Figure 1. ICL Sensing by MutSα (MSH2–MSH6) (A) Plasmid pull-down assay of control plasmids or plasmids containing a single site-specific SJG-136 ICL lesion. Plasmids were incubated for 40 min in HSS and then purified from extracts using recombinant Bio-LacR protein coupled to streptavidin beads. Plasmid-bound proteins were analyzed by western blot (WB) using the indicated antibodies. (B) Plasmid pull-down assay and WB of SJG-136-treated plasmids. In the last lane, labeled (10×2), twice the amount of 10 μM-treated plasmids was incubated in HSS. Input was plasmid DNA run on an agarose gel. (C) MSH2–MSH6 immunodepletion. Mock- and MSH2-depleted HSS was analyzed by WB. (D) Quantification of ICL repair in mock- and MSH2-depleted HSS at 3 hr. Results represent mean ± SEM from n = 7 independent experiments. (E) Quantification of ICL repair in HSS and HSS supplemented with MutSαWT at 3 hr. Results represent mean ± SEM from n = 11 independent experiments.
Image published in: Kato N et al. (2017)
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