Snf1p phosphorylation is required for Cd tolerance. (A) Cells lacking Snf1p or proteins regulating its activity (Snf4p or the upstream kinases Sak1p, Tos3p, Elm1p) are cadmium sensitive. 10-fold serial dilutions of exponentially growing cells were spotted on YNB + 2% glucose agar plates containing cadmium and growth was scored after 2–3 days at 30°C. (B) Snf1p phosphorylation/kinase activity is required for cadmium tolerance.snf1 Δ cells were transformed with a plasmid containing SNF1 or the unphosphorylatable SNF1-T210A allele. Wild-type and snf1 Δ cells were also transformed with the empty vector as a control. 10-fold serial dilutions of exponentially growing cells were spotted on selective medium with or without cadmium, and growth was scored after 2–3 days at 30°C. (C) Snf1p phosphorylation is not induced by cadmium. Snf1p phosphorylation was monitored with a phospho-specific-Snf1p antibody whereas total Snf1p was detected with anti-HA antibody. H: high glucose (2%); L: low glucose (0.05%). (D) Mig1p is not phosphorylated during cadmium exposure. Mig1p was detected with anti-HA antibody and mobility was monitored in the presence of cadmium, low glucose (L: 0.05%) or high glucose (H: 2%).