In Saccharomyces cerevisiae, PHO85 encodes a cyclin-dependent protein kinase (Cdk) with multiple roles in cell cycle and metabolic controls. In association with the cyclin Pho80, Pho85 controls acid phosphatase gene expression through phosphorylation of the transcription factor Pho4. Pho85 has also been implicated as a kinase that phosphorylates and negatively regulates glycogen synthase (Gsy2), and deletion of PHO85 causes glycogen overaccumulation. We report that the Pcl8/Pcl10 subgroup of cyclins directs Pho85 to phosphorylate glycogen synthase both in vivo and in vitro. Disruption of PCL8 and PCL10 caused hyperaccumulation of glycogen, activation of glycogen synthase, and a reduction in glycogen synthase kinase activity in vivo. However, unlike pho85 mutants, pcl8 pcl10 cells had normal morphologies, grew on glycerol, and showed proper regulation of acid phosphatase gene expression. In vitro, Pho80-Pho85 complexes effectively phosphorylated Pho4 but had much lower activity toward Gsy2. In contrast, Pcl10-Pho85 complexes phosphorylated Gsy2 at Ser-654 and Thr-667, two physiologically relevant sites, but only poorly phosphorylated Pho4. Thus, both the in vitro and in vivo substrate specificity of Pho85 is determined by the cyclin partner. Mutation of PHO85 suppressed the glycogen storage deficiency of snf1 or glc7-1 mutants in which glycogen synthase is locked in an inactive state. Deletion of PCL8 and PCL10 corrected the deficit in glycogen synthase activity in both the snf1 and glc7-1 mutants, but glycogen synthesis was restored only in the glc7-1 mutant strain. This genetic result suggests an additional role for Pho85 in the negative regulation of glycogen accumulation that is independent of Pcl8 and Pcl10.
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