Sit4p-mediated dephosphorylation of Atp2p regulates ATP synthase activity and mitochondrial function

Sit4p is a type 2A-related protein phosphatase in Saccharomyces cerevisiae involved in a wide spectrum of cellular functions, including the glucose repression of mitochondrial transcription. Here we report that Sit4p is also involved in post-translational regulation of mitochondrial proteins and identified 9 potential targets. One of these, the ATP synthase (FoF1 complex) beta subunit Atp2p, was characterized and two phosphorylation sites, T124 and T317, were identified. Expression of Atp2p-T124 or T317 phosphoresistant versions in sit4Δ cells decreased Atp2p phosphorylation confirming these as Sit4p-regulated sites. Moreover, Sit4p and Atp2p interacted both physically and genetically. Mimicking phosphorylation at T124 or T317 increased Atp2p levels, resulting in higher abundance/activity of ATP synthase. Similar changes were observed in sit4Δ cells in which Atp2p is endogenously more phosphorylated. Expression of Atp2-T124 or T317 phosphomimetics also increased mitochondrial respiration and ATP levels and extended yeast lifespan. These results suggest that Sit4p-mediated dephosphorylation of Atp2p-T124/T317 downregulates Atp2p alongside with ATP synthase and mitochondrial function. Combination of transcriptional with post-translational regulation during fermentative growth may allow for a more efficient Sit4p repression of mitochondrial respiration.