Implications of phosphorylation on Ser 179 of the C-subunit isoform TPK1 from yeast PKA

SOLARI, CLARITA; TUDISCA VANESA; MORENO, SILVIA; PORTELA PAULA

Mendoza

Congreso; XVLIII Reunion Anual de SAIB; 2012

Sociedad Argentina de Investigacio en Bioquimica y Biologia Molecular

Many protein kinases are themselves phosphoproteins, and their biological function and activity are frequently regulated by phosphorylation. PKA regulatory subunit is encoded by the BCY1 gene, and the catalytic subunits are encoded by three genes: TPK1, TPK2 and TPK3. Previously we have reported that following cAMP-PKA pathway activation, Tpk1 changes its phosphorylation status toward more phosphorylated isoforms by an intramolecular phosphorylation mechanism. Tpk1 increases its specific activity toward kemptide. Using mass spectrometry and array peptides derived from Tpk1 we identified mainly Ser as a putative target residue. Here we report the role of Ser phosphorylation of Tpk1 by several readouts of PKA. We have constructed strains containing Tpk1wt or Tpk1 or Tpk1 as sole source of PKA activity. Our results show that Tpk1 strain is deficient on non-fermentable carbon source growth, glycogen content, tolerance to heat stress, growth on rapamycin containing medium and shows a reduced life span on stationary phase. Tpk1-GFP and Tpk1 -GFP show a nucleo-cytoplasmic distribution in glucose growing cells whereas Tpk1 -GFP was localized in the nucleus. Our results suggest that the unphosphorylated state of Ser reduces the Tpk1 kinase activity improving the respiratory metabolism, survival to stress and life span of the yeast cells. 179 179 S179A S179D