a b s t r a c tThe stress-activated kinase Hog1p mediates arsenic tolerance by decreasing arsenite influx through the aquaglyceroporin Fps1p in Saccharomyces cerevisiae. Unexpectedly, we found that overexpression of FPS1 increased arsenite tolerance suggesting a physiological role of Fps1p in arsenic detoxification. Consistently, during arsenite treatment transcription of FPS1 gene was strongly upregulated, while Fps1p was not degraded and remained localized to the plasma membrane. Moreover, deletion of FPS1 gene resulted in arsenate sensitivity. Finally, transport experiments revealed that Fps1p in concert with the arsenite transporter Acr3p mediates arsenite efflux.

Cells slow down cell cycle progression in order to adapt to unfavorable stress conditions. Yeast (Saccharomyces cerevisiae) responds to osmotic stress by triggering G 1 and G 2 checkpoint delays that are dependent on the mitogen-activated protein kinase (MAPK) Hog1. The high-osmolarity glycerol (HOG) pathway is also activated by arsenite, and the hog1⌬ mutant is highly sensitive to arsenite, partly due to increased arsenite influx into hog1⌬ cells. Yeast cell cycle regulation in response to arsenite and the role of Hog1 in this process have not yet been analyzed. Here, we found that long-term exposure to arsenite led to transient G 1 and G 2 delays in wild-type cells, whereas cells that lack the HOG1 gene or are defective in Hog1 kinase activity displayed persistent G 1 cell cycle arrest. Elevated levels of intracellular arsenite and "cross talk" between the HOG and pheromone response pathways, observed in arsenite-treated hog1⌬ cells, prolonged the G 1 delay but did not cause a persistent G 1 arrest. In contrast, deletion of the SIC1 gene encoding a cyclin-dependent kinase inhibitor fully suppressed the observed block of G 1 exit in hog1⌬ cells. Moreover, the Sic1 protein was stabilized in arsenite-treated hog1⌬ cells. Interestingly, Sic1-dependent persistent G 1 arrest was also observed in hog1⌬ cells during hyperosmotic stress. Taken together, our data point to an important role of the Hog1 kinase in adaptation to stress-induced G 1 cell cycle arrest.

The existence of a proteolytic system which can specifically recognize and cleave proteins in mitochondria is now well established. The components of this system comprise processing peptidases, ATP-dependent peptidases and oligopeptidases. A short overview of experimentally confirmed proteases mainly from Arabidopsis thaliana is provided. The role of the mitochondrial peptidases in plant growth and development is emphasized. We also discuss the possibility of existence of as yet unidentified plant homologs of yeast mitochondrial ATP-independent proteases.

The Saccharomyces cerevisiae High-Osmolarity Glycerol (HOG) pathway is a conserved mitogen-activated protein kinase (MAPK) signal transduction system that often serves as a model to analyze systems level properties of MAPK signaling. Hog1, the MAPK of the HOG-pathway, can be activated by various environmental cues and it controls transcription, translation, transport, and cell cycle adaptations in response to stress conditions. A powerful means to study signaling in living cells is to use kinase inhibitors; however, no inhibitor targeting wild-type Hog1 exists to date. Herein, we describe the design, synthesis, and biological application of small molecule inhibitors that are cell-permeable, fast-acting, and highly efficient against wild-type Hog1. These compounds are potent inhibitors of Hog1 kinase activity both in vitro and in vivo. Next, we use these novel inhibitors to pinpoint the time of Hog1 action during recovery from G1 checkpoint arrest, providing further evidence for a specific role of Hog1 in regulating cell cycle resumption during arsenite stress. Hence, we describe a novel tool for chemical genetic analysis of MAPK signaling and provide novel insights into Hog1 action.