RIBOSOMAL GENES OF YEAST AND THEIR REGULATION

描述（由申请人提供）：由于细胞的生长速度取决于蛋白质的合成，而蛋白质的合成又依赖于核糖体，因此对细胞生长的调控最终必须依赖于对核糖体合成的调控，而核糖体的合成消耗了细胞资源的很大一部分。因此，我们对核糖体合成调控的研究是理解生长控制的基础。我们将试图确定137个核糖体蛋白基因（编码78种核糖体蛋白，负责约30%的细胞mrna）如何在酿酒酵母中转录到如此高的程度、如此精确的定量和如此协调的调控。关键的转录因子Rap1负责激活和沉默RP基因，我们的假设是整个上游序列已经进化到满足这些需求，这意味着激活子和启动子元件之间存在相互作用。这些研究将基于在核糖体蛋白基因上发现的Rap1位点与自身或与他人的近端启动子区域的功能相互作用的系统分析，反之亦然，测试在其他基因或端粒中发现的Rap1位点。特别有价值的是，我们可以独立分析对至少三种信号通路的反应，PKA， PKC和TOR，以及热休克和其他影响核糖体合成的应激。我们将解剖Rap1本身，包括确定DNA结合区域的作用，询问特定区域是否可以“抑制”Rap1的功能，并确定Rap1的翻译后修饰可能在这种调节中发挥作用。我们最近的遗传研究结果引起了我们对细胞生理学中一个重要但被忽视的领域的关注，不仅对酵母，而且对所有细胞，错误组装或不完整的核糖体亚基的识别和降解，如果保持完整，可能会阻塞翻译或核糖体组装的途径。我们提出了一个实验，以询问是否有功能校对完成的核糖体。我们将研究蛋白质降解途径和RNA降解途径的几个部分，以确定哪个部分参与了这一重要任务。我们提出了一个突变体筛选来识别关键参与者。

DESCRIPTION (provided by applicant): Since the rate of growth of a cell depends on the synthesis of proteins, and the synthesis of proteins depends on ribosomes, the regulation of growth must ultimately depend on the regulation of the synthesis of ribosomes, which consumes a large fraction of the cell's resources. Thus our study of the regulation of ribosome synthesis is fundamental to the understanding of growth control. We will attempt to determine how 137 ribosomal protein genes, encoding 78 ribosomal proteins and responsible for some 30 percent of the cell's mRNAs, are transcribed to such a high degree, with such quantitative precision, and with such coordinate regulation in S. cerevisiae. The critical transcription factor, Rap1, is responsible both for activating and for silencing the RP genes, Our hypothesis is that the entire upstream sequence has evolved to meet these demands, implying interaction between activator and promoter elements. These studies will be based on a systematic analysis of the functional interaction of Rap1 sites found on ribosomal protein genes with their own or with others' proximal promoter regions, and vice versa, testing the Rap1 sites found in other genes or in the telomeres. Of particular value is that we can independently assay the response to at least three signaling pathways, PKA, PKC, and TOR, as well as heat shock and other stresses that affect ribosome synthesis. We will dissect Rap1, itself, including determining the role of the DNA binding region, asking whether specific regions can 'squelch' Rap1 function, and determining the post-translational modifications of Rap1 that might play a role in this regulation. A new project arises from our recent genetic results that have drawn our attention to an important but neglected area of cell physiology, not only for S. cerevisiae, but for all cells, the identification and degradation of mal-assembled or incomplete ribosomal subunits, that, if left intact, could clog the pathways for translation or ribosome assembly. We propose an experiment to ask whether there is functional proofreading of completed ribosomes. We will examine the several parts of both the protein degradation pathway and the RNA degradation pathway to determine which is involved in this important task. We propose a mutant screen to identify key players.