Pheromone-dependent ubiquitination of the mitogen-activated protein kinase kinase Ste7

Many cell signaling pathways are regulated by phosphorylation, ubiquitination, and degradation of constituent proteins. As with phosphorylation, protein ubiquitination can be reversed, through the action of ubiquitin-specific processing proteases (UBPs). Here we have analyzed 15 UBP disruption mutants in the yeast Saccharomyces cerevisiae and identified one (ubp3Delta) that acts specifically in the pheromone response pathway. Upon pheromone stimulation, ubp3Delta mutants accumulate unconjugated polyubiquitin chains as well as polyubiquitinated forms of the mitogen-activated protein kinase kinase Ste7. The ubp3Delta mutants exhibit a potentiated response to pheromone, as measured by in vivo MAP kinase activity, transcriptional induction, and cell cycle arrest. Signaling is likewise enhanced upon direct activation of Ste4 (G protein beta subunit) and Ste11 (Ste7 kinase) but not the downstream transcription factor Ste12. These findings reveal a mechanism by which pheromone-triggered ubiquitination of Ste7 can modulate the pheromone response in vivo.