REGULATION OF FILAMENTOUS GROWTH IN YEAST

这项建议的目的是了解提供 对收敛的MAP激酶通路的特异性。信息素反应 酿酒酵母中负责传播的途径 在单倍体接合过程中，胞外交配信号进入细胞核。 它包括信息素受体，一种异三聚体G蛋白，一种高度 保守的PAK/Mekk/MEK/MAPK激酶级联(Ste20p、Ste11p、Ste7p和 Fus3p/Kss1p)和转录因子Ste12p。中的元素 还需要路径，包括Ste20p、Ste11p、Ste7p和Ste12p 用于二倍体丝状生长(细长细胞链的发育 氮素饥饿时)和单倍体入侵生长。因此，一个激酶 在单倍体和单倍体中，不同的发育途径使用级联 二倍体和途径的输出涉及相同的转录 因数，STE12P。这项拟议的研究使用了分子和基因 确定Ste12p调节器和目标的方法，并了解 STE_(12)P在丝状细胞生长中的调控作用及其比较 在单倍体交配中。第一个具体目标是识别和描述 Ste12p调控丝状二倍体细胞中的基因。基因 丝状细胞生长过程中Ste12p的转录调控 二倍体将用酵母LacZ融合文库克隆。他们的 表达模式、蛋白质定位和突变表型将是 分析过了。一些克隆基因的启动子将被定位以定义 STE 12P诱导的上游激活序列 抄写。第二个目的是了解STE12P的机制 对丝状菌生长的监管，并确定其监管机构。我们建议 研究丝状化条件下Ste12p的磷酸化。遗传 筛选的目的是识别二倍体中的Ste12p激酶，因为 信息素反应途径的MAP激酶Fus3p/Kss1不是必需的 用于丝状生长。我们建议增加监管机构的职能 与Stel2p一起提供了二倍体的特异性调控。两者都有 基因和生化实验将被用来鉴定这些 监管者。保守的MAP激酶对Ste12p调控的研究 路径解决了一个有趣的问题，涉及到对 在这些途径中具有特异性。这项研究将加强我们的 对MAP激酶通路复杂调控机制的认识 哺乳动物细胞。基因二态调控的研究进展 易驯化的酿酒酵母也将加深我们对真菌的了解 二形性在白色念珠菌的发病机制中很重要， 人类常见的真菌病原体。

The aim of this proposal is to understand mechanisms that provide the specificity to converging MAP kinase pathways. The pheromone response pathway in Saccharomyces cerevisiae is responsible for transmitting extracellular mating signals into the nucleus during haploid conjugation. It includes pheromone receptors, a hetero-trimeric G protein, a highly conserved PAK/MEKK/MEK/MAPK kinase cascade (Ste20p, Ste11p, Ste7p, and Fus3p/Kss1p), and a transcription factor, Ste12p. Elements of the pathway, including Ste20p, Ste11p, Ste7p, and Ste12p, are also required for diploid filamentous growth (development of chains of elongated cells upon nitrogen starvation) and haploid invasive growth. Thus, one kinase cascade is used by different developmental pathways in haploids and diploids and the outputs of the pathways involve the same transcription factor, Ste12p. The proposed research uses molecular and genetic approaches to identify Ste12p regulators and targets, and to understand Ste 12p regulation in filamentous growth in comparison to its regulation in haploid mating. The first specific aim is to identify and characterize Ste12p regulated genes in filamentous diploid cells. Genes transcriptionally regulated by Ste12p during filamentous growth in diploids will be cloned with a yeast lacZ fusion library. Their expression pattern, protein localization and mutant phenotypes will be analyzed. Promoters of some of the cloned genes will be mapped to define the upstream activation sequences responsible for Ste 12p induced transcription. The second aim is to understand the mechanism of Ste12p regulation in filamentous growth and identify its regulators. We propose to study Ste12p phosphorylation under filamentation conditions. Genetic screens are designed to identify a Ste12p kinase in diploids because the MAP kinases Fus3p/Kss1 of the pheromone response pathway are not required for filamentous growth. We propose additional regulators functioning together with Stel2p provide the diploid specific regulation. Both genetic and biochemical experiments will be utilized to identify these regulators. Our research on Ste12p regulation by the conserved MAP kinase pathway addresses an interesting question concerning the regulation of specificity among these pathways. This study will enhance our understanding of the complicated regulation by MAP kinase pathways in mammalian cells. Investigation of dimorphic regulation in genetically tractable S. cerevisiae will also further our knowledge of fungal dimorphism, which is important in the pathogenesis of Candida albicans, a common fungal pathogen of humans.