Two activating phosphorylation sites of Pbs2 MAP2K in the yeast HOG pathway are differentially dephosphorylated by four PP2C phosphatases Ptc1-Ptc4

To cope with an increased external osmolarity, the budding yeast Saccharomyces cerevisiae activates the Hog1 mitogen-activated protein kinase (MAPK) through the high-osmolarity glycerol (HOG) pathway, which governs adaptive responses to osmostress. In the HOG pathway, two apparently redundant upstream branches, termed SLN1 and SHO1, activate cognate MAP3Ks (MAPKK kinase) Ssk2/22 and Ste11, respectively. These MAP3Ks, when activated, phosphorylate and thus activate the Pbs2 MAP2K (MAPK kinase), which in turn phosphorylates and activates Hog1. Previous studies have shown that protein tyrosine phospha-tases and the serine/threonine protein phosphatases type 2C negatively regulate the HOG pathway to prevent its excessive and inappropriate activation, which is detrimental to cell growth. The tyrosine phosphatases Ptp2 and Ptp3 dephos-phorylate Hog1 at Tyr-176, whereas the protein phosphatase type 2Cs Ptc1 and Ptc2 dephosphorylate Hog1 at Thr-174. In contrast, the identities of phosphatases that dephosphorylate Pbs2 remained less clear. Here, we examined the phosphor-ylation status of Pbs2 at the activating phosphorylation sites Ser-514 and Thr-518 (S514 and T518) in various mutants, both in the unstimulated and osmostressed conditions. Thus, we found that Ptc1-Ptc4 collectively regulate Pbs2 nega-tively, but each Ptc acts differently to the two phosphoryla-tion sites in Pbs2. T518 is predominantly dephosphorylated by Ptc1, while S514 can be dephosphorylated by any of Ptc1-4 to an appreciable extent. We also show that Pbs2 dephosphorylation by Ptc1 requires the adaptor protein Nbp2 that recruits Ptc1 to Pbs2, thus highlighting the complex processes involved in regulating adaptive responses to osmostress.

Automated summary

To adapt to increased osmolarity, the yeast Saccharomyces cerevisiae activates the Hog1 MAPK through the HOG pathway, which governs osmostress response. The SLN1 and SHO1 branches of the pathway activate the MAP3Ks Ssk2/22 and Ste11, respectively, which activate Pbs2 MAP2K to phosphorylate and activate Hog1. Protein phosphatases, including Ptp2, Ptp3, Ptc1, and Ptc2, negatively regulate the pathway. Dephosphorylation of Pbs2 at specific sites, particularly T518 by Ptc1, is crucial for modulating the adaptive response to osmostress. Nbp2 is required for Ptc1-mediated Pbs2 dephosphorylation. Rating: 8/10