FIG. 5.

A strain lacking PTC1 shows defects in Hog1 activation and inactivation. (A) Hog1 kinase activity and the level of Hog1 protein were examined in the wild-type strain IMY102 and in ptc1Δ strain IMY103. Hog1-HA was isolated from yeast cells that were untreated or exposed to 0.4 M sodium chloride for the indicated times, and Hog1 kinase activity was assessed as described in the legend to Fig. 4. The level of Hog1-HA was assessed using anti-HA antibody. (B) Dual phosphorylation of kinase-inactive Hog1-K52M is rapid in both wild-type ptc1Δ strains. PTC1 hog1Δ and ptc1Δ hog1Δ strains expressing Hog1-K52M-HA were exposed to 0.4 M NaCl for the indicated times. Activation of Hog1-K52M-HA was monitored using antibody specific for dually phosphorylated Hog1, and its relative level of expression was assessed using anti-HA antibody.