A dynamic model for the GAL genetic regulatory network of Saccharomyces cerevisiae

Eukaryotic organisms carry tremendous amount of genetic information and possess exquisite cellular structures just to produce the 6000 to 40,000 proteins necessary to give rise to the extraordinary complexity of life spanning from yeast to human. In order to perpetuate life, an organism must achieve appropriate temporal and spatial regulation of gene activity. Studies of the genetic regulatory networks that enable cells to activate and repress expression of specific groups of genes in response to environmental cues have enriched our understanding of the intricate cellular processes that regulate gene activities. One well characterized genetic regulatory network of archetypical importance is the GAL network that dictates synthesis of galactose metabolic enzymes in the yeast Saccharomyces cerevisiae.; The GAL genetic regulatory network is composed of three regulatory genes, GAL4, GAL80, and GAL3 as well as eleven target genes that are involved in galactose metabolism and other cellular functions. Over four decades of genetic dissecting and molecular probing have established that the GAL genes are efficiently transcribed only when the sequence specific transcription factor Gal4p is activated. Activation of Gal4p requires the interaction between the Gal4p inhibitory protein Gal80p and the signal transducer protein Gal3p. A prevailing model specifies that Gal3p binds to a Gal80p-Gal4p complex in the nucleus to activate Gal4p. The cellular behavior of the Gal4, Gal80 and Gal3 proteins however were poorly defined. In this thesis study, I first determined the subcellular distribution profiles of Gal3p and Gal80p. Two independent methods showed that Gal3p is a cytoplasmic protein, whilst Gal80p is located in both the cytoplasm and nucleus. Further analysis demonstrated that Gal3p can exert its induction function solely in the cytoplasm; Gal80p is a nucleocytoplasmic shuttling protein; and GAL gene activation is a result of decreased binding between Gal80p and Gal4p. I also initiated a structure/function analysis for the Gal3 protein. Taken together, this thesis work indicates that the tightly controlled and highly efficient galactose-responsive mechanism in yeast is achieved by modulating protein-protein interactions located in two different cellular compartments. A dynamic model for the GAL genetic regulatory network is proposed that contrasts with the previous prevailing model.