Date of Award

Fall 12-2008

Degree Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Biological Sciences

Committee Chair

Glenmore Shearer Jr. & Dr. Jeong-Ho Kim

Committee Chair Department

Biological Sciences

Committee Member 2

Mohamed Elasri

Committee Member 2 Department

Biological Sciences

Committee Member 3

Yanlin Guo

Committee Member 3 Department

Biological Sciences

Committee Member 4

Youping Deng

Committee Member 4 Department

Biological Sciences

Abstract

Glucose is the most abundant monosaccharide and preferred carbon and energy source for most cells. Many organisms have evolved sophisticated means to sense glucose and respond to it appropriately. The budding yeast, Saccharomyces cerevisiae senses glucose through two transmembrane proteins, Snf3 and Rgt2. In the presence of extracellular glucose Snf3 and Rgt2 generate an intracellular signal that leads to the degradation of Mthl and Stdl, thereby inducing the expression of hexose transporter genes (EXT) by inhibiting the function of Rgtl, a transcriptional repressor of HXT genes. Mthl and Stdl are degraded via the Yckl/2 Kinase-SCFGrrl-26S proteasome pathway triggered by the glucose sensors. RGT2-1 and SNF3-1 induce expression of HXT genes even in the absence of glucose. I show that RGT2-1 promotes ubiquitination and subsequent degradation of Mthl and Stdl regardless of the presence of glucose. Sitespecific mutagenesis reveals that conserved lysine residues of Mthl and Stdl might serve as attachment sites for ubiquitin, and that the potential casein kinase (Yckl/2) consensus sites in Mthl and Stdl are needed for their phosphorylation. The data provides biochemical evidence for glucose independent degradation of Mthl and Stdl. I further identified, the subcellular localization and the cellular compartment in which of Mthl and Stdl are degraded in response to glucose. The data shows that, Mthl and Stdl are present in nucleus when they are not degraded due to mutational blocks in the SnO/Rgt2-Rgtl pathway. Mthl and Stdl could be degraded in both the nucleus and cytoplasm when its subcellular localization is artificially manipulated; however, glucose-induced degradation occurs only in the nucleus. I also demonstrate that membrane tethering of Yckl/2 plays no or little role in the degradation of Mthl. Transcriptomic analysis of mthlAstdlA mutant identified new target genes for Mthl and Stdl in new functional categories including mitochondrial/respiration genes, transporter genes and amino acid pathway genes in addition to HXT genes. This analysis provided insights into understanding the new functions of the two paralogous proteins Mthl and Stdl.

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