s6 ribosomal protein 5g10 2217 cell signaling  (Cell Signaling Technology Inc)

Journal: JCI Insight

Article Title: Loss of genome maintenance is linked to mTOR complex 1 signaling and accelerates podocyte damage

doi: 10.1172/jci.insight.172370

Figure Lengend Snippet: ( A ) Volcano plot depicting differentially expressed genes of Ercc1 pko versus control glomeruli. Significantly differentially expressed genes depicted in black, pseudogenes depicted in red. FDR, false discovery rate. ( B ) Gene length analysis of genes differentially expressed in Ercc1 pko versus control glomeruli. Downregulated genes (blue) are significantly larger than upregulated genes (red). P value: 1.52 × 10 –7 (Mann-Whitney U test). ( C ) Correlation analysis plotting the NES of Kyoto Encyclopedia of Genes and Genomes pathway genes in Tsc2 ko versus ctrl kidneys and Ercc1 pko versus ctrl glomeruli. ( D ) GSEA of differentially expressed genes (upregulation, fold-change ≥ 2) in Ercc1 -pko glomeruli classified by the gene rank in the mTORC1 hyperactivation ( Tsc2 ko) data set. The running enrichment score is shown as a green line. The upregulated genes in Ercc1 -pko glomeruli are significantly (adjusted P value: 1.67 × 10 –5 ) more enriched (NES: 1.74) in the genes upregulated upon mTORC1 hyperactivation. ( E ) Representative immunofluorescence staining of SNP, p-S6RP, and DAPI in sections of 9-week-old Ercc1 ctrl and pko kidneys with quantification of SNP and p-S6RP double-positive cells per glomerulus and per total SNP-positive cells, scale bar indicating 20 μm & 10 μm in zoom (unpaired t test, n = 5, 10 glomeruli per sample). **** P ≤ 0.0001.

Article Snippet: SDS-PAGE was used for protein size separation with subsequent blotting onto PVDF membranes and visualized with enhanced chemiluminescence after incubation of the blots with corresponding antibodies: Phospho-Histone H2A.X (Ser139); Phospho-S6 Ribosomal Protein (Ser235/236) (D57.2.2E); S6 Ribosomal Protein (5G10) from Cell Signaling Technology; α-actin (JLA20) from Developmental Studies Hybridoma Bank; and β-tubulin (E7) from Developmental Studies Hybridoma Bank.

Techniques: Control, MANN-WHITNEY, Immunofluorescence, Staining

Journal: JCI Insight

Article Title: Loss of genome maintenance is linked to mTOR complex 1 signaling and accelerates podocyte damage

doi: 10.1172/jci.insight.172370

Figure Lengend Snippet: ( A ) Schematic in vitro protocol for the induction of DNA damage. ( B ) Representative immunoblot images of gH2A.X and loading control β-tubulin of immortalized murine podocyte lysates ( n = 3). ( C ) Representative immunofluorescence images for p53 and DAPI in immortalized murine podocytes, scale bar indicating 20 μm ( n = 3). ( D ) Representative immunoblot images of p-S6RP, S6RP, and loading control protein α-actin of immortalized murine podocyte lysates; p-S6RP and S6RP gels run in parallel ( n = 3). All cells imaged or lysed after treatment with MMC or UV-C irradiation ± 10 ng/mL rapamycin or serum starvation (SS) ( n ≥ 4). ( E ) Representative immunoblot images of p-S6RP, S6RP, and loading control protein α-actin of immortalized murine podocyte lysates ( n = 3 MMC; n = 6 UV-C). All cells imaged or lysed after treatment with MMC or UV-C irradiation ± ATM inhibitor KU60019 or DNA-PK inhibitor nedisertib. ( F ) Representative immunofluorescence staining of synaptopodin (gray), p–DNA PKc (green), and DAPI (red) in sections of 9-week-old Ercc1 ctrl and pko kidneys, with quantification of SNP-positive cells with p–DNA PKc–positive nuclei per total SNP-positive cells. Yellow circles indicating positive nuclei, red circles indicating negative nuclei, scale bar indicating 50 μm (unpaired t test, n = 4–5, 10 glomeruli per sample). ( G ) Representative immunofluorescence staining of SNP (gray), p-ATM (green), and DAPI (red) in sections of 9-week-old Ercc1 ctrl and pko kidneys, with quantification of SNP-positive cells with p–ATM-positive nuclei per total SNP-positive cells. Yellow circles indicating positive nuclei, red circles indicating negative nuclei, scale bar indicating 10 μm (unpaired t test, n = 4–5, 10 glomeruli per sample). All violin plots indicate median (black) and upper and lower quartile (gray). *** P ≤ 0.001, **** P ≤ 0.0001. DNA-PK, DNA-dependent protein kinase; ATM, ataxia telangiectasia mutated serine/threonine kinase.

Article Snippet: SDS-PAGE was used for protein size separation with subsequent blotting onto PVDF membranes and visualized with enhanced chemiluminescence after incubation of the blots with corresponding antibodies: Phospho-Histone H2A.X (Ser139); Phospho-S6 Ribosomal Protein (Ser235/236) (D57.2.2E); S6 Ribosomal Protein (5G10) from Cell Signaling Technology; α-actin (JLA20) from Developmental Studies Hybridoma Bank; and β-tubulin (E7) from Developmental Studies Hybridoma Bank.

Techniques: In Vitro, Western Blot, Control, Immunofluorescence, Irradiation, Staining

Journal: JCI Insight

Article Title: Loss of genome maintenance is linked to mTOR complex 1 signaling and accelerates podocyte damage

doi: 10.1172/jci.insight.172370

Figure Lengend Snippet: ( A ) Representative PAS staining of Ercc1 -pko mice treated with vehicle ( Ercc1 pko) or 2 mg rapamycin/kg body weight ( Ercc1 pko Rapa) from 6 weeks of age and glomerulosclerosis assessment of all glomeruli depicted as parts of a whole and scatterplot, scale bar: 50 μm (2-way ANOVA with Šídák’s multiple comparisons test, n ≥ 9, 50 glomeruli per sample). ( B ) Representative PAS staining of end-of-life Ercc1 –/Δ mice treated with 14 mg rapamycin/kg food from 8 weeks of age and glomerulosclerosis assessment of all glomeruli depicted as parts of a whole and scatterplot, scale bar: 50 μm (2-way ANOVA with Šídák’s multiple comparisons test, n = 8, 50 glomeruli per sample). ( C ) Representative immunofluorescence staining of SNP, p-S6RP, and DAPI in sections of end-of-life Ercc1 –/Δ kidneys treated (according to B ) with quantification of SNP and p-S6RP double-positive cells per glomerulus and SNP-positive area per glomerulus, scale bar: 20 μm, 5 μm in zoom (unpaired t test, n = 5, 10 glomeruli per sample). ( D ) STED images of cleared kidney tissue of Ercc1 –/Δ kidneys treated (according to B ) after immunolabeling with an anti-nephrin antibody with quantification of slit diaphragm length, scale bar: 2 μm (unpaired t test, 5 areas per animal, n = 4 animals). ( E ) Representative immunofluorescence staining of SNP (gray), gH2A.X (green), and Draq5 (red) in sections of end-of-life Ercc1 –/Δ kidneys treated (according to B ) with quantification of gH2A.X foci per nuclear area. Yellow dotted line, nuclear border ( n = 5, 10 glomeruli per sample, 5 podocytes per glomerulus); scale bar: 2 μm. All violin plots indicate median (black) and upper and lower quartile (gray); scatterplots indicate mean plus 95% confidence interval. * P ≤ 0.05, ** P ≤ 0.01, *** P ≤ 0.001, **** P ≤ 0.0001.

Article Snippet: SDS-PAGE was used for protein size separation with subsequent blotting onto PVDF membranes and visualized with enhanced chemiluminescence after incubation of the blots with corresponding antibodies: Phospho-Histone H2A.X (Ser139); Phospho-S6 Ribosomal Protein (Ser235/236) (D57.2.2E); S6 Ribosomal Protein (5G10) from Cell Signaling Technology; α-actin (JLA20) from Developmental Studies Hybridoma Bank; and β-tubulin (E7) from Developmental Studies Hybridoma Bank.

Techniques: Staining, Immunofluorescence, Immunolabeling

Journal: Communications Biology

Article Title: Lipolysis-derived fatty acids are needed for homeostatic control of sterol element-binding protein-1c driven hepatic lipogenesis

doi: 10.1038/s42003-025-08002-1

Figure Lengend Snippet: Resources table

Article Snippet: Antibody , Anti-S6 ribosomal protein, clone 5G10 (Rabbit polyclonal) , Cell Signaling , AB_331355 , 1:1000.

Techniques: Knock-Out