GENOME-WIDE DEPENDENCY OF TRANSCRIPTION MACHINERY ON GCN5 IN SACCHAROMYCES CEREVISIAE

Open Access
Author:
Cantor, David John
Area of Honors:
Biochemistry and Molecular Biology
Degree:
Bachelor of Science
Document Type:
Thesis
Thesis Supervisors:
  • Benjamin Franklin Pugh, Thesis Supervisor
  • David Scott Gilmour, Honors Advisor
  • Wendy Hanna Rose, Faculty Reader
Keywords:
  • transcription machinery genome-wide mapping
  • Gcn5 deletion
  • ChIP-exo
  • SOLiD sequencing
Abstract:
Eukaryotic gene regulation has largely been evolutionary conserved. The basic transcription machinery present in Saccharomyces cerevisiae, budding yeast, have close homologs in higher eukaryotic organisms. The Spt-Ada-Gcn5 (SAGA) complex is the major H3 histone acetyltransferase (HAT). While certain factor binding locations are known on a genomic scale, there exists little data on the functional dependencies of the transcriptional machinery during PIC assembly genome-wide. In the present study, genome-wide occupancies of 12 gene regulatory proteins in Gcn5 deletion strains were mapped using chromatin immunoprecipitation followed by next generation SOLiD sequencing (ChIP-seq) to better elucidate their dependencies on the HAT activity of SAGA. Gcn5 was deleted from strains with 12 gene regulatory proteins TAP-tagged. Stress state occupancy was compared to non-stress state occupancy by putting the cells through a heat shock. Evidence suggests that SAGA is involved in the stress response, and so a better elucidation of functional dependency on Gcn5 may be assessed upon heat shock stress. The following study focuses on analyzing the initial sequencing data from the deletion strains. We also show different genome-wide occupancy patterns for Rsc9, a subunit of RSC (Remodel the Structure of Chromatin) complex, and RNA polymerase II when at stress state compared to non-stress state in the Gcn5 deletion models suggesting a unique response to stress separate from the SAGA dependent pathway.