Transcription factor positioning and function during promoter opening

描述（由申请人提供）：在这些生活领域中指导监管演变的约束条件的差异。本研究旨在了解转录因子在RNA聚合酶（RNAP）辅助启动基因表达中的作用。它具体解决了两个转录因子，TFB和TFE，从古细菌到人类的结构和功能保守的位置和功能。了解古细菌转录因子的位置和功能对人类健康的重要性在于这些转录因子的人类同源物具有正确基因表达所需的相同或非常相似的功能，并且改变其功能或相互作用伴侣功能的扰动可能导致基因失调疾病，包括癌症和先天性发育缺陷。这项工作旨在确定转录起始过程背后的结构相互作用。我们期望获得新的洞察力的精细尺度定位和功能的TFB和TFE。特别是，我们将有足够的关于B指定位的信息来创建其在启动动态过程中的功能模型，我们还将能够使用这些模型来测试B指中特定突变的影响。我们还将获得前所未有的洞察TFE多肽在转录复合物的方向，并将能够确定假定的相互作用的表面，也可以诱变和测试，以阐明更详细的功能。从这里提出的实验结果将有助于推动更深入的了解不仅是古菌的机制，而且TFIIB和TFIIE（或更具体地说，其α亚基的N-末端）在转录启动真核RNAP II的作用。我们还预计我们的工作将有助于促进真核类型和细菌启动子开放机制之间的比较。虽然非常不同的转录因子（TFB和TFE与sigma）在这个过程中帮助它们各自的RNAP，但最终，从转录泡，RNA-DNA杂交体和活性位点本身的角度来看，结果，延伸RNA聚合酶在几何上是相同的。当两个系统之间的功能相似性和差异被更好地理解时，我们将更好地理解它们的影响。 在人类发育障碍和癌症中，基因组特定部分的变化会导致基因表达的改变;因此，细胞可能失去与邻居合作的能力，并可能导致出生缺陷，遗传疾病或癌症。对这些变化如何影响基因表达过程的详细理解是不完整的，部分原因是基因表达的过程尚未完全理解。为了解决这个问题，我正在研究一组称为古细菌的生物体中基因表达的详细机制。红细胞具有与人类细胞中的转录系统非常相似的转录系统，尽管复杂性要低得多。因此，它们为理解人类基因表达的基本机制提供了一个相对简单的模型。

DESCRIPTION (provided by applicant): Differences on the constraints guiding the evolution of regulation in these domains of life. This research proposal seeks an understanding of the roles for transcription factors in assisting initiation of gene expression by RNA polymerases (RNAPs). It specifically addresses the position and function of two transcription factors, TFB and TFE, structurally and functionally conserved from archaea to humans. The significance of understanding the position and function of archaeal transcription factors to human health rests in the likelihood that human homologs of these transcription factors have the same or very similar functions that are required for proper gene expression, and that perturbations that alter their function or the function of interacting partners can cause diseases of gene dysregulation, including cancer and congenital developmental defects. This work aims to identify the structural interactions that underlie the process of transcription initiation. We expect to gain novel insight into the fine-scale positioning and function of TFB and TFE. In particular, we will have enough information about the positioning of the B-finger to create models for its function in the dynamic process of initiation, and we will also be able to test the effects of specific mutations in the B-finger using these models. We will also gain unprecedented insight into the orientation of the TFE polypeptide in transcription complexes, and will be able to identify putative interacting surfaces that can also be mutagenized and tested to elucidate function in more detail. Results from the experiments proposed here will help to drive a deeper understanding not just of the archaeal mechanism, but also of the roles for TFIIB and TFIIE (or, more specifically, the N-terminus of its alpha subunit) in transcription initiation by eukaryotic RNAP II. We also anticipate our work will help facilitate a comparison between eukaryotic-type and bacterial promoter opening mechanisms. While very different transcription factors (TFB and TFE versus sigma) assist their respective RNAPs in the process, in the end, the outcome, an elongating RNA polymerase, is geometrically the same from the point of view of the transcription bubble, the RNA-DNA hybrid, and the active site itself. When the functional similarities and differences between the two systems are understood better, we will be poised to better understand the effects of these. PUBLIC HEALTH RELEVANCE In human developmental disorders and cancers, changes in specific parts of the genome cause alterations in gene expression; as a consequence of this, cells may lose the ability to cooperate with their neighbors, and birth defects, genetic disease, or cancer can result. A detailed understanding of how these changes affect the gene expression process is incomplete, in part because the process of gene expression is not completely understood. To address this problem, I am studying the detailed mechanism of gene expression in a group of organisms called the archaea. Archaea have a transcription system that is very similar to the transcription system in human cells, though much less complicated. Therefore, they provide a relatively simple model for understanding the basic mechanisms of human gene expression.