Systematic analysis of essential genes reveals important regulators of G protein signaling.
The yeast pheromone pathway consists of a canonical heterotrimeric G protein and MAP kinase cascade. To identify additional signaling components, we systematically evaluated 870 essential genes using a library of repressible-promoter strains. Quantitative transcription-reporter and MAPK activity assays were used to identify strains that exhibit altered pheromone sensitivity. Of the 92 newly identified essential genes required for proper G protein signaling, those involved with protein degradation were most highly represented. Included in this group are members of the Skp, Cullin, F box (SCF) ubiquitin ligase complex. Further genetic and biochemical analysis reveals that SCF(Cdc4) acts together with the Cdc34 ubiquitin-conjugating enzyme at the level of the G protein; promotes degradation of the G protein alpha subunit, Gpa1, in vivo; and catalyzes Gpa1 ubiquitination in vitro. These insights to the G protein signaling network reveal the essential genome as an untapped resource for identifying new components and regulators of signal transduction pathways.
Cappell
Steven D
SD
Department of Pharmacology, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA.
Baker
Rachael
R
Skowyra
Dorota
D
Dohlman
Henrik G
HG
D064173:Anaphase-Promoting Complex-Cyclosome; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism; D016000:Cluster Analysis; D044783:F-Box Proteins / Q000235:genetics / Q000378:metabolism; D043802:GTP-Binding Protein alpha Subunits, Gq-G11 / Q000235:genetics / Q000378:metabolism; D044387:GTP-Binding Protein beta Subunits / Q000235:genetics / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D020869:Gene Expression Profiling; D015966:Gene Expression Regulation, Fungal; D016681:Genome, Fungal; D006801:Humans; D010641:Phenotype; D010675:Pheromones / Q000235:genetics / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D015203:Reproducibility of Results; D044843:SKP Cullin F-Box Protein Ligases / Q000235:genetics / Q000378:metabolism; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction / Q000235:genetics*; D013379:Substrate Specificity; D044763:Ubiquitin-Conjugating Enzymes; D043743:Ubiquitin-Protein Ligase Complexes / Q000235:genetics / Q000378:metabolism; D044767:Ubiquitin-Protein Ligases / Q000235:genetics / Q000378:metabolism
C080443:CDC4 protein, S cerevisiae; D018797:Cell Cycle Proteins; D044783:F-Box Proteins; D044387:GTP-Binding Protein beta Subunits; D010675:Pheromones; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; C067576:Ste4 protein, S cerevisiae; C577449:CDC34 protein, S cerevisiae; C577447:CDC34 protein, human; D044763:Ubiquitin-Conjugating Enzymes; D043743:Ubiquitin-Protein Ligase Complexes; D064173:Anaphase-Promoting Complex-Cyclosome; D044843:SKP Cullin F-Box Protein Ligases; D044767:Ubiquitin-Protein Ligases; D019204:GTP-Binding Proteins; C084293:GPA1 protein, S cerevisiae; D043802:GTP-Binding Protein alpha Subunits, Gq-G11