The KlSTE2 and KlSTE3 genes encode MATα‐ and MATa‐specific G‐protein‐coupled receptors, respectively, which are required for mating of Kluyveromyces lactis haploid cells

Abstract

Mating in yeast is initiated by binding of pheromone to G-protein-coupled receptors expressed in haploid cells. We analysed the role of KlSte2p and KlSte3p receptors in the Kluyveromyces lactis mating pathway. By sequence analysis, KlSte2p and KlSte3p are the homologues of the Saccharomyces cerevisiae α-pheromone and a-pheromone receptors, respectively. However, by expression experiments, we determined that KlSTE2 gene is expressed in the cells typified as MAT α and therefore is the receptor for the K. lactis a-pheromone and KlSTE3 gene is expressed in the MAT a cells and binds the α-pheromone. The KlSTE2 gene is silent in MAT a cells, while it is highly expressed in MATα cells, and conversely the KlSTE3 gene is expressed in MAT a cells and repressed in MATα cells. Disruption mutants of both genes were found to be sterile, and this defect is reversed by plasmidic copies of each gene. The cytoplasmic C-terminus of KlSte3p interacts strongly with the KlGpa1p (Gα) subunit, which is involved in the transduction of the pheromone stimulus to induce mating. Remarkably, this same domain does not interact with a constitutive active allele of the Gα subunit, indicating that the C-terminus is able to discriminate between the active (GTP-bound) and inactive (GDP-bound) forms of the Gα subunit.

Kluyveromyces lactissexual pheromones. Gene structures and cellular responses to α-factor

“…cerevisiae , each pheromone is encoded by two different genes (Kurjan & Herskowitz, 1982; Singh et al , 1983; Michaelis & Herskowitz, 1988). All of the above observations are in agreement with the fact that K. lactis MAT α cells express the KlSte2p receptor, whereas the MAT a cells express the KlSte3p receptor (Torres‐Quiroz et al , 2007).…”

“…According to this model, the nature of the putative interactions involved in receptor and pheromone recognition are conserved in K. lactis . First, the W1 and W3 present in both pheromones may contact the conserved aromatic group of the Y residue present in the interface of the sixth transmembrane region (TM6) of both receptors (Torres‐Quiroz et al , 2007). Second, Q10 of both factors may contact the receptor's polar residues present on the outside surface of the first transmembrane segment (S and T in Sa.…”

“…cerevisiae α‐factor might contact the aromatic F and Y residues located at the extracellular side of the fifth and sixth ScSte2p transmembrane regions (TM5 and TM6), respectively (Naider & Becker, 2004). Interestingly, position 13 of the K. lactis pheromone is F, and its putative receptor‐interacting residues are Y in TM5 and Y in TM6 (Torres‐Quiroz et al , 2007). Regardless of the moderate conservation of the primary sequence of α‐factors (50%), some of the nonconserved residues could play important roles in species‐specific interactions with cognate receptors, and should contribute to the sterility barrier between these two yeasts.…”

“…The Gα (Saviñón‐Tejeda et al , 2001) and the Gβ (Kawasaki et al , 2005) subunits of the heterotrimeric G‐protein trigger the pheromone signal to downstream effectors. Sex‐specific G‐Protein‐coupled (GPC) receptors located at the plasma membrane are essential for this process (Torres‐Quiroz et al , 2007). Both G‐protein subunits function in the mating pathway via the KlSte12p transcription factor.…”

Protein Kinases Involved in Mating and Osmotic Stress in the Yeast Kluyveromyces lactis

“…Our preliminary studies indicate that both KlSte50p and KlSte20p interact with KlSte11p, thus activating the MAPK cascade that elicits mating in K. lactis. The actual picture of the pheromone response pathway in K. lactis points to a model where activation of G protein by binding of pheromone to GPCR (32) triggers two branches that converge in the MAPKKK KlSte11p; one is essential for mating and is formed by G␤-KlSte20p, and the second is dispensable and is formed by G␣-KlSte50p. Additional support for this model is given by the disruption phenotypes of mutants that inactivate the G protein subunits.…”