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J Biol Chem. 2019 May 31;294(22):8760-8772. doi: 10.1074/jbc.RA119.007832. Epub 2019 Apr 22.

Systematic proteomics of endogenous human cohesin reveals an interaction with diverse splicing factors and RNA-binding proteins required for mitotic progression.

Author information

1
From the Departments of Oncology and Biochemistry & Molecular Biology, Georgetown University School of Medicine, Washington, D. C. 20057.
2
the Department of Genome Sciences.
3
Institute for Stem Cell and Regenerative Medicine, and.
4
Division of Hematology, University of Washington, Seattle, Washington 98195.
5
the Section of Hematology, Yale Cancer Center, Yale University School of Medicine, New Haven, Connecticut 06510, and.
6
the Mass Spectrometry and Proteomics Resource Laboratory, Harvard University, Cambridge, Massachusetts 02138.
7
From the Departments of Oncology and Biochemistry & Molecular Biology, Georgetown University School of Medicine, Washington, D. C. 20057, waldmant@georgetown.edu.

Abstract

The cohesin complex regulates sister chromatid cohesion, chromosome organization, gene expression, and DNA repair. Cohesin is a ring complex composed of four core subunits and seven regulatory subunits. In an effort to comprehensively identify additional cohesin-interacting proteins, we used gene editing to introduce a dual epitope tag into the endogenous allele of each of 11 known components of cohesin in cultured human cells, and we performed MS analyses on dual-affinity purifications. In addition to reciprocally identifying all known components of cohesin, we found that cohesin interacts with a panoply of splicing factors and RNA-binding proteins (RBPs). These included diverse components of the U4/U6.U5 tri-small nuclear ribonucleoprotein complex and several splicing factors that are commonly mutated in cancer. The interaction between cohesin and splicing factors/RBPs was RNA- and DNA-independent, occurred in chromatin, was enhanced during mitosis, and required RAD21. Furthermore, cohesin-interacting splicing factors and RBPs followed the cohesin cycle and prophase pathway of cell cycle-regulated interactions with chromatin. Depletion of cohesin-interacting splicing factors and RBPs resulted in aberrant mitotic progression. These results provide a comprehensive view of the endogenous human cohesin interactome and identify splicing factors and RBPs as functionally significant cohesin-interacting proteins.

KEYWORDS:

CKAP5; MGA; RNA splicing; RNA-binding protein; STAG2; cell cycle; cell division; cohesin; gene editing; genome structure; mitosis; protein–protein interaction

PMID:
31010829
PMCID:
PMC6552432
[Available on 2020-05-31]
DOI:
10.1074/jbc.RA119.007832

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