Promoter Regulation in Response to Environmental Stress

描述（由申请人提供）：真核生物将数百种（在某些情况下数千种）蛋白质用于基因表达的调控。然而，我们对所有这些蛋白质如何在组成一个典型基因组的数千个基因中协调它们的行为知之甚少。当细胞对包括环境压力在内的信号事件做出反应时，这种协调是如何改变的，人们对此知之甚少。这里提出的工作使用酿酒酵母作为模型细胞系统进行广泛的调查转录调控蛋白位于整个基因组中，以及当基因组被环境信号重新编程时，如热休克和其他压力，它们移动到哪里。热冲击为单元提供快速且简单的编程事件。该项目的初步研究已经揭示了新的见解基因调控，通过证明许多基因进行部分组装的转录机制在启动子。部分复合物等待信号事件，驱使它们进入完全组装。通过染色质免疫沉淀测定来评估参与转录的各种蛋白质的位置，其中微阵列用于检测全基因组结合事件（所谓的chIP芯片）。将在正常生长条件下和广泛的环境应力下对位置进行评估，特别强调热冲击。结合事件之间的关系将提供新的见解转录复合体的组装和调控。通过从细胞分离的天然转录复合物的全基因组生物化学解剖，将提供额外的机制见解。我们的细胞经常面临极端环境，包括温度，饥饿，辐射和有害化学物质。我们如何处理这种压力取决于我们的转录机制的行动。因此，广泛了解我们的转录机制如何在面对各种压力时工作，对于从生理学角度了解人类健康至关重要。

DESCRIPTION (provided by applicant): Eukaryotes dedicate hundreds (and in some cases thousands) of proteins towards the regulation of gene expression. Yet very little is known about how all of these proteins coordinate their behavior at the many thousands of genes that comprise a typical genome. Little is known about how this coordination changes as cells reprogram their genome in response to signaling events including environmental stress. The work proposed here uses Saccharomyces cerevisiae as a model cellular system to undertake a broad survey of where transcriptional regulatory proteins are located throughout the genome, and where they move to when the genome is reprogrammed by environmental signals, such as heat shock and other stresses. Heat shock provides a rapid and simple programming event for the cell. Preliminary studies on this project have already revealed novel insights into gene regulation by demonstrating that many genes undergo partial assembly of the transcription machinery at promoters. Partial complexes await signaling events that drive them into full assembly. The location of a wide range of proteins involved in transcription will be evaluated by chromatin immunoprecipitation assays in which microarrays are used to detected genome-wide binding events (so called chlP-chip). Location will be assessed under normal growth conditions and under a wide range of environmental stresses, with particular emphasis on heat shock. Relationships among binding events will provide new insights into transcription complex assembly and regulation. Additional mechanistic insight will be provided through genome-wide biochemical dissection of native transcription complexes isolated from cells. Our cells are constantly faced with environmental extremes, involving temperature, starvation, radiation and harmful chemicals. How we deal with this stress depends upon the action of our transcription machinery. Therefore, a broad understanding of how our transcription machinery works in the face of various stresses is essential for a physiological understanding of human health.