 0.038 for 30 min, p = 0.057 for 45 min, p = 0.198 for 60 min, Dunnett post hoc test vs mock treatment ( N = 3); (c) p-mTOR (4,12) = 14.25, p < 0.001, one-way ANOVA, p < 0.001 for 15 and 30 min, p = 0.008 for 45 min, p = 0.007 for 60 min, Dunnett post hoc test vs mock treatment ( N = 4); (d) p-ERK (4,20) = 134.963, p < 0.001, one-way ANOVA, p < 0.001 for 15, 30 and 45 min, p = 0.001 for 60 min, Dunnett post hoc test vs mock treatment ( N = 6); (e) p-eIF2a (4,8) = 3.162, p = 0.078, one-way ANOVA, p = 0.614 for 15 min, p = 0.164 for 30 min, p = 0.035 for 45 min, p = 0.132 for 60 min, Dunnett post hoc test vs mock treatment ( N = 3). b Effect of ghrelin concentration: (a) p-AKT (4,8) = 4.92, p = 0.019, one-way ANOVA, p = 0.995 for 3 nM, p = 0.421 for 11 nM, p = 0.033 for 33 nM, p = 0.022 for 100 nM, Dunnett post hoc test vs mock treatment ( N = 3); (b) p-TSC2 (4,8) = 2.872, p = 0.08, one-way ANOVA, p = 0.424 for 3 nM, p = 0.26 for 11 nM, p = 0.023 for 33 nM, p = 0.246 for 100 nM, Dunnett post hoc test vs mock treatment ( N = 3); (c) p-mTOR (4,12) = 8.382, p < 0.001, one-way ANOVA, p = 0.214 for 3 nM, p = 0.025 for 11 nM, p < 0.001 for 33 and 100 nM, Dunnett post hoc test vs mock treatment ( N = 4); (d) p-ERK (4,20) = 80.687, p < 0.001, one-way ANOVA, p = 0.311 for 3 nM, p < 0.001 for 11 nM, 33 nM and 100 nM, Dunnett post hoc test vs mock treatment ( N = 6); e) p-eIF2a (4,8) = 0.523, p = 0.722, one-way ANOVA, p = 0.77, 0.989, 1 and 0.993 for 3 nM, 11 nM, 33 nM and 100 nM, accordingly, Dunnett post hoc test vs mock treatment ( N = 3). c RPPA analysis, N = 3, independent samples T test for: a) AMPK (pT172): t(4) = – 2.507, p = 0.066; b) ACC (pS79): t (4) = – 2.053, p = 0.109. d Confocal analysis of Fura Red signal (30 min after ghrelin treatment), independent samples T test, ( N = 3), T(24) = – 26.197, p < 0.001. f Effect of ghrelin treatment on the cap-binding capacity of eIF4G1 and 4E-BP1: (a) eIF4G1 (4,16) = 0.391, p = 0.812, one-way ANOVA for the effect of treatment time, p = 0.863, 0.916, 1 and 0.753 for 15, 30, 60 and 124E-BP1 eIF4G (4,16) = 5.456, p = 0.006, one-way ANOVA for the effect of treatment time, p = 0.271, 0.482, 0.939 and 0.055 for 15, 30, 60 and 120 min, respectively, Dunnett post hoc test vs mock (0 time point) treatment ( N = 5 for 15 and 30 min and N = 4 for 60 and 120 min of ghrelin treatment)
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Effects of GHS-R1α activation by ghrelin on the pathways regulating translation initiation and on the translation initiation complex in HEK293 GHS-R1α-EGFP+ cells. a , b Western blotting analysis of time- ( a ) and concentration- ( b ) dependent effects of ghrelin on the activity of AKT/mTOR and ERK signalling cascades, eIF2 phosphorylation and PDCD4 levels. In ( a ), 100 nM ghrelin was used; in ( b ), cells were treated for 30 min. c RPPA analysis of the effects of ghrelin treatment (500 nM, 1 h) on AMPK activity, reported by p-AMPKα (T172) and p-ACC (S79) levels ( N = 3). d Effect of ghrelin on GHS-R1α internalisation and cytosolic Ca 2+ levels: live cell confocal imaging analysis of translocation of GFP-tagged ghrelin receptor to cytosol and changes in Fura Red fluorescence upon ghrelin treatment. Note that a large proportion of GHS-R1α undergoes internalisation and aggregates in non-treated cells due to the constitutive activity of the receptor. Fura Red signals decrease upon Ca 2+ elevation; photo-bleaching of the dye is shown as mock. In all experiments ( N = 3), cells were pre-incubated in DMEM supplemented with 1% FBS for 12–14 h and then treated with 100 nM ghrelin. Fluorescence images are stacks of three (GFP) and five confocal planes (Fura Red) taken with 0.5 µm steps. e RPPA analysis of the effect of ghrelin treatment (500 nM, 1 h) on p-4E-BP1 (S65) levels; phosphorylation blocks the capacity of 4E-BP1 to inhibit initiation. f Composition of cap-bound proteins-regulators of translation initiation; changes in eIF4G and 4E-BP1 protein levels during 2 h treatment with ghrelin are shown. Input protein analysis is used as control; α-tubulin and eIF4G2 are detected only in the input samples, demonstrating specificity of cap binding. Data are shown as individual data points (gradient circles), mean values (horizontal bars) and mean ± SD. Asterisks have not been used because of the complexity of plots; see statistical significance report below. Statistical details: a effect of ghrelin treatment time: (a) p-AKT (4,8) = 39.625, p < 0.001, one-way ANOVA, p = 0.011 for 15 min, p = 0.024 for 30 min, p = 0.005 for 45 min, p = 0.068 for 60 min, Dunnett post hoc test vs mock treatment ( N = 3); (b) p-TSC2 (4,8) = 12.738, p = 0.002, one-way ANOVA, p = 0.046 for 15 min, p = 0.038 for 30 min, p = 0.057 for 45 min, p = 0.198 for 60 min, Dunnett post hoc test vs mock treatment ( N = 3); (c) p-mTOR (4,12) = 14.25, p < 0.001, one-way ANOVA, p < 0.001 for 15 and 30 min, p = 0.008 for 45 min, p = 0.007 for 60 min, Dunnett post hoc test vs mock treatment ( N = 4); (d) p-ERK (4,20) = 134.963, p < 0.001, one-way ANOVA, p < 0.001 for 15, 30 and 45 min, p = 0.001 for 60 min, Dunnett post hoc test vs mock treatment ( N = 6); (e) p-eIF2a (4,8) = 3.162, p = 0.078, one-way ANOVA, p = 0.614 for 15 min, p = 0.164 for 30 min, p = 0.035 for 45 min, p = 0.132 for 60 min, Dunnett post hoc test vs mock treatment ( N = 3). b Effect of ghrelin concentration: (a) p-AKT (4,8) = 4.92, p = 0.019, one-way ANOVA, p = 0.995 for 3 nM, p = 0.421 for 11 nM, p = 0.033 for 33 nM, p = 0.022 for 100 nM, Dunnett post hoc test vs mock treatment ( N = 3); (b) p-TSC2 (4,8) = 2.872, p = 0.08, one-way ANOVA, p = 0.424 for 3 nM, p = 0.26 for 11 nM, p = 0.023 for 33 nM, p = 0.246 for 100 nM, Dunnett post hoc test vs mock treatment ( N = 3); (c) p-mTOR (4,12) = 8.382, p < 0.001, one-way ANOVA, p = 0.214 for 3 nM, p = 0.025 for 11 nM, p < 0.001 for 33 and 100 nM, Dunnett post hoc test vs mock treatment ( N = 4); (d) p-ERK (4,20) = 80.687, p < 0.001, one-way ANOVA, p = 0.311 for 3 nM, p < 0.001 for 11 nM, 33 nM and 100 nM, Dunnett post hoc test vs mock treatment ( N = 6); e) p-eIF2a (4,8) = 0.523, p = 0.722, one-way ANOVA, p = 0.77, 0.989, 1 and 0.993 for 3 nM, 11 nM, 33 nM and 100 nM, accordingly, Dunnett post hoc test vs mock treatment ( N = 3). c RPPA analysis, N = 3, independent samples T test for: a) AMPK (pT172): t(4) = – 2.507, p = 0.066; b) ACC (pS79): t (4) = – 2.053, p = 0.109. d Confocal analysis of Fura Red signal (30 min after ghrelin treatment), independent samples T test, ( N = 3), T(24) = – 26.197, p < 0.001. f Effect of ghrelin treatment on the cap-binding capacity of eIF4G1 and 4E-BP1: (a) eIF4G1 (4,16) = 0.391, p = 0.812, one-way ANOVA for the effect of treatment time, p = 0.863, 0.916, 1 and 0.753 for 15, 30, 60 and 124E-BP1 eIF4G (4,16) = 5.456, p = 0.006, one-way ANOVA for the effect of treatment time, p = 0.271, 0.482, 0.939 and 0.055 for 15, 30, 60 and 120 min, respectively, Dunnett post hoc test vs mock (0 time point) treatment ( N = 5 for 15 and 30 min and N = 4 for 60 and 120 min of ghrelin treatment)

Journal: Cellular and Molecular Life Sciences

Article Title: Ghrelin rapidly elevates protein synthesis in vitro by employing the rpS6K-eEF2K-eEF2 signalling axis

doi: 10.1007/s00018-022-04446-4

Figure Lengend Snippet: Effects of GHS-R1α activation by ghrelin on the pathways regulating translation initiation and on the translation initiation complex in HEK293 GHS-R1α-EGFP+ cells. a , b Western blotting analysis of time- ( a ) and concentration- ( b ) dependent effects of ghrelin on the activity of AKT/mTOR and ERK signalling cascades, eIF2 phosphorylation and PDCD4 levels. In ( a ), 100 nM ghrelin was used; in ( b ), cells were treated for 30 min. c RPPA analysis of the effects of ghrelin treatment (500 nM, 1 h) on AMPK activity, reported by p-AMPKα (T172) and p-ACC (S79) levels ( N = 3). d Effect of ghrelin on GHS-R1α internalisation and cytosolic Ca 2+ levels: live cell confocal imaging analysis of translocation of GFP-tagged ghrelin receptor to cytosol and changes in Fura Red fluorescence upon ghrelin treatment. Note that a large proportion of GHS-R1α undergoes internalisation and aggregates in non-treated cells due to the constitutive activity of the receptor. Fura Red signals decrease upon Ca 2+ elevation; photo-bleaching of the dye is shown as mock. In all experiments ( N = 3), cells were pre-incubated in DMEM supplemented with 1% FBS for 12–14 h and then treated with 100 nM ghrelin. Fluorescence images are stacks of three (GFP) and five confocal planes (Fura Red) taken with 0.5 µm steps. e RPPA analysis of the effect of ghrelin treatment (500 nM, 1 h) on p-4E-BP1 (S65) levels; phosphorylation blocks the capacity of 4E-BP1 to inhibit initiation. f Composition of cap-bound proteins-regulators of translation initiation; changes in eIF4G and 4E-BP1 protein levels during 2 h treatment with ghrelin are shown. Input protein analysis is used as control; α-tubulin and eIF4G2 are detected only in the input samples, demonstrating specificity of cap binding. Data are shown as individual data points (gradient circles), mean values (horizontal bars) and mean ± SD. Asterisks have not been used because of the complexity of plots; see statistical significance report below. Statistical details: a effect of ghrelin treatment time: (a) p-AKT (4,8) = 39.625, p < 0.001, one-way ANOVA, p = 0.011 for 15 min, p = 0.024 for 30 min, p = 0.005 for 45 min, p = 0.068 for 60 min, Dunnett post hoc test vs mock treatment ( N = 3); (b) p-TSC2 (4,8) = 12.738, p = 0.002, one-way ANOVA, p = 0.046 for 15 min, p = 0.038 for 30 min, p = 0.057 for 45 min, p = 0.198 for 60 min, Dunnett post hoc test vs mock treatment ( N = 3); (c) p-mTOR (4,12) = 14.25, p < 0.001, one-way ANOVA, p < 0.001 for 15 and 30 min, p = 0.008 for 45 min, p = 0.007 for 60 min, Dunnett post hoc test vs mock treatment ( N = 4); (d) p-ERK (4,20) = 134.963, p < 0.001, one-way ANOVA, p < 0.001 for 15, 30 and 45 min, p = 0.001 for 60 min, Dunnett post hoc test vs mock treatment ( N = 6); (e) p-eIF2a (4,8) = 3.162, p = 0.078, one-way ANOVA, p = 0.614 for 15 min, p = 0.164 for 30 min, p = 0.035 for 45 min, p = 0.132 for 60 min, Dunnett post hoc test vs mock treatment ( N = 3). b Effect of ghrelin concentration: (a) p-AKT (4,8) = 4.92, p = 0.019, one-way ANOVA, p = 0.995 for 3 nM, p = 0.421 for 11 nM, p = 0.033 for 33 nM, p = 0.022 for 100 nM, Dunnett post hoc test vs mock treatment ( N = 3); (b) p-TSC2 (4,8) = 2.872, p = 0.08, one-way ANOVA, p = 0.424 for 3 nM, p = 0.26 for 11 nM, p = 0.023 for 33 nM, p = 0.246 for 100 nM, Dunnett post hoc test vs mock treatment ( N = 3); (c) p-mTOR (4,12) = 8.382, p < 0.001, one-way ANOVA, p = 0.214 for 3 nM, p = 0.025 for 11 nM, p < 0.001 for 33 and 100 nM, Dunnett post hoc test vs mock treatment ( N = 4); (d) p-ERK (4,20) = 80.687, p < 0.001, one-way ANOVA, p = 0.311 for 3 nM, p < 0.001 for 11 nM, 33 nM and 100 nM, Dunnett post hoc test vs mock treatment ( N = 6); e) p-eIF2a (4,8) = 0.523, p = 0.722, one-way ANOVA, p = 0.77, 0.989, 1 and 0.993 for 3 nM, 11 nM, 33 nM and 100 nM, accordingly, Dunnett post hoc test vs mock treatment ( N = 3). c RPPA analysis, N = 3, independent samples T test for: a) AMPK (pT172): t(4) = – 2.507, p = 0.066; b) ACC (pS79): t (4) = – 2.053, p = 0.109. d Confocal analysis of Fura Red signal (30 min after ghrelin treatment), independent samples T test, ( N = 3), T(24) = – 26.197, p < 0.001. f Effect of ghrelin treatment on the cap-binding capacity of eIF4G1 and 4E-BP1: (a) eIF4G1 (4,16) = 0.391, p = 0.812, one-way ANOVA for the effect of treatment time, p = 0.863, 0.916, 1 and 0.753 for 15, 30, 60 and 124E-BP1 eIF4G (4,16) = 5.456, p = 0.006, one-way ANOVA for the effect of treatment time, p = 0.271, 0.482, 0.939 and 0.055 for 15, 30, 60 and 120 min, respectively, Dunnett post hoc test vs mock (0 time point) treatment ( N = 5 for 15 and 30 min and N = 4 for 60 and 120 min of ghrelin treatment)

Article Snippet: Antibodies were from: Alomone Labs, Israel (ghrelin receptor, № AGR-031); Becton Dickinson, NJ (eIF4G2 aka NAT1, № 610,742); Cell Signalling Technology, MA (CREB, № 1385; phospho-CREB (Ser133), № 9198; mTOR, № 2972; phospho-mTOR (Ser2448), № 2971; phospho-ERK1/2 (Thr202/Tyr204), № 9101; eEF2, № 2332; phospho-eEF2 (Thr56), № 2331; eEF2K, № 3692; phospho-eEF2K (Ser366), № 3691; eIF4G1, № 2498; 4E-BP1, № 9452; eIF2α, № 5324; phospho-eIF2α (Set51), № 3398; p-AKT (Ser473), № 4060; tuberin/TSC2, № 4304; phospho-TSC2 (Thr1462), № 3617; rpS6, №2217; phospho-rpS6 (Ser235/236), № 2271, PDCD4, № 9535); ECM Biosciences, KY (eEF2K Phospho-Regulation Antibody Sampler Kit, № EK6910, which includes antibodies against Ser78, Ser359, Thr348, Ser398, Ser500 in eEF2K and eEF2K C-terminus); Millipore, CA (ERK1/2, № 06–182); Proteintech, IL (AKT, № 10,176–2-AP); Sigma, MO (α-tubulin №T5168, HRP-conjugated anti-rabbit and anti-mouse antibodies, № A1949 and A0168); Thermo Fisher Scientific, MA (anti-rabbit Alexa Fluor 555-conjugated, № A-21428).

Techniques: Activation Assay, Western Blot, Concentration Assay, Activity Assay, Imaging, Translocation Assay, Fluorescence, Incubation, Binding Assay