Abstract
BACKGROUND: MutS Homolog 3 (MSH3), part of the MutSβ DNA mismatch repair complex with MSH2, can reversibly translocate from the nucleus to cytosol via IL-6 signaling, abrogating nuclear MutSβ function and is associated with metastasis and poor patient survival. A polymorphism consisting of deletion of 27-bp proximate to the nuclear localization signal (NLS) (Δ27bpMSH3) allows MSH3 cytosolic retention with IL-6 or oxidative stress. Here, we examined for IL-6-induced post-translational modifications associated with MSH3 cytosolic translocation. METHODS: We utilized MSH3-genotyped colon cancer cell lines after IL-6 treatment to assess post-translational modification of MSH3 via Western blots (WB). We modified sequences within the MSH3-NLS-EGFP reporter construct to assess MSH3 localization via immunofluorescent microscopy and WB after nuclear-cytosolic fractionation. Immunoprecipitation (IP) followed by WB was used to study post-IL-6-induced interactions with MSH3. RESULTS: MSH3 and Δ27bpMSH3 increased serine phosphorylation after 2 hours followed by tyrosine phosphorylation 18 hours post IL-6 treatment, with Δ27bpMSH3 showing more robust phosphorylation than MSH3 likely due to increased cytosolic translocation. MSH3 cytosolic localization was enhanced by acetylation of lysine residues within MSH3's NLS, specifically at residues K(99), K(100) and K(103). With the observed acetylation control for MSH3 cytosolic localization, IP experiments demonstrate binding of cytosolic-located histone deacetylase 6 (HDAC6) to acetylated Δ27bpMSH3. CONCLUSIONS: Polymorphic MSH3 undergoes serine/tyrosine phosphorylation and NLS acetylation upon IL-6 signaling for its nuclear-cytosolic shift and binds HDAC6 in the cytosol which may contribute to anticipated deacetylation and MSH3 protein stability when separated from MSH2. These modifications might be targeted to regulate MSH3's intracellular localization.