Off-campus WSU users: To download campus access dissertations, please use the following link to log into our proxy server with your WSU access ID and password, then click the "Off-campus Download" button below.
Non-WSU users: Please talk to your librarian about requesting this dissertation through interlibrary loan.
Access Type
WSU Access
Date of Award
1-1-2003
Degree Type
Dissertation
Degree Name
Ph.D.
Department
Biological Sciences
First Advisor
John Lopes
Abstract
Gene expression is coordinately controlled by a network of regulatory proteins and mis-regulation of these proteins can affect cell metabolism globally. For example, mis-regulation of mammalian C-MYC proto-oncogene expression was identified in all cases of Burkitt's lymphoma and in several cases of breast cancers and prostrate cancers. Myc is a basic helix-loop-helix (bHLH) protein that activates the expression of numerous genes. Therefore, it is important that we understand how bHLH proteins function and how their expression is regulated. bHLH proteins are highly conserved from yeast to humans. The yeast phospholipid biosynthetic pathway provides a unique opportunity to study the regulation of a bHLH gene and its targets. Two bHLH proteins, Ino2p and Ino4p, regulate phospholipid biosynthetic gene expression. Ino2p heterodimerizes with Ino4p, binds to the bHLH consensus site (5'-CANNTG-3') located in the promoter of target genes, and activates their transcription by recruiting chromatin remodeling complexes. The expression of INO2 gene itself is auto-regulated and is also regulated by two regulatory proteins, Ume6p and Opi1p. The goal of this dissertation is to understand how Ume6p and Opi1p regulate the expression of INO2 bHLH gene and the phospholipid biosynthetic structural genes. In a ume6Delta strain, the expression of most phospholipid biosynthetic genes and the INO2 regulatory gene is drastically reduced. Mutants that over-express the INO2 gene and rescue a ume6Delta phenotype were isolated using INO2-HIS3 reporter. The mutant collection contained mutations in OPI1 and SIN3 genes. Ume6p, Sin3p, and Rpd3p are known to form a complex and repress numerous genes. Consistent with this, I found that a sin3Delta mutant over-expressed an INO2-cat reporter, but surprisingly an rpd3Delta mutant had no effect. Further, an OPI1-cat expression study demonstrated that UME6 does not regulate OPI1 gene expression. Therefore mutations in OPI1 and SIN3 genes bypass the effect of ume6Delta mutation on INO2 gene expression. Characterization of opi1 mutants revealed that a mutant Opi1p is synthesized; however, the opi1 mutants misregulate INO1-lacZ (INO1 is an Ino2p target gene) expression. Therefore, the work presented here sheds light on the complex regulation of the INO2 bHLH gene and its target genes.
Recommended Citation
Kaadige, Mohan R., "Characterization of two repression mechanisms in Saccharomyces cerevisiae" (2003). Wayne State University Dissertations. 3404.
https://digitalcommons.wayne.edu/oa_dissertations/3404