Fig. 6: TbRAP1 RRM mutants have an increased amount of DNA damage at the telomere and the subtelomere.
From: The RRM-mediated RNA binding activity in T. brucei RAP1 is essential for VSG monoallelic expression
a TbRAP1-/2FL exhibits an increased VSG switching rate. Average and standard deviation were calculated from three (WT) and four (TbRAP1-/2FL) independent experiments. P values of two-sided unpaired t-tests are shown (TbRAP1-/2FL vs TbRAP1+/+). b, d, e Western analyses to examine the γH2A protein level in WT cells before and after phleomycin treatment (as a positive control) and in TbRAP1F/2FL (b), TbRAP1F/2FQ (d), and TbRAP1F/2FA&5A (e) cells before and after a 30–48 h Cre induction. A γH2A rabbit antibody23 and the tubulin antibody TAT-149 were used. Molecular marker was run on the left lane in each gel and their sizes are indicated on the left. c, f ChIP using the γH2A rabbit antibody and IgG in TbRAP1F/2FL (c) and TbRAP1F/2FQ (f) cells after a 30 h Cre induction followed by Southern blotting using a telomere and a tubulin probe. Blots were exposed to a phosphorimager. Images were quantified using ImageQuant and average and standard deviation were calculated from two (γH2A antibody, (TTAGGG)n probe in TbRAP1F/2FL cells) or three (all other samples) independent experiments in (c) and three independent experiments in (f). P values of two-sided unpaired t-tests (mutant vs. control cells) are shown. g ChIP using a γH2A rabbit antibody and IgG in TbRAP1F/2FQ cells followed by quantitative PCR using primers specific to the indicated active and silent ES loci. SNAP50 is a chromosome internal gene. Average enrichment (ChIP/Input) was calculated from three independent experiments. P values of two-sided unpaired t-tests (γH2A ChIP products, +Cre vs. -Cre) are shown. Error bars in (a, c, f, g) represent standard deviation. Source data are provided as a Source Data file.
