PMID:18573873

Citation	Horie, T, Tatebayashi, K, Yamada, R and Saito, H (2008) Phosphorylated Ssk1 prevents unphosphorylated Ssk1 from activating the Ssk2 mitogen-activated protein kinase kinase kinase in the yeast high-osmolarity glycerol osmoregulatory pathway. Mol. Cell. Biol. 28:5172-83
Abstract	In Saccharomyces cerevisiae, external high osmolarity activates the Hog1 mitogen-activated protein kinase (MAPK), which controls various aspects of osmoadaptation. Ssk1 is a homolog of bacterial two-component response regulators and activates the Ssk2 MAPK kinase kinase upstream of Hog1. It has been proposed that unphosphorylated Ssk1 (Ssk1-OH) is the active form and that Ssk1 phosphorylated (Ssk1 approximately P) at Asp554 by the Sln1-Ypd1-Ssk1 multistep phosphorelay mechanism is the inactive form. In this study, we show that constitutive activation of Ssk2 occurs when Ssk1 phosphorylation is blocked by either an Ssk1 mutation at the phosphorylation site or an Ssk1 mutation that inhibits its interaction with Ypd1, the donor of phosphate to Ssk1. Thus, Ssk1-OH is indeed necessary for Ssk2 activation. However, overexpression of wild-type Ssk1 or of an Ssk1 mutant that cannot bind Ssk2 prevents constitutively active Ssk1 mutants from activating Ssk2. Therefore, Ssk1 has a dual function as both an activator of Ssk2 and an inhibitor of Ssk1 itself. We also found that Ssk1 exists mostly as a dimer within cells. From mutant phenotypes, we deduce that only the Ssk1-OH/Ssk1-OH dimer can activate Ssk2 efficiently. Hence, because Ssk1 approximately P binds to and inhibits Ssk1-OH, moderate fluctuation of the level of Ssk1-OH does not lead to nonphysiological and detrimental activation of Hog1.
Links	PubMed PMC2519728 Online version:10.1128/MCB.00589-08
Keywords	Amino Acid Substitution; Aspartic Acid; Dimerization; Enzyme Activation/drug effects; Genes, Dominant; Glycerol/pharmacology; MAP Kinase Kinase Kinases/chemistry; MAP Kinase Kinase Kinases/metabolism; Models, Biological; Mutation/genetics; Osmolar Concentration; Phosphorylation/drug effects; Protein Binding/drug effects; Protein Structure, Tertiary; Protein-Serine-Threonine Kinases/chemistry; Protein-Serine-Threonine Kinases/metabolism; Saccharomyces cerevisiae/drug effects; Saccharomyces cerevisiae/enzymology; Saccharomyces cerevisiae Proteins/chemistry; Saccharomyces cerevisiae Proteins/metabolism; Water-Electrolyte Balance/drug effects
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Annotations

Gene product	Qualifier	GO Term	Evidence Code	with/from	Aspect	Extension	Notes	Status
YEAST:SSK1	enables	GO:0042803: protein homodimerization activity	ECO:0000314: direct assay evidence used in manual assertion		F		Seeded From UniProt	complete
YEAST:SSK1	involved_in	GO:0000185: activation of MAPKKK activity	ECO:0000315: mutant phenotype evidence used in manual assertion		P	part_of:(GO:0071474) has_input:(UniProtKB:P53599) has_input:(UniProtKB:P25390)	Seeded From UniProt	complete

References

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