Accurate chromosome segregation is essential for the successful transmission of genetic information to daughter cells, and deficiencies in this process are associated with miscarriages, congenital disorders, and tumorigenesis. Integral to proper chromosome segregation is the cohesion or physical association of replicated sister chromatids, which is mediated by the evolutionarily conserved cohesin complex. Cohesins are involved not only in chromosome segregation, but also play important roles in DNA repair and gene regulation. Cohesin's spatial distribution on budding yeast chromosomes is highly reproducible, suggesting that it plays important roles in chromosome structure and function. Cohesins are frequently found in intergenic regions between convergently transcribed genes, indicating interplay between cohesin distributions and transcription that is currently not characterized. Notably, extensive cohesin-enriched domains assemble in pericentromeric (kinetochore-flanking) regions, where they promote chromosome biorientation and resist precocious sister chromatid separation. We showed previously that budding yeast kinetochores direct pericentromeric cohesin domain assembly. Our preliminary data indicate that pericentromeric cohesin domains are assembled epigenetically by a nucleation and spreading mechanism. We test the veracity of this model in Specific Aim 1 by determining whether loop formation adjacent to the centromere impedes cohesin recruitment in distal sequences. Pericentromeric chromatin will also be isolated and its protein composition characterized in an unbiased screen for kinetochore-associated factors or epigenetic chromatin modifications that direct cohesin domain assembly. Lastly, insulators that limit cohesin recruitment will be used to delineate the minimally effective pericentromeric domain necessary for high fidelity chromosome transmission. Preliminary data also indicate that cohesin distributions are directed by the prior association of the Scc2/Scc4 cohesin loader with intergenic sequences. In the second aim, we endeavor to determine the mechanisms and significance of loader and cohesin localization on chromosome arms. The order and interdependence of association of the RSC ATP-dependent chromatin remodeler, Scc2/Scc4, and cohesin will be determined at chromosome arm cohesin-associated regions to dissect the pathway for cohesin deposition. The contribution of RNA polymerase II-dependent transcription in the establishment of cohesin loader distributions will be examined following polymerase inactivation and the mechanism and significance of cohesin redistribution in an RNA polymerase II transcription termination mutant are examined to delineate the nature of the relationship between transcription and cohesin localization. The role of budding yeast cohesins in the promotion of RNA polymerase II transcription termination will be determined using conditional cohesin mutants.

Public Health Relevance

Chromosomes must be accurately duplicated and segregated to daughter cells during each cell division, and errors in these events can lead to cancer or genetic disease, such as Down syndrome. The accurate segregation of replicated chromosomes, or sister chromatids, to daughter cells requires that sisters physically associate with one another throughout much of the cell cycle. In this application, we endeavor to understand how the proteins that mediate the physical association of sister chromatids, called cohesins, are recruited to proper locations on the chromosome to mediate chromosome segregation, DNA repair, and gene regulation, each of which is important for the maintenance of genomic integrity.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM066213-09
Application #
8368800
Study Section
Nuclear and Cytoplasmic Structure/Function and Dynamics Study Section (NCSD)
Program Officer
Janes, Daniel E
Project Start
2002-07-01
Project End
2014-11-30
Budget Start
2012-12-01
Budget End
2014-11-30
Support Year
9
Fiscal Year
2013
Total Cost
$287,027
Indirect Cost
$93,569
Name
University of Colorado Denver
Department
Biochemistry
Type
Schools of Medicine
DUNS #
041096314
City
Aurora
State
CO
Country
United States
Zip Code
80045
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Kogut, Igor; Wang, Jianbin; Guacci, Vincent et al. (2009) The Scc2/Scc4 cohesin loader determines the distribution of cohesin on budding yeast chromosomes. Genes Dev 23:2345-57
Eckert, Carrie A; Gravdahl, Daniel J; Megee, Paul C (2007) The enhancement of pericentromeric cohesin association by conserved kinetochore components promotes high-fidelity chromosome segregation and is sensitive to microtubule-based tension. Genes Dev 21:278-91
Weber, Stewart A; Gerton, Jennifer L; Polancic, Joan E et al. (2004) The kinetochore is an enhancer of pericentric cohesin binding. PLoS Biol 2:E260
Glynn, Earl F; Megee, Paul C; Yu, Hong-Guo et al. (2004) Genome-wide mapping of the cohesin complex in the yeast Saccharomyces cerevisiae. PLoS Biol 2:E259