THE SNF1 PROTEIN-KINASE AND ITS ACTIVATING SUBUNIT, SNF4, INTERACT WITH DISTINCT DOMAINS OF THE SIP1 SIP2/GA183 COMPONENT IN THE KINASE COMPLEX/

The Snf1 protein kinase plays a central role in the response to glucos e starvation in the yeast Saccharomyces cerevisiae. Previously, we sho wed that two-hybrid interaction between Snf1 and its activating subuni t, Snf4, is inhibited by high levels of glucose. These findings, toget her with biochemical evidence that Snf1 and Snf4 remain associated in cells grown in glucose, suggested that another protein (or proteins) a nchors Snf1 and Snf4 into a complex. Here, we examine the possibility that a family of proteins, comprising Sip1, Sip2, and Ga183, serves th is purpose. We first show that the fraction of cellular Snf4 protein t hat is complexed with Snf1 is reduced in a sip1 Delta sip2 Delta ga183 Delta triple mutant. We then present evidence that Sipl, Sip2, and Ga 183 each interact independently with both Snf1 and Snf4 via distinct d omains. A conserved internal region binds to the Snf1 regulatory domai n, and the conserved C-terminal ASC domain binds to Snf4. Interactions were mapped by using the two-hybrid system and were confirmed by in v itro binding studies. These findings indicate that the Sip1/Sip2/Ga183 family anchors Snf1 and Snf4 into a complex. Finally, the interaction of the yeast Sip2 protein with a plant Snf1 homolog suggests that thi s function is conserved in plants.