1. Cellular transformation up-regulates proteins that help preserve telomere integrity.
Cellular transformation up-regulates proteins that help preserve telomere integrity. siRNA screens were performed to test the effect of candidate depletion on telomere structure visualized on metaphase chromosome spreads. (A) Heatmap showing behavior of the selected candidates in QTIPs. Reported are mean SILAC ratios from the forward and reverse TRF IP replicates. The asterisks indicate proteins that were classified as significantly “transformation responsive” in the QTIP screen (Table S1, list C). (B) RT–qPCR to verify candidate depletion using siRNA pools in HLF-TSR and HeLa cells. Knockdown efficiency was compared with the mRNA levels in cells transfected with control siRNA against GFP. Plotted is mean + SD from two biological and two technical replicates for HLF-TSR and from two technical replicates from one representative experiment for HeLa. Two-tailed unpaired t test (*P < 0.05; **P < 0.01; ns = not significant). (C) Analysis of fragile telomeres upon candidate depletion. Indicated telomere aberrations were scored on metaphase spreads with telomeres detected by FISH with a Cy3-[CCCTAA]3 probe (red) and DNA stained with DAPI (gray). HLF-TSR: >50 metaphases from two independent experiments (except for siGFP samples, for which data from three independent replicates) were analyzed for each condition. HeLa: >25 metaphases were analyzed per condition in a single experiment. The black line represents median. One-way ANOVA with Dunnett’s multiple comparisons test, comparing all conditions to siGFP (*P < 0.05; **P < 0.01; ****P < ; ns = not significant). See also Table S2.
Jana Majerska et al. LSA 2018;1:e
© 2018 Lingner et al.