Characterization of structurally atypical sigma factors carrying fused domains of unknown function in the plant endosymbiont, Sinorhizobium meliloti

转录是所有生物基因表达的第一个主要步骤。在细菌中，所有的基因表达都是由一个被称为σ (σ)因子的一般转录因子家族调节的。每个σ因子控制一个基因子集（其调控子）的表达，根据σ因子的不同，调控子可能由一个到数千个基因组成。这些蛋白的一个重要组是由胞质外功能(ECF) σ因子组成的，其中许多σ因子的活性受到包括拮抗σ因子蛋白在内的信号转导途径的负调控。ECF σ因子及其相关调节因子构成了一个主要的感觉系统，与特定的转录因子和双组分调节系统一起，介导细胞对细胞内外变化的物理化学条件进行适当的基因表达反应的能力。******在机制上，σ因子执行两个主要功能：它们将RNAP全酶靶向到基因启动子上，并将转录起始位点捕获在称为开放复合物的单链状态。开放复合物的稳定是转录起始途径的重要步骤，σ因子的活性部分类似于古细菌和真核生物RNAP II中TFIIB的活性。了解σ因子函数的机理细节是一个非常活跃的研究领域，它涉及到一些特征良好的模型系统。然而，基因组序列数据的分析表明，在许多细菌中存在结构异常的ECF σ因子相关蛋白，这些非典型或非典型系统仍未得到充分研究。ECF σ因子的几个亚群意外地携带未知或不完全表征功能的融合蛋白结构域。新出现的证据指向这些融合域的调控功能，我的研究计划的长期目标是研究细菌中非规范转录系统的机制细节。对非规范系统的研究有望揭示转录起始的新途径，以及基因表达调控与其他细胞过程之间意想不到的生理关系。*** ***值得注意的是，重要的内共生细菌Sinorhizobium meliloti合成了两个独特的非规范融合域σ因子的例子。本文描述了其中一种蛋白质（RpoE9）对S. meliloti生物学的生理重要性的研究。为此，我们提出了一系列的研究目标，主要集中在识别RpoE9活性与其他细胞调节和稳态系统之间的意外交集。我们希望强调的是，虽然对这种结构非典型σ因子的研究本身很重要，但我们也将其视为从更全局和系统范围的角度建立和完善所有ECF σ因子研究的工具。＊＊＊＊＊

Transcription is the first major step in gene expression in all organisms. In bacteria, all gene expression is regulated by a family of general transcription factors called sigma (σ) factors. Each σ factor controls the expression of a subset (its regulon) of genes and regulons may consist of between one and thousands of genes depending on the σ factor. An important group of these proteins is comprised of Extracytoplasmic Function (ECF) σ factors and the activity of many of these is negatively regulated by a signal transduction pathway that includes an antagonistic anti-σ factor protein. ECF σ factors and their associated regulators constitute a major sensory system that, along with specific transcription factors and two-component regulatory systems, mediate the cellular ability to mount appropriate gene expression responses to changing physicochemical conditions inside and outside of the cell. ******Mechanistically, σ factors carry out two major functions: they target RNAP holoenzyme to the gene promoter and they trap the transcription start site in a single-stranded state called an open-complex. Stabilization of the open-complex is an essential step in the transcription initiation pathway and σ factor activity is partially analogous to the activity of TFIIB in archaeal and eukaryotic RNAP II. Understanding mechanistic detail of σ factor function is a very active area of research that involves a few well-characterized model systems. However, the analysis of genome sequence data has revealed the presence of structurally unusual ECF σ factor-related proteins in many bacteria and these atypical or non-canonical systems remain understudied. Several sub-groups of ECF σ factors unexpectedly carry fused protein domains of unknown or incompletely characterized function. Emerging evidence points to regulatory functions for these fused domains and the long term objective of my research program is to study mechanistic details of non-canonical transcription systems in bacteria. The study of non-canonical systems carries the promise of uncovering novel pathways to transcription initiation and unexpected physiological relationships between the regulation of gene expression and other cellular processes.*** ***Remarkably, the important endosymbiotic bacterium Sinorhizobium meliloti synthesizes two unique examples of non-canonical fused domain σ factors. This current proposal describes the study of the physiological importance of one of these proteins (RpoE9) to S. meliloti biology. To these ends, we are proposing a series of research pursuits mainly focused on discerning unexpected intersections between RpoE9 activity and other cellular regulatory and homeostatic systems. We wish to emphasize that while the study of this structurally atypical σ factor is important in itself, we also view it as a vehicle to establish and refine the study of all ECF σ factors from a more global and systems-wide perspective. *****