A constitutively active G‐protein‐coupled receptor causes mating self‐compatibility in the mushroom Coprinus

In the mushroom Coprinus cinereus, the multiallelic B mating type genes are predicted to encode a large family of seven‐transmembrane domain receptors and CaaX‐modified pheromones. We have shown that a single amino acid change Q229P in transmembrane domain VI of one receptor confers a self‐compatible mating phenotype. Using a heterologous yeast assay, we have demonstrated that this C.cinereus pheromone receptor is a G‐protein‐coupled receptor and that the Q229P mutation is constitutively activating. A C.cinereus pheromone precursor was processed to an active species specifically in yeast MATa cells and activated the co‐expressed wild‐type receptor. Yeast cells expressing the wild‐type receptor were used to test the activity of synthetic peptides, enabling us to predict the structure of the mature C.cinereus pheromone and to show that the Q229P mutation does not compromise normal receptor function.

[1]  R. Kahmann,et al.  Crosstalk between cAMP and pheromone signalling pathways in Ustilago maydis , 1998, Molecular and General Genetics MGG.

[2]  A. Brown,et al.  Mapping of a yeast G protein betagamma signaling interaction. , 1998, Genetics.

[3]  L. Vaillancourt,et al.  Pheromones and pheromone receptors as mating-type determinants in basidiomycetes. , 1996, Genetic engineering.

[4]  G. Fink,et al.  Methods in enzymology vol 194 guide to yeast genetics and molecular biology , 1991 .

[5]  M. Bölker,et al.  Sexual pheromones and mating responses in fungi. , 1993, The Plant cell.

[6]  R. Lefkowitz,et al.  Enhanced myocardial function in transgenic mice overexpressing the beta 2-adrenergic receptor. , 1994, Science.

[7]  J J Clare,et al.  Production of mouse epidermal growth factor in yeast: high-level secretion using Pichia pastoris strains containing multiple gene copies. , 1991, Gene.

[8]  J. Thorner,et al.  Control of yeast mating signal transduction by a mammalian beta 2-adrenergic receptor and Gs alpha subunit. , 1990, Science.

[9]  L. Casselton,et al.  Molecular Genetics of Mating Recognition in Basidiomycete Fungi , 1998, Microbiology and Molecular Biology Reviews.

[10]  G. Sprague,,et al.  Evidence the yeast STE3 gene encodes a receptor for the peptide pheromone a factor: gene sequence and implications for the structure of the presumed receptor. , 1986, Proceedings of the National Academy of Sciences of the United States of America.

[11]  Y. Kang,et al.  Effects of expression of mammalian G alpha and hybrid mammalian-yeast G alpha proteins on the yeast pheromone response signal transduction pathway , 1990, Molecular and cellular biology.

[12]  J. Weijer Genetics of Sexuality in Higher Fungi , 1966 .

[13]  J. Kurjan The pheromone response pathway in Saccharomyces cerevisiae. , 1993, Annual review of genetics.

[14]  Sangram S. Sisodia,et al.  Dual Roles for Ste24p in Yeast a-Factor Maturation: NH2-terminal Proteolysis and COOH-terminal CAAX Processing , 1998, The Journal of cell biology.

[15]  J. Kurjan The Saccharomyces cerevisiae Pheromone Response Pathway , 1993 .

[16]  R. Rothstein Targeting, disruption, replacement, and allele rescue: integrative DNA transformation in yeast. , 1991, Methods in enzymology.

[17]  M. Whiteway,et al.  Pheromone signalling and polarized morphogenesis in yeast. , 1997, Current opinion in genetics & development.

[18]  M. Bölker,et al.  G proteins in Ustilago maydis: transmission of multiple signals? , 1997, The EMBO journal.

[19]  M. Brann,et al.  Pharmacology of a constitutively active muscarinic receptor generated by random mutagenesis. , 1995, The Journal of pharmacology and experimental therapeutics.

[20]  I. Connerton,et al.  A large pheromone and receptor gene complex determines multiple B mating type specificities in Coprinus cinereus. , 1998, Genetics.

[21]  C. Specht,et al.  Multiple genes encoding pheromones and a pheromone receptor define the B beta 1 mating-type specificity in Schizophyllum commune. , 1997, Genetics.

[22]  R. Sikorski,et al.  A system of shuttle vectors and yeast host strains designed for efficient manipulation of DNA in Saccharomyces cerevisiae. , 1989, Genetics.

[23]  L. Casselton,et al.  Dikaryon Formation in Coprinus cinereus : Selection and Identification of B Factor Mutants , 1980 .

[24]  L. Guarente Yeast promoters and lacZ fusions designed to study expression of cloned genes in yeast. , 1983, Methods in enzymology.

[25]  Bruce R. Conklin,et al.  Structural elements of Gα subunits that interact with Gβγ, receptors, and effectors , 1993, Cell.

[26]  I. Herskowitz,et al.  The a-factor pheromone of Saccharomyces cerevisiae is essential for mating , 1988, Molecular and cellular biology.

[27]  J. Konopka,et al.  Identification of a Polar Region in Transmembrane Domain 6 That Regulates the Function of the G Protein-Coupled α-Factor Receptor , 1998, Molecular and Cellular Biology.

[28]  A. Wach PCR‐synthesis of marker cassettes with long flanking homology regions for gene disruptions in S. cerevisiae , 1996, Yeast.

[29]  K. Arai,et al.  Regulation of the yeast pheromone response pathway by G protein subunits. , 1990, The EMBO journal.

[30]  B. Ozenberger,et al.  Functional coupling of a mammalian somatostatin receptor to the yeast pheromone response pathway , 1995, Molecular and cellular biology.

[31]  J. Konopka,et al.  Mutation of Pro-258 in transmembrane domain 6 constitutively activates the G protein-coupled alpha-factor receptor. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[32]  J. Edman,et al.  The alpha-mating type locus of Cryptococcus neoformans contains a peptide pheromone gene , 1993, Molecular and cellular biology.

[33]  J. W. Wells,et al.  Constitutive activation of the m5 muscarinic receptor by a series of mutations at the extracellular end of transmembrane 6. , 1997, Biochemistry.

[34]  M. Simon,et al.  Gα15 and Gα16 Couple a Wide Variety of Receptors to Phospholipase C (*) , 1995, The Journal of Biological Chemistry.

[35]  F. Lottspeich,et al.  Pheromones trigger filamentous growth in Ustilago maydis. , 1994, The EMBO journal.

[36]  L. Lin,et al.  A point mutation in the glucose-dependent insulinotropic peptide receptor confers constitutive activity. , 1997, Biochemical and biophysical research communications.

[37]  S. Rees,et al.  G16 as a universal G protein adapter: implications for agonist screening strategies. , 1996, Trends in pharmacological sciences.

[38]  I. Herskowitz,et al.  Regulation of yeast mating-type interconversion: feedback control of HO gene expression by the mating-type locus. , 1983, Proceedings of the National Academy of Sciences of the United States of America.

[39]  K. Świeżyński,et al.  Heterokaryon formation in Coprinus lagopus , 1960 .

[40]  A. Scheer,et al.  Constitutively active G protein-coupled receptors: potential mechanisms of receptor activation. , 1997, Journal of receptor and signal transduction research.

[41]  E. Kothe,et al.  The mating‐type locus B alpha 1 of Schizophyllum commune contains a pheromone receptor gene and putative pheromone genes. , 1995, The EMBO journal.

[42]  R. Lefkowitz,et al.  Enhanced Myocardial Function inTransgenic Mice Overexpressing theP2-Adrenergic Receptor , 1994 .

[43]  M. Bölker,et al.  The a mating type locus of U. maydis specifies cell signaling components , 1992, Cell.

[44]  H. Bourne,et al.  How receptors talk to trimeric G proteins. , 1997, Current opinion in cell biology.

[45]  I. Herskowitz MAP kinase pathways in yeast: For mating and more , 1995, Cell.

[46]  J M Becker,et al.  Fungal lipopeptide mating pheromones: a model system for the study of protein prenylation. , 1995, Microbiological reviews.

[47]  R. Müller,et al.  Yeast vectors for the controlled expression of heterologous proteins in different genetic backgrounds. , 1995, Gene.

[48]  D. Lewis Genetical analysis of methionine suppressors in Coprinus , 1961 .