A MEMBRANE-ASSOCIATED COMPLEX CONTAINING THE VPS15 PROTEIN-KINASE ANDTHE VPS34 PI 3-KINASE IS ESSENTIAL FOR PROTEIN SORTING TO THE YEAST LYSOSOME-LIKE VACUOLE

The Vps15 protein kinase and the Vps34 phosphatidylinositol 3-kinase ( PI 3-kinase) are required for the sorting of soluble hydrolases to the yeast vacuole. Overproduction of Vps34p suppresses the growth and vac uolar protein sorting defects associated with vps15 kinase domain muta nts, suggesting that Vps15p and Vps34p functionally interact. Subcellu lar fractionation and sucrose density gradients indicate that Vps15p i s responsible for the association of Vps34p with an intracellular memb rane fraction. Chemical cross-linking and native immunoprecipitation e xperiments demonstrate that Vps15p and Vps34p interact as components o f a hetero-oligomeric protein complex. In addition, we show that an in tact Vps15 protein kinase domain is required for activation of the Vps 34 PI 3-kinase, suggesting that the Vps34 lipid kinase is regulated by a Vps15p-mediated protein phosphorylation event. We propose that Vps1 5p and Vps34p function together as components of a membrane-associated signal transduction complex that regulates intracellular protein traf ficking decisions through and lipid phosphorylation events.