Identifying Novel Interaction Between the E3 Ubiquitin Ligase Ring Finger Protein 20 (RNF20) and the Deubiquitinating Enzyme Ubiquitin Specific Protease 7 (USP7)

Abstract

USP7, which is a ubiquitin-specific protease, is an essential deubiquitinating enzyme involved in a plethora of cellular processes such as epigenetic regulation, DNA damage response and oncogenic processes. In this study, RNF20, which is an E3 ligase, has been identified as a novel substrate for USP7. Through various biochemical and functional assays, including endogenous co-immunoprecipitation, comparing parental and USP7 knockout HCT116 cells, reconstitution assay, overexpression with USP7 or the catalytic mutant USP7, cycloheximide chase and a deubiquitination assay, we show that RNF20 stability is dependent upon USP7 removing ubiquitin chains that would mark RNF20 for proteasomal degradation. This interaction was seen within multiple cell lines, with the catalytic activity of USP7 being crucial to the stability of RNF20. These findings provide the foundation for future studies on how the regulation of RNF20 through USP7 can contribute to epigenetic regulation, DNA damage response, and oncogenic processes.