Design, Synthesis, and Characterization of a Highly Effective Hog1 Inhibitor: A Powerful Tool for Analyzing MAP Kinase Signaling in Yeast
Figure 8
Hog1 kinase activity is required to relieve As(III)-induced G1 checkpoint arrest.
(A) Kinetics of Hog1 activation during G1 checkpoint adaptation in response to As(III) stress. Hog1 phosphorylation was monitored as in Figure 6. (B) HOG1 deletion or the addition of 4b resulted in persistent G1 arrest in the presence of As(III). (C) As(III)-induced G1 checkpoint delay can be prolonged by addition of 4b until just before onset of the S phase. (D) Removal of 4b quickly relieves G1 arrest. Wild-type (W303-1A) and the isogenic HOG1 deletion mutant (hog1Δ) were synchronized in G1 with 5 µM α-factor and released in fresh medium in the presence or absence of 0.5 mM sodium arsenite [As(III)]. 4b (1 µM) was added as indicated. After washing out the inhibitor (in 7D), the cells were resuspended in fresh medium containing 0.5 mM As(III). The percentage of cells that remained in G1 was determined by the α-factor-nocodazole trap assay.