The Role Of Alternate Sigma Factors In The Transmission

sigma是sigma70家族的一员，负责大肠杆菌中多种基因的转录，这些基因在细胞进入固定期或在某些类型的饥饿和应激（如酸胁迫或高渗胁迫）期间表达。沙门氏菌质粒毒力基因spvR和spvABCD在包括鼠伤寒沙门氏菌、霍乱沙门氏菌和都柏林沙门氏菌在内的多种沙门氏菌的大（50至100 kb）质粒上携带。这些质粒（SpvR、SpvC和SpvD或sigma）的缺失会导致小鼠模型中毒力的丧失和网状内皮系统中细胞繁殖能力的丧失。这些基因的表达随着细胞进入站位期而急剧增加，并依赖于sigma。目前的模型提出，在生长期、应激或饥饿条件下，sigmas的积累会增加spvR的转录，然后sigmas和spvR共同作用，增加spv操纵子的转录。虽然额外的调节因子通常可以调节特定的sigmaS依赖基因的表达，但这些基因的表达主要是由于细胞中sigmaS水平的增加。在大肠杆菌中，sigmaS水平通过多种方式控制，包括转录起始和翻译延伸。然而，影响sigma水平的一些主要因素是导致蛋白质稳定性增加的翻译后事件。在37℃的指数生长过程中，sigma的半衰期小于2分钟。当细胞进入固定期或遇到一定的应激条件时，sigma的半衰期增加到大于30min。ClpXP蛋白酶是一种依赖atp的细胞质蛋白酶，在指数生长过程中负责sigma的快速降解。ClpXP对sigma的快速降解需要RssB（也称为SprE），这是一种与反应调节因子家族具有同源性的蛋白质，似乎可以特异性地调节sigma的活性及其降解。热休克蛋白DnaK也被证明在sigma的翻译后控制中具有积极作用。DnaK似乎参与了至少两种信号的转导，即热休克和碳饥饿，这两种信号导致sigma周转率降低。在细菌中，sigmaS-RNA聚合酶全酶在从指数生长过渡到固定阶段或对某些类型的应激作出反应后负责基因表达。伯氏疏螺旋体含有rpoS（编码sigmaS），我们的初步数据表明，rpoS的表达随着培养进入固定期而增加。我们已经将伯氏疏螺旋体rpoS的转录起始位点定位到潜在的sigma54依赖性启动子。此外，对burgdorferi sigma54突变体rpoS的转录分析表明，当细胞进入固定生长阶段时，sigma54会调控sigmaS。因此，sigma54-全酶是sigmaS表达所必需的，这两种蛋白在蜱虫中肠和哺乳动物宿主中伯氏疏螺旋体的生存中起着关键作用。(3) sigma54对基因表达的调控。如上所述，我们已经确定了一个潜在的依赖于sigma54的启动子，它参与了rpoS的表达。由rpoN基因编码的Sigma54，首次被证明是肠道细菌中参与氮代谢的基因表达所必需的。它已被证明是多种细菌转录基因所必需的，这些基因的产物涉及多种功能，如氢代谢、c4 -二羧酸运输、毛蛋白和鞭毛生物合成以及芳香族化合物的降解。与其他可选的0因子不同，sigma54与sigma70家族不具有同源性。此外，sigma54-RNA聚合酶全酶（sigma54- holo酶）启动转录的机制与其他形式的RNA聚合酶全酶不同。sigma54-Holoenzyme可以识别在？12和？24个区，一致序列为5?-TGGCACN4TTTGC(A/T)-3？保守的GG和GC双偶体之间的间距（在一致序列中下划线）是至关重要的，因为间距的任何变化都会导致sigma54-全酶无法识别启动子。Sigma54-Holoenzyme与启动子结合形成封闭的启动子复合物，但在缺乏激活蛋白的情况下无法启动转录。激活子通常与转录起始位点上游100- 200bp的位点结合，并通过DNA环与sigma54-全酶进行短暂接触。激活剂和sigma54-全酶之间的有效相互作用导致封闭启动子复合物异构化为具有转录能力的开放复合物。为了催化这种异构化反应，活化剂必须水解ATP。激活剂结合ATP水解打开复合物形成的机制尚不清楚，但它似乎不涉及激活剂或sigma54全酶的磷酸化。sigma54全酶活化剂的活性受环境信号的调控。许多sigma54全酶的激活因子是双组分调节系统中的反应调节因子，这些蛋白的磷酸化导致它们的激活。这些反应调节因子被其同源蛋白组氨酸激酶磷酸化以响应环境信号。一旦磷酸化，反应调节因子就会激活其他基因的转录。伯氏疏螺旋体sigma54全酶的激活剂也是双组分系统的响应调节剂。编码激活子的基因与编码其同源蛋白组氨酸激酶的基因在一个操纵子中。我们的初步数据表明，激活剂控制rpoS的表达。因此，我们将这种激活剂称为sigmaS调节剂（SisR），将其同源蛋白组氨酸激酶称为sigmaS调节蛋白组氨酸激酶（SisK）。

Regulation of gene expression by sigmaS, a member of the sigma70 family, is responsible for the transcription of a variety of genes in E. coli that are expressed as the cells enter stationary phase or during certain types of starvation and stress, such as acid stress or hyperosmotic stress. A role for sigmaS in the regulation of virulence factors has also been established in Salmonella sp. Salmonella plasmid virulence genes, spvR and spvABCD are carried on large (50 to 100-kb) plasmids in a variety of Salmonella species including S. typhimurium, S. choleraesuis, and S. dublin. Loss of these plasmids, of SpvR, SpvC and SpvD, or ofsigmaS, results in loss of virulence in mouse models and the ability of the cells to multiply in the reticuloendothelial system. Expression of the genes increases dramatically as cells enter station phase and is dependent onsigmaS. The current model proposes that accumulation ofsigmaS in response to growth phase, stress, or starvation conditions increases transcription of spvR, andsigmaS and SpvR then act together to increase transcription of the spv operon. Although additional regulatory factors often function to modulate the expression of specific sigmaS-dependent genes, the expression of these genes results largely from increased levels of sigmaS in the cell. In E. coli, sigmaS levels are controlled in a variety of ways, including transcription initiation and translational elongation. Some of the major factors that influence sigmaS levels, however, are post-translational events that lead to increased stability of the protein. During exponential growth at 37oC,sigmaS has a half-life of less than 2 min. As the cells enter stationary phase or encounter certain stress conditions, the half-life of sigmaS increases to greater than 30 min. The ClpXP protease is a cytoplasmic, ATP-dependent protease that is responsible for the rapid degradation of sigmaS during exponential growth. This rapid degradation of sigmaS by ClpXP requires RssB (also called SprE), a protein that shares homology with the family of response regulators and appears to specifically modulate the activity of sigmaS as well as its degradation. The heat shock protein DnaK has also been shown to have a positive role in the post-translational control of sigmaS. DnaK appears to be involved in the transduction of at least two signals, heat shock and carbon starvation, that result in reduced sigmaS turnover. In bacteria, sigmaS-RNA polymerase holoenzyme is responsible for gene expression following the transition from exponential growth to stationary phase or in response to certain types of stress. B. burgdorferi contains rpoS (encoding sigmaS) and our preliminary data indicate that expression of rpoS increases as cultures enter stationary phase. We have mapped a transcriptional start site of rpoS from B. burgdorferi to a potential sigma54-dependent promoter. Furthermore, analysis of the transcription of rpoS in a B. burgdorferi sigma54 mutant indicates that sigmaS is regulated by sigma54 as cells enter stationary phase of growth. Thus sigma54-holoenzyme is required for sigmaS expression and both proteins play key roles in the survival of B. burgdorferi in the tick midgut and in survival in mammalian hosts. (3) Regulation of gene expression by sigma54. As indicated above, we have identified a potential sigma54-dependent promoter that is involved in expression of rpoS. Sigma54, encoded by the rpoN gene, as first shown to be required for the expression of genes involved in nitrogen metabolism in enteric bacteria. It has since been shown to be required in various bacteria for the transcription of genes whose products are involved in such diverse functions as hydrogen metabolism, C4-dicarboxylic acid transport, pilin and flagellar biosynthesis, and degradation of aromatic compounds. Unlike other alternative o factors, sigma54 does not share homology with the sigma70 family. Moreover, the mechanism by which sigma54-RNA polymerase holoenyzme (sigma54-holoenzyme) initiates transcription differs from that of other forms of RNA polymerase holoenzyme. sigma54-Holoenzyme recognizes promoters that have conserved elements in the ?12 and ?24 regions, having the consensus sequence 5?-TGGCACN4TTTGC(A/T)-3?. The spacing between the conserved GG and GC doublets (underlined in the consensus sequence) is critical, as any changes in the spacing result in failure of sigma54-holoenzyme to recognize the promoter. Sigma54-Holoenzyme binds to the promoter to form a closed promoter complex, but it is unable to initiate transcription in the absence of an activator protein. The activator binds to sites that are usually located 100-200 bp upstream of the transcriptional start site and makes transient contact with sigma54-holoenyzme through DNA looping. Productive interactions between the activator and sigma54-holoenzyme lead to the isomerization of the closed promoter complex to an open complex that is transcriptionally competent. To catalyze this isomerization reaction, the activator must hydrolyze ATP. The mechanism by which the activator couples ATP hydrolysis to open complex formation is not known, but it does not appear to involve phosphorylation of either the activator or sigma54-holoenzyme. The activities of activators of sigma54-holoenzyme are regulated in response to environmental signals. Many of the activators of sigma54-holoenzyme are response regulators in two-component regulatory systems, and phosphorylation of these proteins results in their activation. These response regulators are phosphorylated by their cognate protein histidine kinases in response to an environmental signal. Once phosphorylated, the response regulator activates transcription of other genes. The activator of sigma54-holoenzyme from B. burgdorferi is also a response regulator of a two-component system. The gene encoding the activator is in an operon with a gene encoding its cognate protein histidine kinase. Our preliminary data indicates that the activator controls expression of rpoS. Therefore, we refer to this activator as sigmaS regulator (SisR) and its cognate protein histidine kinase as sigmaS regulatory protein histidine kinase (SisK).