title: Quantification of mutation-derived bias for alternate mating functionalities of the Saccharomyces cerevisiae Ste2p pheromone receptor.
abstract: Although well documented for mammalian G-protein-coupled receptors, alternate functionalities and associated alternate signalling remain to be unequivocally established for the Saccharomyces cerevisiae pheromone Ste2p receptor. Here, evidence supporting alternate functionalities for Ste2p is re-evaluated, extended and quantified. In particular, strong mating and constitutive signalling mutations, focusing on residues S254, P258 and S259 in TM6 of Ste2p, are stacked and investigated in terms of their effects on classical G-protein-mediated signal transduction associated with cell cycle arrest, and alternatively, their impact on downstream mating projection and zygote formation events. In relative dose response experiments, accounting for systemic and observational bias, mutational-derived functional differences were observed, validating the S254L-derived bias for downstream mating responses and highlighting complex relationships between TM6-mutation derived constitutive signalling and ligand-induced functionalities. Mechanistically, localization studies suggest that alterations to receptor trafficking may contribute to mutational bias, in addition to expected receptor conformational stabilization effects. Overall, these results extend previous observations and quantify the contributions of Ste2p variants to mediating cell cycle arrest versus downstream mating functionalities.
authors.affiliation: Department of Biochemistry, University of Saskatchewan, 107 Wiggins Road, Saskatoon, SK S7N 5E5, Canada and.
authors.forename: Pooja
authors.initials: P
authors.lastname: Choudhary
authors.affiliation: Department of Biochemistry, University of Saskatchewan, 107 Wiggins Road, Saskatoon, SK S7N 5E5, Canada and Aquatic and Crop Resources Development, National Research Council of Canada, 110 Gymnasium Place, Saskatoon, SK S7N 0W9, Canada michele.loewen@nrc.ca.
authors.forename: Michele C
authors.initials: MC
authors.lastname: Loewen
mesh_terms: D059447:Cell Cycle Checkpoints / Q000235:genetics / Q000502:physiology; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D021381:Protein Transport; D051964:Receptors, Mating Factor / Q000235:genetics* / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000166:cytology / Q000235:genetics* / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics* / Q000378:metabolism*; D015398:Signal Transduction
chemical_list: D051964:Receptors, Mating Factor; C498031:STE2 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins
keywords: G-protein-coupled receptor; alternate functionalities; pheromone mating; site-directed mutagenesis; yeast