Complementation of cell stress, polarity, and cell cycle phenotypes in a pho85Δ strain by CDK5 expression. (A) Salt sensitivity. Serial dilutions of log phase cultures of pho85 or wild-type cells expressing either CDK5 or CDK5-I plasmids were plated on SG-URA medium with 0.75 M NaCl or on an SD-URA plate (control) and incubated for 5 days at 30°C. (B) Overexpression of CDK5 in a cln1Δ cln2Δ pho85Δ strain. A cln1Δ cln2Δ pho85Δ strain carrying a pho85 mutant allele (pho85ts17) expressed from the MET25 promoter (BY794) and either pCDK5 (right side of plates) or pCDK5-I (left side of plates) was streaked on a galactose-containing plate plus methionine (+ Gal + Met, Right; repress pho85ts17, induce CDK5) or a galactose-containing plate minus methionine (control, Left) and incubated for 5 days at 30°C. Two independent isolates are shown. (C) Overexpression of CDK5 in a slt2Δ pho85Δ strain. A slt2Δ pho85Δ strain expressing either PHO85 (left side of plates) or CDK5 (right side of plates) from the GAL promoter was streaked onto a yeast extract/peptone/dextrose (YPD) or yeast extract/peptone/galactose (YPG) plate as indicated and incubated for 5 days at 25°C. (D) Pho4 kinase activity associated with HA-Pcl2-Cdk5. Ha-Pcl2 was immunoprecipitated from yeast extracts and used in kinase assays with Pho4 as a substrate. The following strains were used for extract preparation: lane 1, pho85Δ HA-PCL2 with pCDK5; lane 2, pho85Δ HA-PCL2 with pCDK5-I; lane 3, pho85Δ HA-PCL2 with pPHO85 (positive control).