N‐terminal residues of the yeast pheromone receptor, Ste2p, mediate mating events independently of G1‐arrest signaling

Shi, Chunhua; Kendall, Stephanie; Grote, Eric; Kaminskyj, Susan G. W.; Loewen, Michèle C.

doi:10.1002/jcb.22129

Cited by 17 publications

(21 citation statements)

References 35 publications

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“…Haploid cells locate mating partners by polarizing their growth in the direction of the highest pheromone concentration [11,12]. Mutations in either pheromones or receptors can alter mating efficiency [13][14][15], and pheromone-receptor specificity has been proposed as a possible mechanism for mate discrimination between those species whose pheromone peptide sequences differ [3,16]. Attraction to pheromones produced by incompatible partners is particularly costly for yeast because each cell can mate only once; zygote inviability or sterility is equivalent to death for the mating haploids.…”

Section: Resultsmentioning

confidence: 99%

Experimental Evolution of Species Recognition

et al. 2015

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Sex with another species can be disastrous, especially for organisms that mate only once, like yeast. Courtship signals, including pheromones, often differ between species and can provide a basis for distinguishing between reproductively compatible and incompatible partners. Remarkably, we show that the baker's yeast Saccharomyces cerevisiae does not reject mates engineered to produce pheromones from highly diverged species, including species that have been reproductively isolated for up to 100 million years. To determine whether effective discrimination against mates producing pheromones from other species is possible, we experimentally evolved pheromone receptors under conditions that imposed high fitness costs on mating with cells producing diverged pheromones. Evolved receptors allowed both efficient mating with cells producing the S. cerevisiae pheromone and near-perfect discrimination against cells producing diverged pheromones. Sequencing evolved receptors revealed that each contained multiple mutations that altered the amino acid sequence. By isolating individual mutations, we identified specific amino acid changes that dramatically improved discrimination. However, the improved discrimination conferred by these individual mutations came at the cost of reduced mating efficiency with cells producing the S. cerevisiae pheromone, resulting in low fitness. This tradeoff could be overcome by simultaneous introduction of separate mutations that improved mating efficiency alongside those that improved discrimination. Thus, if mutations occur sequentially, the shape of the fitness landscape may prevent evolution of the optimal phenotype--offering a possible explanation for the poor discrimination of receptors found in nature.

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Section: Resultsmentioning

confidence: 99%

Experimental Evolution of Species Recognition

et al. 2015

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“…For example, the Ste2p mutantsY266C, Q149R and W295C exhibited robust FUS1-lacZ activity, but little to no growth arrest activity [43] [65] [66]. Similarly, a divergence in correlation between mating efficiency and FUS1-lacZ induction has been reported for I24C and I29 mutants which exhibited approximately 50% and 100% of the WT FUS1-lacZ activities, respectively, but only <0.09% and 4.6% of the WT mating efficiency[29]. Mutations of receptors resulting in biased activation of specific signaling pathways has been described for other GPCRs [67–70].…”

Section: Discussionmentioning

confidence: 99%

“…The N-terminus of Ste2p is ~48 amino acids long, and it harbors two glycosylation sites (N25 and N32) which were eliminated by mutation (N25A and N32A) without affecting receptor function [25]. Other studies indicated that the N-terminus contributed to receptor dimerization [26, 27], and three residues (Pro 15, Ile24, and Ile29) were found to be essential for mating but not for signaling as measured by growth arrest and reporter gene ( FUS1 ) activation assays [28, 29]. Truncation of parts of the N-terminus implicated this domain in cellular fusion (mating) during late stages of conjugation of opposite mating types [29].…”

Section: Introductionmentioning

confidence: 99%

The N-terminus of the yeast G protein-coupled receptor Ste2p plays critical roles in surface expression, signaling, and negative regulation

Uddin

Hauser

Naider

et al. 2016

Biochimica et Biophysica Acta (BBA) - Biomembranes

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G protein-coupled receptors (GPCRs) are found in all eukaryotic cells examined to date where they function as membrane-bound proteins that bind a multitude of extracellular ligands to initiate intracellular signal transduction systems controlling cellular physiology. GPCRs have seven heptahelical membrane spanning domains connected by extracellular and intracellular loops with an extracellular N-terminus and an intracellular C-terminus. The N-terminus has been the least studied domain of most GPCRs. The yeast Ste2p protein, the receptor for the thirteen amino acid peptide pheromone α-factor, has been used extensively as a model to study GPCR structure and function. In this study we constructed a number of deletions of the Ste2p N-terminus and uncovered an unexpected function as a negative regulatory domain. We examined the role of the N-terminus in expression, signaling function and ligand-binding properties and found that the residues 11-30 play a critical role in receptor expression on the cell surface. The studies also indicated that residues 2-10 of the N-terminus are involved in negative regulation of signaling as shown by the observation that deletion of these residues enhanced mating and gene induction. Furthermore, our results indicated that the residues 21-30 are essential for optimal signaling. Overall, we propose that the N-terminus of Ste2p plays multiple regulatory roles in controlling receptor function.

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“…Towards validating reported alternate functionalities of Ste2p (10,12,14,16,18) according to current standards (6), a relative mutational-derived bias method was applied. This method accounts for systemic and observational bias that makes direct comparison of data between assays otherwise impossible.…”

Section: Discussionmentioning

confidence: 99%

“…Some examples of these include mutations I24C and I29C in the N-terminus, yielding a phenotype that has strong G-protein-dependent MAPK signalling and cell cycle arrest activity, but no actual mating functionality (18). Similarly, mutation S251L, in the central region of transmembrane (TM) 6, also yielded normal signalling, but a weak-mating phenotype (19) (Fig.…”

mentioning

confidence: 99%

Quantification of mutation-derived bias for alternate mating functionalities of theSaccharomyces cerevisiaeSte2p pheromone receptor

Choudhary

Loewen

2015

J Biochem

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Although well documented for mammalian G-proteincoupled 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 ligandinduced 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.

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N‐terminal residues of the yeast pheromone receptor, Ste2p, mediate mating events independently of G1‐arrest signaling

Cited by 17 publications

References 35 publications

Experimental Evolution of Species Recognition

Experimental Evolution of Species Recognition

The N-terminus of the yeast G protein-coupled receptor Ste2p plays critical roles in surface expression, signaling, and negative regulation

Quantification of mutation-derived bias for alternate mating functionalities of theSaccharomyces cerevisiaeSte2p pheromone receptor

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