The phosphotyrosyl phosphatase activator, ncs1p (Rrd1p), functions with cla4p to regulate the G(2)/M transition in Saccharomyces cerevisiae

The Saccharomyces cerevisiae p21-activated kinases, Ste20p and Cla4p, have individual functions but appear to share an essential function(s) as well b ecause a strain lacking both kinases is inviable. To learn more about the s hared function, we sought new mutations that were lethal in the absence of CLA4. This approach led to the identification of at least 10 complementatio n groups designated NCS (need CLA4 to survive). As for ste20 cla4-75 mutant s, most ncs cla4-75 double mutants were defective for septin localization d uring budding. One group, NCS1/RRD1 (YIL153w), did not confer this defect, however, and we investigated its function further. ncs1 Delta cla4 Delta ce lls arrested with elongated buds and short mitotic spindles. The morphologi cal defects and lethality were suppressed by mutations that abrogate the ce ll cycle morphogenetic checkpoint, CDC28Y19F or swel Delta. The connection to the cell cycle may be direct, as we detected a Cla4p-Cdc28p complex. NCS 1 encodes a protein with significant similarity to a mammalian phosphotyros yl phosphatase activator (PTPA) regulatory subunit for type 2A protein phos phatases (PP2As). Genetic and biochemical evidence suggested that the phosp hatase Sit4p is a target for Ncs1p. First, CLA4 and SIT4 were synthetically lethal. Second, Ncs1p and its yeast paralog, Noh1p (Rrd2p), bound to the c atalytic domain of Sit4p in vitro, and Ncs1p could be immunoprecipitated wi th Sit4p but not with another PP2A (Pph21p) from yeast cell extracts. Strai ns lacking both NCS1 and NOH1 were inviable and arrested as unbudded cells, implying that PTPA function is required for proper G(1) progression.