期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：2022
卷号：119
期号：33
DOI：10.1073/pnas.2208004119
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：Significance
Cohesin is a heteropentameric protein complex consisting of two structural maintenance of chromosomes (SMC) subunits and three non-SMC subunits. The two SMC subunits form a heterodimer with an ATPase head and hinge that are connected by long coiled coils. Isolation of ATPase mutants followed by comprehensive identification of suppressor mutations in SMC subunits that can bypass ATPase defects was performed. Locations and properties of mutant alleles reflect how ATPase activities could be compromised by structural adaptation. ATP-driven conformational changes may enhance DNA anchoring by the head, alter interactions of coiled coils at the head with other subunits for DNA to go through, and fold/extend coiled coils near break sites around midpoint to bring together DNA elements far from each other.
The cohesin complex is required for sister chromatid cohesion and genome compaction. Cohesin coiled coils (CCs) can fold at break sites near midpoints to bring head and hinge domains, located at opposite ends of coiled coils, into proximity. Whether ATPase activities in the head play a role in this conformational change is yet to be known. Here, we dissected functions of cohesin ATPase activities in cohesin dynamics in
Schizosaccharomyces pombe. Isolation and characterization of cohesin ATPase temperature-sensitive (ts) mutants indicate that both ATPase domains are required for proper chromosome segregation. Unbiased screening of spontaneous suppressor mutations rescuing the temperature lethality of cohesin ATPase mutants identified several suppressor hotspots in cohesin that located outside of ATPase domains. Then, we performed comprehensive saturation mutagenesis targeted to these suppressor hotspots. Large numbers of the identified suppressor mutations indicated several different ways to compensate for the ATPase mutants: 1) Substitutions to amino acids with smaller side chains in coiled coils at break sites around midpoints may enable folding and extension of coiled coils more easily; 2) substitutions to arginine in the DNA binding region of the head may enhance DNA binding; or 3) substitutions to hydrophobic amino acids in coiled coils, connecting the head and interacting with other subunits, may alter conformation of coiled coils close to the head. These results reflect serial structural changes in cohesin driven by its ATPase activities potentially for packaging DNAs.
