INTERACTIONS BETWEEN CAMP-DEPENDENT AND SNF1 PROTEIN-KINASES IN THE CONTROL OF GLYCOGEN ACCUMULATION IN SACCHAROMYCES-CEREVISIAE

The synthesis of glycogen in Saccharomyces cerevisiae is stimulated by nutrient limitation and requires both glycogen synthase and the glyco gen branching enzyme. Of the two glycogen synthase genes present in ye ast, GSY2 appears to be more important for the accumulation of glycoge n upon entry into stationary phase. In cells grown on glucose, GSY2 mR NA levels increased approximately 10-fold during the transition from l ogarithmic to stationary phase. Growth of cells in glycerol, however, resulted in constitutive expression of GSY2 mRNA and the corresponding protein, GS-2, suggestive of glucose repression of GSY2. Mutants defe ctive in the SNF1 gene, which encodes a protein kinase important in gl ucose repression mechanisms, are known not to accumulate glycogen. A m odest 2-4-fold decrease in total GS-2 level was observed, and upon ent ry into stationary phase, the enzyme was blocked in the inactive, phos phorylated state in snf1 strains. The GS-2 protein is thought to be re gulated by covalent phosphorylation of three COOH-terminal sites (Hard y, T.A., and Roach, P. J. (1993) J. Biol. Chem. 268, 23799-23805), rem oval of which results in constitutively active glycogen synthase that bypasses phosphorylation controls. Expression of COOH-terminally trunc ated GS-2 in snf1 cells restored glycogen accumulation, and so we prop ose that the SNF1 kinase controls the phosphorylation state of GS-2. C yclic AMP pathways also exert control over glycogen accumulation. In b cy1 cells, which have constitutively active cyclic AMP-dependent prote in kinase, greatly reduced levels of both GS-2 message and protein wer e observed. With wild type GSY2 placed under control of the ADH1 promo ter, bcy1 cells did not accumulate glycogen despite increased GS-2. Ov erexpression of truncated GS-2, however, resulted in definite though r educed glycogen accumulation; the glycogen synthesized was structurall y distinct from wild type with properties characteristic of less branc hed polysaccharide. We conclude that the cAMP pathway controls both th e expression and the phosphorylation state of GS-2. Furthermore, other factor(s) necessary for glycogen biosynthesis, such as the branching enzyme GLC3, must also be under negative control by the cAMP pathway. The results demonstrate interactive controls of GS-2 by the cAMP-depen dent and SNF1 protein kinases.