title: Sck1 negatively regulates Gpa2-mediated glucose signaling in Schizosaccharomyces pombe.
abstract: Schizosaccharomyces pombe detects extracellular glucose via a G protein-mediated cyclic AMP (cAMP)-signaling pathway activating protein kinase A (PKA) and regulating transcription of genes involved in metabolism and sexual development. In this pathway, Gpa2 Gα binds to and activates adenylyl cyclase in response to glucose detection by the Git3 G protein-coupled receptor. Using a two-hybrid screen to identify extrinsic regulators of Gpa2, we isolated a clone that expresses codons 471 to 696 of the Sck1 kinase, which appears to display a higher affinity for Gpa2(K270E)-activated Gα relative to Gpa2(+) Gα. Deletion of sck1(+) or mutational inactivation of the Sck1 kinase produces phenotypes reflecting increased PKA activity in strains expressing Gpa2(+) or Gpa2(K270E), suggesting that Sck1 negatively regulates PKA activation through Gpa2. In contrast to the Gpa2(K270E) GDP-GTP exchange rate mutant, GTPase-defective Gpa2(R176H) weakly binds Sck1 in the two-hybrid screen and a deletion of sck1(+) in a Gpa2(R176H) strain confers phenotypes consistent with a slight reduction in PKA activity. Finally, deleting sck1(+) in a gpa2Δ strain results in phenotypes consistent with a second role for Sck1 acting in parallel with PKA. In addition to this parallel role with PKA, our data suggest that Sck1 negatively regulates Gpa2, possibly targeting the nucleotide-free form of the protein that may expose the one and only AKT/PKB consensus site in Gpa2 for Sck1 to bind. This dual role for Sck1 may allow S. pombe to produce distinct biological responses to glucose and nitrogen starvation signals that both activate the Wis1-Spc1/StyI stress-activated protein kinase (SAPK) pathway.
authors.affiliation: Biology Department, Boston College, Chestnut Hill, Massachusetts, USA.
authors.forename: Dayna K
authors.initials: DK
authors.lastname: Mudge
authors.forename: Fan
authors.initials: F
authors.lastname: Yang
authors.forename: Brian M
authors.initials: BM
authors.lastname: Currie
authors.forename: James M
authors.initials: JM
authors.lastname: Kim
authors.forename: Kelly
authors.initials: K
authors.lastname: Yeda
authors.forename: Varoon K
authors.initials: VK
authors.lastname: Bashyakarla
authors.forename: F Douglas
authors.initials: FD
authors.lastname: Ivey
authors.forename: Charles S
authors.initials: CS
authors.lastname: Hoffman
mesh_terms: D044385:GTP-Binding Protein alpha Subunits / Q000235:genetics / Q000378:metabolism*; D005947:Glucose / Q000378:metabolism*; D009154:Mutation; D011485:Protein Binding; D011494:Protein Kinases / Q000235:genetics / Q000378:metabolism*; D012568:Schizosaccharomyces / Q000235:genetics* / Q000378:metabolism; D029702:Schizosaccharomyces pombe Proteins / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction
chemical_list: D044385:GTP-Binding Protein alpha Subunits; D029702:Schizosaccharomyces pombe Proteins; D011494:Protein Kinases; C107804:Sck1 protein, S pombe; C540695:gpa2 protein, S pombe; D005947:Glucose