COORDINATED ACTIVATION OF THE FOUR MAPKS BY MATING PHEROMONE

RODRIGO BALTANÁS; DAVID LEVIN; STEFAN HOHMANN; ALEJANDRO COLMAN-LERNER

Cold Spring Harbor, New York, USA

Congreso; 2009 CSHL Meeting on Yeast Cell Biology; 2009

Cold Spring Harbor Laboratory

In this work we investigate how the coordinated activation of the mating pheromone and the cell wall integrity MAPK cascades modulate a third MAPK cascade, the “High Osmolarity Glycerol” response in S. cerevisiae. We studied these pathways at the single cell level using quantitative cytometry of transcriptional and relocalization-based fluorescent reporters, and volume changes; and, at the population level, using quantitative western blots.  The HOG pathway has always been thought of as a transient, homeostatic response to osmoshock. Here we found that in yeast adapted to grow in high osmolarity (e.g, 1M sorbitol or 0.5M NaCl), activation of the cell wall integrity pathway (CWIP) MAPK Mpk1/Slt2 leads to a persistent activation HOG pathway. We observed this behavior activating the CWIP with several different stimuli, including mating pheromone, butanol induced filamentation and high temperature shock.  The activation of the HOG pathway is seen at the level of phosphorilation of Hog1p, translocation of Hog1-YFP, and also at a transcriptional level using PSTL1-YFP. The output of the HOG response follows the dynamics typical of the inducing stimulus, continuous, stable over time when using pheromone and butanol, transient when using temperature. Pheromone-induced activation of HOG does not disappear in the sensor deletion mutants of the CWI pathway Mid2, Wsc1 and Mtl1 and it required essential components of the cascade Bck1 and the MAPK Slt2/Mpk1, but not the transcription factor Rlm1. Our data suggests a model in which the CWIP activates the HOG pathway indirectly, by opening the glyceroporin Fps1, leading to glycerol loss, and subsequent reduction of turgor pressure, leading to activation of the Sln1 branch of the HOG pathway. This hypothesis is supported by a) the observation that cells adapted to 1M glycerol (instead of 1M sorbitol) do not activate the HOG pathway after α factor treatment; and b) ΔFPS1 cells and Δssk1 cells fail to activate the HOG pathway. Finally, we found that the correct and coordinated activation of the MAPK network is biologically significant, since disruption of it leads to reduced mating efficiency in cells adapted to grow in high osmolarity.