K Hirota1, K Tanaka, Y Watanabe, M Yamamoto. 1. Department of Biophysics and Biochemistry, Graduate School of Science, University of Tokyo, Hongo, Tokyo 113-0033, Japan.
Abstract
BACKGROUND: Yeast mating-pheromone receptors facilitate the study of G protein-coupled signal transduction. To date, molecular dissection of the budding yeast alpha-factor receptor has been done extensively, but little analysis has been performed with pheromone receptors of fission yeast, another genetically tractable yeast species. RESULTS: We analysed the fission yeast M-factor receptor Map3p. Truncation of the C-terminal 54 amino acids made Map3p dominant-negative over the wild-type. This form, called Map3-dn9p, was competent in the induction of pheromone-dependent gene expression, although it could not direct proper conjugation. Map3-dn9p failed both to provoke the orientated projection of conjugation tubes and to induce adaptation to the pheromone signal associated with endocytosis of the receptor. Deletion and substitution analyses suggested that the integrity of the C-terminal region, rather than a specific subgroup of amino acid residues therein, was vital for the respective Map3p activities. Ubiquitination of the C-terminus was not absolutely essential for Map3p function. CONCLUSIONS: The C-terminal region of Map3p is dispensable for the pheromone signalling per se, but is pivotal for adaptation and pheromone-induced conjugation tube formation, as is true with the budding yeast alpha-factor receptor. However, the mechanisms which induce adaptation appear to differ between fission and budding yeast concerning the necessity of ubiquitination.
BACKGROUND:Yeast mating-pheromone receptors facilitate the study of G protein-coupled signal transduction. To date, molecular dissection of the budding yeast alpha-factor receptor has been done extensively, but little analysis has been performed with pheromone receptors of fission yeast, another genetically tractable yeast species. RESULTS: We analysed the fission yeast M-factor receptor Map3p. Truncation of the C-terminal 54 amino acids made Map3p dominant-negative over the wild-type. This form, called Map3-dn9p, was competent in the induction of pheromone-dependent gene expression, although it could not direct proper conjugation. Map3-dn9p failed both to provoke the orientated projection of conjugation tubes and to induce adaptation to the pheromone signal associated with endocytosis of the receptor. Deletion and substitution analyses suggested that the integrity of the C-terminal region, rather than a specific subgroup of amino acid residues therein, was vital for the respective Map3p activities. Ubiquitination of the C-terminus was not absolutely essential for Map3p function. CONCLUSIONS: The C-terminal region of Map3p is dispensable for the pheromone signalling per se, but is pivotal for adaptation and pheromone-induced conjugation tube formation, as is true with the budding yeast alpha-factor receptor. However, the mechanisms which induce adaptation appear to differ between fission and budding yeast concerning the necessity of ubiquitination.
Authors: Laura Merlini; Bita Khalili; Felipe O Bendezú; Daniel Hurwitz; Vincent Vincenzetti; Dimitrios Vavylonis; Sophie G Martin Journal: Curr Biol Date: 2016-03-24 Impact factor: 10.834