Phosphorylation of the MAPKKK regulator Ste50p in Saccharomyces cerevisiae: a casein kinase I phosphorylation site is required for proper mating function.
The Ste50 protein of Saccharomyces cerevisiae is a regulator of the Ste11p protein kinase. Ste11p is a member of the MAP3K (or MEKK) family, which is conserved from yeast to mammals. Ste50p is involved in all the signaling pathways that require Ste11p function, yet little is known about the regulation of Ste50p itself. Here, we show that Ste50p is phosphorylated on multiple serine/threonine residues in vivo. Threonine 42 (T42) is phosphorylated both in vivo and in vitro, and the protein kinase responsible has been identified as casein kinase I. Replacement of T42 with alanine (T42A) compromises Ste50p function. This mutation abolishes the ability of overexpressed Ste50p to suppress either the mating defect of a ste20 ste50 deletion mutant or the mating defect of a strain with a Ste11p deleted from its sterile-alpha motif domain. Replacement of T42 with a phosphorylation-mimetic aspartic acid residue (T42D) permits wild-type function in all assays of Ste50p function. These results suggest that phosphorylation of T42 of Ste50p is required for proper signaling in the mating response. However, this phosphorylation does not seem to have a detectable role in modulating the high-osmolarity glycerol synthesis pathway.
Wu
Cunle
C
Genetics Group, Biotechnology Research Institute, National Research Council of Canada, Montreal, Quebec, Canada H4P 2R2. cunle.wu@cnrc-nrc.gc.ca
Arcand
Mathieu
M
Jansen
Gregor
G
Zhong
Mei
M
Iouk
Tatiana
T
Thomas
David Y
DY
Meloche
Sylvain
S
Whiteway
Malcolm
M
D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D047388:Casein Kinases; D002455:Cell Division / Q000187:drug effects / Q000502:physiology; D017353:Gene Deletion; D047908:Intracellular Signaling Peptides and Proteins; D020930:MAP Kinase Kinase Kinases / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D008969:Molecular Sequence Data; D010766:Phosphorylation; D011401:Promoter Regions, Genetic / Q000235:genetics; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism*; D011498:Protein Precursors / Q000494:pharmacology; D017346:Protein Serine-Threonine Kinases / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000187:drug effects / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology / Q000502:physiology; D012694:Serine / Q000378:metabolism; D015398:Signal Transduction; D013012:Sorbitol / Q000494:pharmacology; D013347:Subcellular Fractions / Q000737:chemistry; D013912:Threonine / Q000378:metabolism
D047908:Intracellular Signaling Peptides and Proteins; C048632:MF(ALPHA)1 protein, S cerevisiae; D011498:Protein Precursors; C079506:STE50 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D013912:Threonine; D012694:Serine; D013012:Sorbitol; D011494:Protein Kinases; D047388:Casein Kinases; D017346:Protein Serine-Threonine Kinases; D020930:MAP Kinase Kinase Kinases; C472815:SSK2 protein, S cerevisiae; C476806:SSK22 protein, S cerevisiae; C078754:STE20 protein, S cerevisiae; C067018:Ste11 protein, S cerevisiae