title: TORC1 promotes phosphorylation of ribosomal protein S6 via the AGC kinase Ypk3 in Saccharomyces cerevisiae.
abstract: The target of rapamycin complex 1 (TORC1) is an evolutionarily conserved sensor of nutrient availability. Genetic and pharmacological studies in the yeast Saccharomyces cerevisiae have provided mechanistic insights on the regulation of TORC1 signaling in response to nutrients. Using a highly specific antibody that recognizes phosphorylation of the bona fide TORC1 target ribosomal protein S6 (Rps6) in yeast, we found that nutrients rapidly induce Rps6 phosphorylation in a TORC1-dependent manner. Moreover, we demonstrate that Ypk3, an AGC kinase which exhibits high homology to human S6 kinase (S6K), is required for the phosphorylation of Rps6 in vivo. Rps6 phosphorylation is completely abolished in cells lacking Ypk3 (ypk3Δ), whereas Sch9, previously reported to be the yeast ortholog of S6K, is dispensable for Rps6 phosphorylation. Phosphorylation-deficient mutations in regulatory motifs of Ypk3 abrogate Rps6 phosphorylation, and complementation of ypk3Δ cells with human S6 kinase restores Rps6 phosphorylation in a rapamycin-sensitive manner. Our findings demonstrate that Ypk3 is a critical component of the TORC1 pathway and that the use of a phospho-S6 specific antibody offers a valuable tool to identify new nutrient-dependent and rapamycin-sensitive targets in vivo.
authors.affiliation: Biozentrum, University of Basel, Basel, Switzerland.
authors.forename: Asier
authors.initials: A
authors.lastname: González
authors.affiliation: Biozentrum, University of Basel, Basel, Switzerland.
authors.forename: Mitsugu
authors.initials: M
authors.lastname: Shimobayashi
authors.affiliation: Institute of Molecular Biosciences, University of Graz, Graz, Austria.
authors.forename: Tobias
authors.initials: T
authors.lastname: Eisenberg
authors.affiliation: Institute of Molecular Biosciences, University of Graz, Graz, Austria.
authors.forename: David Adrian
authors.initials: DA
authors.lastname: Merle
authors.affiliation: Institute of Molecular Biosciences, University of Graz, Graz, Austria.
authors.forename: Tobias
authors.initials: T
authors.lastname: Pendl
authors.affiliation: Biozentrum, University of Basel, Basel, Switzerland.
authors.forename: Michael N
authors.initials: MN
authors.lastname: Hall
authors.affiliation: Institute of Molecular Biosciences, University of Graz, Graz, Austria.
authors.forename: Tarek
authors.initials: T
authors.lastname: Moustafa
mesh_terms: D000704:Analysis of Variance; D017867:Cyclic Nucleotide-Regulated Protein Kinases / Q000378:metabolism*; D004926:Escherichia coli; D015151:Immunoblotting; D010766:Phosphorylation; D010957:Plasmids / Q000235:genetics; D016133:Polymerase Chain Reaction; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D019893:Ribosomal Protein S6 Kinases / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism*; D014157:Transcription Factors / Q000378:metabolism*
chemical_list: D029701:Saccharomyces cerevisiae Proteins; C561842:TORC1 protein complex, S cerevisiae; D014157:Transcription Factors; D017867:Cyclic Nucleotide-Regulated Protein Kinases; D017346:Protein Serine-Threonine Kinases; D019893:Ribosomal Protein S6 Kinases; C000604510:Ypk3 protein, S cerevisiae