Regulation of Enhancers by SUMOylation Through TFAP2C Binding and Recruitment of HDAC Complex to the Chromatin.

Enhancers are fundamental to gene regulation. Post-translational modifications by the small ubiquitin-like modifiers (SUMO) modify chromatin regulation enzymes, including histone acetylases and deacetylases. However, it remains unclear whether SUMOylation regulates enhancer marks, acetylation at the 27th lysine residue of the histone H3 protein (H3K27Ac). To investigate whether SUMOylation regulates H3K27Ac, we performed genome-wide ChIP-seq analyses and discovered that knockdown (KD) of the SUMO activating enzyme catalytic subunit UBA2 reduced H3K27Ac at most enhancers. Bioinformatic analysis revealed that TFAP2C-binding sites are enriched in enhancers whose H3K27Ac was reduced by UBA2 KD. ChIP-seq analysis in combination with molecular biological methods showed that TFAP2C binding to enhancers increased upon UBA2 KD or inhibition of SUMOylation by a small molecule SUMOylation inhibitor. However, this is not due to the SUMOylation of TFAP2C itself. Proteomics analysis of TFAP2C interactome on the chromatin identified histone deacetylation (HDAC) and RNA splicing machineries that contain many SUMOylation targets. TFAP2C KD reduced HDAC1 binding to chromatin and increased H3K27Ac marks at enhancer regions, suggesting that TFAP2C is important in recruiting HDAC machinery. Taken together, our findings provide insights into the regulation of enhancer marks by SUMOylation and TFAP2C and suggest that SUMOylation of proteins in the HDAC machinery regulates their recruitments to enhancers.

Phosphorylation of ETS1 by Src family kinases prevents its recognition by the COP1 tumor suppressor.

Oncoproteins and tumor suppressors antagonistically converge on critical nodes governing neoplastic growth, invasion, and metastasis. We discovered that phosphorylation of the ETS1 and ETS2 transcriptional oncoproteins at specific serine or threonine residues creates binding sites for the COP1 tumor suppressor protein, which is an ubiquitin ligase component, leading to their destruction. In the case of ETS1, however, phosphorylation of a neighboring tyrosine residue by Src family kinases disrupts COP1 binding, thereby stabilizing ETS1. Src-dependent accumulation of ETS1 in breast cancer cells promotes anchorage-independent growth in vitro and tumor growth in vivo. These findings expand the list of potential COP1 substrates to include proteins whose COP1-binding sites are subject to regulatory phosphorylation and provide insights into transformation by Src family kinases.