Pbp1 is a component of glucose deprivation induced stress granules and is involved in P-body-dependent granule assembly. We have recently shown that Pbp1 plays an important role in the interplay between three sensory protein kinases in yeast: AMP-regulated kinase (Snf1 in yeast), PAS kinase 1 (Psk1 in yeast), and the target of rapamycin complex 1 (TORC1), to regulate glucose allocation during nutrient depletion. This signaling cascade occurs through the SNF1-dependent phosphorylation and activation of Psk1, which phosphorylates and activates poly(A)- binding protein binding protein 1 (Pbp1), which then inhibits TORC1 through sequestration at stress granules. In this study we further characterized the regulation of Pbp1 by PAS kinase through the characterization of the role of the Psk1 homolog (Psk2) in Pbp1 regulation, and the identification of functional Pbp1 binding partners. Human ataxin-2 (ATXN2) is the homolog of yeast Pbp1 and has been shown to play an important role in the development of several ataxias. In this study we have also provided the evidence that human ataxin-2 can complement Pbp1 in yeast, and that human PAS kinase can phosphorylate human ataxin-2. Further characterizing this interplay between PAS kinase and Pbp1/ATXN2 aid in understanding pathways required for proper glucose allocation during nutrient depletion, including reducing cell growth and proliferation when energy is low. In addition, it yields valuable insights into the role of ataxin-2 in the development of devastating ataxias.

Nutrient sensing kinases sense available nutrients and regulate cell activity accordingly. Three of these enzymes are AMP regulated kinase (AMPK, or Snf1 in yeast), PAS kinase, and target of rapamycin (TOR), are conserved from yeast to man and have overlapping function. AMPK and Snf1 are important in sensing when nutrient status in the cell is low and down regulating energy consuming pathways. PAS kinase is required for glucose homeostasis in the cell, and responds to glucose levels. TOR senses nutrients such as amino acids and upregulates cell growth pathways primarily through protein synthesis. Due to the varying nature of these enzymes, cross talk is expected in order for the cell to properly regulate cellular metabolism and growth in response to energy and nutrient availability. Previous studies have shown that activation of yeast PAS kinase under nutrient stress conditions requires the presence of Snf1. The aim of this thesis is to determine whether Snf1 directly phosphorylates and activates PAS kinase through both in vivo and in vitro approaches. PAS kinase was found to require Snf1 for both activation and phosphorylation in vivo. In vitro kinase assays were also performed to confirm a direct phosphorylation event. The results from this study support the direct phosphorylation and activation of PAS kinase by Snf1, linking cellular energy status to glucose allocation.