Fig. 6: Morc3-ATPase cycle and SUMOylation are needed for its interaction with Daxx.
From: Morc3 silences endogenous retroviruses by enabling Daxx-mediated histone H3.3 incorporation
a ChIP-MS identification of Morc3-associated proteins. Morc3 ChIP-MS experiments (n = 3) were performed with Morc3-3xFLAG knock-in ES cells and wild-type ES cells using FLAG antibody. Dot plot shows log fold change enrichment vs. −log(p value) of proteins in Morc3-3xFLAG vs. wild type (background IP) cells. Positions of labeled proteins are indicated by red dots. b Panther protein class enrichment analysis of proteins which are significantly enriched in the Morc3 ChIP-MS. Bar graph shows the percentage of total mouse proteins (black) or Morc3 ChIP-MS proteins (blue) in significantly enriched Panther protein classes (Fisher’s exact test, p value < 0.05). c ChIP-MS analysis of mutant Morc3 proteins. Dot plots show relative iBAQ values of proteins quantified in wild type vs. mutant Morc3 ChIP-MS data. Positions of ERV regulators are indicated. d Morc3 interaction with Daxx requires the ATPase cycle and SUMOylation. 3xFLAG tagged Morc3 wild type and mutant proteins were immunoprecipitated from nuclear extract of rescue cell lines using anti-FLAG magnetic beads. Wild-type cells without FLAG epitope served as negative control. Western blot analysis of Morc3, Daxx, and Nanog (negative control) revealed interaction of Morc3 wild type protein with Daxx. Mutant Morc3 proteins fail to co-immunoprecipitate Daxx. Uncropped blots in Source data. e ChIP-MS analysis of Daxx wild type vs. DaxxΔSIM mutant proteins. Dot plot shows relative iBAQ values of proteins quantified in Daxx rescue cell lines expressing wild-type Daxx or DaxxΔSIM mutant protein. Positions of ERV regulators are indicated by red dots. f Daxx requires the C-terminal SIM domain for Morc3 interaction. 3xFLAG tagged Daxx wild type and DaxxΔSIM proteins were immunoprecipitated from nuclear extract of rescue cell lines using anti-FLAG magnetic beads. Wild-type cells without FLAG epitope served as negative control. Western blot analysis of Daxx, Morc3, and Nanog (negative control) revealed interaction of Daxx wild type protein with Morc3. Mutant DaxxΔSIM failed to co-immunoprecipitate Morc3. Uncropped blots in Source data.
