USP13 interacts with cohesin and regulates its ubiquitination in human cells
| Autor: | Jung-Sik Kim, Bogdan Budnik, Xiaoyuan He, Todd Waldman, Cecil Han, William S. Lane, Laura A. Díaz-Martínez |
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| Jazyk: | angličtina |
| Rok vydání: | 2020 |
| Předmět: |
0301 basic medicine
cell division DNA Replication DNA Repair DNA repair Immunoprecipitation Chromosomal Proteins Non-Histone ac-SMC3 acetylated-SMC3 cohesin Cell Cycle Proteins Biochemistry FSBP FLAG-streptavidin-binding peptide ZnF zinc finger domain genome structure 03 medical and health sciences Chromosome Segregation Humans Protein Interaction Domains and Motifs Molecular Biology Mitosis mitosis 030102 biochemistry & molecular biology Cohesin Chemistry Ubiquitin DNA replication Ubiquitination SBP streptavidin binding peptide USP13 USP13 Ubiquitin-Specific Protease 13 Cell Biology Cell cycle HCT116 Cells Chromatin Cell biology Establishment of sister chromatid cohesion TBS Tris-buffered saline 030104 developmental biology deubiquitylation (deubiquitination) protein–protein interaction cell cycle Ubiquitin-Specific Proteases biological phenomena cell phenomena and immunity Research Article HeLa Cells |
| Zdroj: | The Journal of Biological Chemistry |
| ISSN: | 1083-351X 0021-9258 |
| Popis: | Cohesin is a multiprotein ring complex that regulates 3D genome organization, sister chromatid cohesion, gene expression, and DNA repair. Cohesin is known to be ubiquitinated, although the mechanism, regulation, and effects of cohesin ubiquitination remain poorly defined. We previously used gene editing to introduce a dual epitope tag into the endogenous allele of each of 11 known components of cohesin in human HCT116 cells. Here we report that mass spectrometry analysis of dual-affinity purifications identified the USP13 deubiquitinase as a novel cohesin-interacting protein. Subsequent immunoprecipitation/Western blots confirmed the endogenous interaction in HCT116, 293T, HeLa, and RPE-hTERT cells; demonstrated that the interaction occurs specifically in the soluble nuclear fraction (not in the chromatin); requires the ubiquitin-binding domains (UBA1/2) of USP13; and occurs preferentially during DNA replication. Reciprocal dual-affinity purification of endogenous USP13 followed by mass spectrometry demonstrated that cohesin is its primary interactor in the nucleus. Ectopic expression and CRISPR knockout of USP13 showed that USP13 is paradoxically required for both deubiquitination and ubiquitination of cohesin subunits in human cells. USP13 was dispensable for sister chromatid cohesion in HCT116 and HeLa cells, whereas it was required for the dissociation of cohesin from chromatin as cells transit through mitosis. Together these results identify USP13 as a new cohesin-interacting protein that regulates the ubiquitination of cohesin and its cell cycle regulated interaction with chromatin. |
| Databáze: | OpenAIRE |
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