Mating Type Silencing and Switching in Yeast

描述(由申请人提供)：我们建议在三个不同但相互关联的领域研究酵母的交配类型。首先，我们计划使用一种新的交配型基因座沉默的视觉方法来探索异染色质沉积和维持的机制，以确定活跃状态和抑制状态之间的相互转换动力学，并评估影响这些转化率的因素。这些结果应该有助于阐明影响X失活和染色体印记等不同事件的调控过程，这些事件是各种疾病(如散发性癌症)的基础。其次，交配型相互转换过程中的供体选择需要酵母基因组的远距离区域之间受发育调节的选择性相互作用。我们计划通过完成对选择过程中改变的突变的基因组筛选，并通过检查活细胞中交配型相互转换的动力学来测试这一重组过程的遗传基础，来探索这一过程的机制基础。这些观察应该对大型真核生物中类似的发育事件有深入的了解，例如免疫球蛋白类别转换和顺序的丙珠蛋白激活。最后，酵母细胞通过启动转录和发育程序对信息素做出反应，最终达到交配。我们最近发现，种群中的单个细胞以两种不同的模式之一对信息素做出反应：一些细胞根本没有反应，而其余的细胞则表现出完全的交配反应。这种分叉的反应部分归因于不同MAP激酶信号通路之间的抑制串扰。我们建议扩展这些研究，以探索负责信息素反应的MAP激酶通路与假菌丝和侵袭性生长之间的串扰。这些研究的结果应该有助于阐明细胞信息处理中信号整合的机制。所有这些研究，在探索酵母细胞类型和交配反应的不同方面时，都被共同的技术和理论方法统一起来，即检查单个细胞，并从它们不同的个体行为中提取生物学和机械原理。

DESCRIPTION (provided by applicant): We propose to investigate mating type in yeast in three distinct but interrelated areas. First, we plan to explore the mechanism by which heterochromatin is deposited and maintained using a novel visual assay for silencing at the mating type loci to determine the dynamics of interconversion between the active and repressed states and to assess the factors that influences those rates of conversion. These results should shed light on the regulatory processes affecting such diverse events as X-inactivation and chromosome imprinting, which underlies various diseases such as sporadic cancers. Second, donor selection during mating type interconversion requires developmentally-regulated, selective interaction between distant regions of the yeast genome. We plan to explore the mechanistic basis of this process by completing a genome-based screen for mutants altered in the selection process and by examining the dynamics of mating type interconversion in living cells to test the genetic underpinnings of this recombination process. These observations should yield insights into similar developmental events in larger eukaryotes, such as immunoglobulin class switching and sequential Pglobin activation. Finally, yeast cells respond to pheromone by initiating a transcriptional and developmental program culminating in mating. We recently found that individual cells within the population respond to pheromone in one of two distinct patterns: some cells show no response at all while the remaining cells exhibiting a full mating response. This bifurcated response is due in part to inhibitory cross talk among different MAP kinase signaling pathways. We propose to extend these studies to explore the crosstalk between the MAP kinase pathways responsible for pheromone response and for pseudohyphal and invasive growth. The results of these studies should shed light on the mechanism for signal integration in information processing in cells. All these studies, while probing different aspects of cell type and mating response in yeast, are unified by the common technical and theoretical approaches of examining individual cells and extracting biological and mechanistic principles from their distinct individual behaviors.