RNA sequencing of pheromone stimulated and unstimulated MATa and MATα Saccharomyces cerevisiae

Haploid budding yeast has two mating types, defined by the alleles of the MAT locus, MATa and MATα. Mating occurs when two haploid cells of opposite mating types signal to each other using reciprocal pheromones and receptors, polarize and grow towards each other, and eventually fuse to form a single diploid cell. The pheromones and receptors are necessary and sufficient to define a mating type, but other mating type-specific proteins make mating more efficient. We examined the role of these proteins by genetically engineering “transvestite” cells that swap the pheromone, pheromone receptor, and pheromone processing factors of one mating type for another. These cells can mate with each other, but their mating is inefficient. By characterizing their mating defects and examining their transcriptomes, we found Afb1 (a-factor barrier), a novel MATα-specific protein that interferes with a-factor, the pheromone secreted by MATa cells. We show that strong pheromone secretion is essential for efficient mating and that the weak mating of transvestites can be improved by boosting their pheromone production. Using synthetic biology, it is possible to characterize the factors that control efficiency in biological processes. In the case of budding yeast mating, selection for increased mating efficiency is likely to have continually boosted pheromone levels and the ability to discriminate between partners who make more (potentially fitter) and less (potentially less fit) pheromones. This sensitivity to which partner makes more pheromone comes at a cost: it means mating is not robust in situations where all potential partners make less pheromone. 4 conditions were analysed, each with 3 biological replicates. The conditions were unstimulated MATa cells in YPD. Stimulated MATa cells in YPD+10nM α-factor. Unstimulated MATα cells in YPD. Stimulated MATα cells in YPD+10nM α-factor.单倍体出芽酵母存在两种交配型，由MAT位点的等位基因MATa与MATα决定。当两种相反交配型的单倍体细胞通过相互对应的信息素与受体进行信号交流、彼此极化并向对方生长，最终融合形成单个二倍体细胞时，便发生交配。信息素与受体是定义交配型的必要且充分条件，但其他交配型特异性蛋白可提升交配效率。本研究通过基因工程构建了"跨交配型"细胞——将某一交配型的信息素、信息素受体及信息素加工因子替换为另一交配型的对应元件，以此探究这类蛋白的功能。这类细胞可进行相互交配，但交配效率低下。通过表征其交配缺陷并分析转录组，我们发现了Afb1（a-factor barrier，a因子屏障）——一种全新的MATα特异性蛋白，可干扰MATa细胞分泌的信息素a因子。我们证实，强信息素分泌是高效交配的必要条件，且可通过提升这类跨交配型细胞的信息素产量来改善其低效交配表型。借助合成生物学技术，可解析调控生物学过程效率的关键因子。以出芽酵母交配为例，对更高交配效率的选择压力可能持续提升了信息素分泌水平，以及区分"分泌更多信息素（潜在适应度更优）"与"分泌更少信息素（潜在适应度更差）"配偶的能力。但这种对信息素分泌量的敏感性存在代价：当所有潜在配偶的信息素分泌量均较低时，交配过程将无法稳定进行。本研究共分析了4组实验条件，每组均设置3次生物学重复。4组条件分别为：YPD培养基中未受刺激的MATa细胞、YPD+10nM α-因子培养基中受刺激的MATa细胞、YPD培养基中未受刺激的MATα细胞，以及YPD+10nM α-因子培养基中受刺激的MATα细胞。