This proposal examines the molecular basis of differential gene expression in the unicellular eucaryotic organism, the yeast Saccharomyces cerevisiae. Yeast cells exhibit one of three specialized cell types -- a, alpha, and a alpha -- each of which produces a unique set of proteins that create the distinctive cell types. Expression of the structural genes that encode these cell- type-specific proteins is regulated by alleles of a single genetic locus, the mating-type locus (MATa and MATalpha). Our goal is to understand the mechanism by which the regulatory protein alphal, encoded by MATalpha, activates expression of alpha- specific genes. We have focused on a particular alpha-specific gene, STE3, whose transcription is subject to two inputs in addition to the requirement for the activator alphal. In particular, the products of five others genes (STE4, STE5, STE7, STE11, and STE12) are required for efficient transcription and part of the response to the a-factor peptide pheromone secreted by a cells is increased transcription of STE3. We have delimited a segment of STE3 (called its UAS) that is necessary and sufficient to impart alpha-specific expression. We have shown that alphal protein, in conjunction with an as yet uncharacterized protein, binds to the STE3 UAS element. We will use gel mobility shift assays and DNAase footprinting assays to study the formation of this protein-DNA complex. Specifically we will determine which nucleotides within the UAS are essential, and whether the STE4/12 genes or response to a-factor affect the formation of the complex. We will determine whether alphal makes specific contacts with the UAS DNA and also identify the other protein(s) that is part of the complex. Our views is that this other protein is a general transcription factor required for expression of many genes, and that it can interact with the STE3 UAS only with the help of alphal. A second goal of this proposal is to understand the mechanism by which a-factor pheromone alters the physiology of alpha cells, for instance to cause arrest of the cell division cycle. We have shown that STE3 encodes the cell surface receptor for a-factor. We will use in vitro mutagenesis of the STE3 gene in an effort to identify the part of the STE3 protein that is involved in production of the intracellular second-messenger. Isolation of phenotype suppressors of STE3 mutations will identify genes that play a role in the normal response to a-factor.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Research Project (R01)
Project #
Application #
Study Section
Genetics Study Section (GEN)
Project Start
Project End
Budget Start
Budget End
Support Year
Fiscal Year
Total Cost
Indirect Cost
University of Oregon
Organized Research Units
United States
Zip Code
Cullen, Paul J; Sprague Jr, George F (2012) The regulation of filamentous growth in yeast. Genetics 190:23-49
Cullen, Paul J; Xu-Friedman, Rufeng; Delrow, Jeffrey et al. (2006) Genome-wide analysis of the response to protein glycosylation deficiency in yeast. FEMS Yeast Res 6:1264-73
Sprague, George F; Cullen, Paul J; Goehring, April S (2004) Yeast signal transduction: regulation and interface with cell biology. Adv Exp Med Biol 547:91-105
Cullen, Paul J; Sabbagh Jr, Walid; Graham, Ellie et al. (2004) A signaling mucin at the head of the Cdc42- and MAPK-dependent filamentous growth pathway in yeast. Genes Dev 18:1695-708
Keniry, Megan E; Kemp, Hilary A; Rivers, David M et al. (2004) The identification of Pcl1-interacting proteins that genetically interact with Cla4 may indicate a link between G1 progression and mitotic exit. Genetics 166:1177-86
Kemp, Hilary A; Sprague Jr, George F (2003) Far3 and five interacting proteins prevent premature recovery from pheromone arrest in the budding yeast Saccharomyces cerevisiae. Mol Cell Biol 23:1750-63
Goehring, April S; Mitchell, David A; Tong, Amy Hin Yan et al. (2003) Synthetic lethal analysis implicates Ste20p, a p21-activated potein kinase, in polarisome activation. Mol Biol Cell 14:1501-16
Rivers, D M; Sprague Jr, G F (2003) Autocrine activation of the pheromone response pathway in matalpha2- cells is attenuated by SST2- and ASG7-dependent mechanisms. Mol Genet Genomics 270:225-33
Goehring, April S; Rivers, David M; Sprague Jr, George F (2003) Urmylation: a ubiquitin-like pathway that functions during invasive growth and budding in yeast. Mol Biol Cell 14:4329-41
Keniry, Megan E; Sprague Jr, George F (2003) Identification of p21-activated kinase specificity determinants in budding yeast: a single amino acid substitution imparts Ste20 specificity to Cla4. Mol Cell Biol 23:1569-80

Showing the most recent 10 out of 52 publications