About GSBS   |  FAQ  |  Job Opportunities  |  Search UMDNJ
     




Cohesion of Silenced Chromatin by Sirtuins in Saccharomyces cerevisiae

by
Yu-Fan Chen
M.S., National Yang Ming University - 2008

Thesis Advisor: Marc Gartenberg, Ph.D.
Graduate Program in Cellular & Molecular Pharmacology

RWJMS Research Building/School of Public Health
Conference Room 258
Piscataway

Tuesday, April 28, 2015
9:00 a.m.


Abstract

Assembly of yeast heterochromatin requires Sir2, the founding member of the conserved family of NAD+-dependent protein deacetylases known as the sirtuins. These proteins serve as metabolic sensors involved in cellular lifespan. They have also been linked to cancer and numerous human diseases. Previous work in the Gartenberg lab showed an unanticipated role for yeast Sir2: the protein is required for cohesion of yeast heterochromatin. Tethered Sir2 fragments mediate cohesion in the absence of other Sir proteins whereas synthetic heterochromatin domains that lack Sir2 do not mediate cohesion at all. The domain of Sir2 responsible for cohesin recruitment maps to the C-terminus. Using mutagenesis guided by Sir2 crystal structures, I identified a small surface patch within this domain that is required for cohesin recruitment and cohesion. At least one additional domain of Sir2 contributes to cohesin recruitment but the secondary domain is not sufficient for cohesion.

The Sir2 surface mutations that block cohesion do not disrupt heterochromatic repression in most standard phenotypic assays. In more specialized assays, however, defects in silencing can be detected. Elevated expression of Sir4, a Sir2 binding partner, suppresses the silencing defects without restoring heterochromatic cohesion. These studies show that the silencing and cohesion functions of Sir2 can be uncoupled, and that silencing can be achieved without cohesion of heterochromatin.

To investigate how Sir2 mediates cohesin recruitment, I used two approaches to find proteins that interact with the C-terminus: yeast two-hybrid screening and copurification followed by mass spectroscopy. Candidate factors from these approaches are discussed.


Return to Dissertation list

 

Newark Campus - Piscataway Campus - Stratford Campus
About GSBS - FAQ - Job Opportunities - Search UMDNJ