Control of Transcription in Eukaryotic Cells

描述(申请人提供)：越来越明显的是，RNA聚合酶II从起始到转录延长的转变是真核基因表达的一个主要调节检查点。在目前的支持期内，我们的实验室在表征这一转变方面取得了重大进展。我们设计了一个启动子清除的工作模型，这是这一过程的第一步。我们的清除模型包含了新的概念，即转录起始因子的重要作用。我们还证明了从转录起始下游的25-35个碱基开始发生明显的清除后转变。我们对RNA聚合酶II通过清除后阶段的能力的分析表明，在转录的这一点上，聚合酶可以被捕获的机制有了新的见解。在目前的应用中，我们提出了四个具体的目标，通过这些目标，我们将(I)测试我们最初的清除和清除后转换模型，(Ii)确定单个转录因子在这些转换中的作用，以及(Iii)识别对这些转换具有重要影响的潜在DNA序列。与公共健康相关：许多基因的正确表达对人类健康至关重要，它们是通过在基因信息转录启动后延长转录机制的暂停来控制的。目前还不知道这些“受监管的闸门”是如何运作的。为了最终操纵这些门以造福人类健康，揭示其中的分子机制是至关重要的。这里提出的研究旨在提供这方面的知识。我们研究的清除和清除后转换是细胞中观察到的调节点的试管模型。

DESCRIPTION (provided by applicant): It is increasingly evident that the transition from initiation to transcript elongation by RNA polymerase II is a major regulatory checkpoint in eukaryotic gene expression. During the current support period our laboratory has made significant advances in characterizing this transition. We have devised a working model for promoter clearance, the initial step in this process. Our clearance model includes the novel concept of an important role for transcript initiation factors. We have also demonstrated that a distinct postclearance transition occurs from 25-35 bases downstream of transcription start. Our analysis of the ability of RNA polymerase II to pass through the postclearance stage has suggested new insights into mechanisms through which polymerase can be trapped at this point in transcription. In the current application, we propose four specific aims, through which we will (i) test our initial models for the clearance and postclearance transitions (ii) determine the roles of individual transcription factors in these transitions, and (iii) identify underlying DNA sequences, which have important effects on these transitions. Relevance to Public Health: Many genes whose correct expression is essential for human health are controlled through the extended pausing of the transcriptional machinery at a point just after the transcription of genetic information has initiated. It is not currently known how these "regulated gates" operate. In order to ultimately manipulate these gates for the benefit of human health, it is essential to uncover the molecular mechanisms involved. The research proposed here is aimed at providing this knowledge. The clearance and postclearance transitions which we study are test tube models for the regulatory points observed in the cell.