Journal of Biological Chemistry: Cohesin SA2 is a sequence-independent DNA-binding protein that recognizes DNA replication and repair intermediates
Preston Countryman 1, Yanlin Fan 2, Aparna Gorthi 3 4, Hai Pan 1, Jack Strickland 1, Parminder Kaur 1, Xuechun Wang 5, Jiangguo Lin 1 6, Xiaoying Lei 2 7, Christian White 1, Changjiang You 8, Nicolas Wirth 9, Ingrid Tessmer 9, Jacob Piehler 8, Robert Riehn 1, Alexander J R Bishop 3 4, Yizhi Jane Tao 2, Hong Wang 10 11
PMID: 29175904
PMCID: PMC5777247
DOI: 10.1074/jbc.M117.806406
Abstract
Proper chromosome alignment and segregation during mitosis depend on the cohesion between sister chromatids, mediated by the cohesin protein complex, which also plays crucial roles in diverse genome maintenance pathways. Current models attribute DNA binding by cohesin to dsDNA’s entrapment by the cohesin ring subunits (SMC1, SMC3, and RAD21 in humans). However, the biophysical properties and activities of the fourth core cohesin subunit SA2 (STAG2) are largely unknown. Here, using single-molecule atomic force and fluorescence microscopy imaging as well as fluorescence anisotropy measurements, we established that SA2 binds to both dsDNA and ssDNA, albeit with a higher binding affinity for ssDNA. We observed that SA2 could switch between the 1D diffusing (search) mode on dsDNA and stable binding (recognition) mode at ssDNA gaps. Although SA2 does not specifically bind to centromeric or telomeric sequences, it does recognize DNA structures often associated with DNA replication and double-strand break repairs, such as a double-stranded end, single-stranded overhang, flap, fork, and ssDNA gap. SA2 loss leads to a defect in homologous recombination-mediated DNA double-strand break repair. These results suggest that SA2 functions at intermediate DNA structures during DNA transactions in genome maintenance pathways. These findings have important implications for understanding the function of cohesin in these pathways.
Keywords: DNA-binding protein; SA2; STAG2; atomic force microscopy (AFM); cohesin DNA binding; fluorescence anisotropy; fluorescence microscopy; genomic instability; protein-DNA interaction; single-molecule biophysics.
© 2018 by The American Society for Biochemistry and Molecular Biology, Inc.
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