Modulation of the DNA-binding activity of Saccharomyces cerevisiae MSH2–MSH6 complex by the high-mobility group protein NHP6A, in vitro

DNA mismatch repair corrects mispaired bases and small insertions/deletions in DNA. In eukaryotes, the mismatch repair complex MSH2-MSH6 binds to mispairs with only slightly higher affinity than to fully paired DNA in vitro. Recently, the high-mobility group box1 protein, (HMGB1), has been shown to stimulate the mismatch repair reaction in vitro. In yeast, the closest homologs of HMGB1 are NHP6A and NHP6B. These proteins have been shown to be required for genome stability maintenance and mutagenesis control. In this work, we show that MSH2-MSH6 and NHP6A modulate their binding to DNA in vitro. Binding of the yeast MSH2-MSH6 to homoduplex regions of DNA significantly stimulates the loading of NHP6A. Upon binding of NHP6A to DNA, MSH2-MSH6 is excluded from binding unless a mismatch is present. A DNA binding-impaired MSH2-MSH6F337A significantly reduced the loading of NHP6A to DNA, suggesting that MSH2-MSH6 binding is a requisite for NHP6A loading. MSH2-MSH6 and NHP6A form a stable complex, which is responsive to ATP on mismatched substrates. These results suggest that MSH2-MSH6 binding to homoduplex regions of DNA recruits NHP6A, which then prevents further binding of MSH2-MSH6 to these sites unless a mismatch is present.