Transcriptional Control of the Helicobacter pylori RpoN Regulon

描述（由申请人提供）：幽门螺杆菌是人类消化性溃疡疾病和多种其他疾病的病原体，包括胃癌和粘膜相关淋巴瘤。幽门螺杆菌穿过胃粘膜层并定植胃上皮需要动力，而幽门螺杆菌使用一簇极性鞭毛来完成这一过程。细菌中鞭毛基因表达的调控是复杂的，涉及到鞭毛基因表达与鞭毛组装耦合的转录层次。研究的长期目标是确定幽门螺旋杆菌调节鞭毛基因表达的机制。备选的sigma因子RpoN是几个幽门螺旋杆菌鞭毛基因转录所必需的。这些rpo依赖基因的转录也需要FlgS/FlgR双组分系统。幽门螺杆菌rpo依赖性鞭毛基因的表达与鞭毛蛋白输出装置有关，因为编码输出装置成分的基因突变干扰了rpo调控子的表达。出口装置影响RpoN调控子表达的机制尚不清楚。本项目的目的是确定出口装置控制幽门螺杆菌RpoN鞭毛调控子表达的分子机制。工作假设是，出口装置通过分泌RpoN调节子的抑制剂或通过蛋白质-蛋白质相互作用影响传感器激酶FlgS的活性来影响幽门螺杆菌RpoN调节子的转录。这一假设将在三个具体目标中进行客观检验。第一个具体目标是确定FlgR的组成活性形式是否绕过输出装置突变，从而允许rpo依赖性报告基因的表达。在第二个特定目标中，将检查具有将出口装置锁定在有利于杆/钩型底物构象的突变的幽门螺杆菌菌株，以了解它们如何影响rpo依赖性报告基因的表达。第三个具体目标是通过破坏编码该蛋白的基因来确定新型蛋白HP1042是否是RpoN调控子的抑制剂，并确定该突变如何影响幽门螺杆菌中ron依赖性报告基因的表达。这项提议的工作与NIH促进美国公众健康的使命有关，因为研究结果可能导致确定控制或检测幽门螺杆菌的新目标。这一信息对于开始降低与幽门螺杆菌感染相关的发病率和死亡率至关重要。公共卫生相关性：幽门螺杆菌是消化性溃疡疾病和其他胃部疾病的病原体，在美国每年导致大约100万人住院和6500人死亡。所提出的研究结果可能会导致确定新的靶点来控制或检测幽门螺杆菌，这对于开始降低与这一重要病原体相关的发病率和死亡率是重要的。

DESCRIPTION (provided by applicant): Helicobacter pylori is the causative agent of peptic ulcer disease and a variety of other diseases in humans, including gastric cancer and mucosal-associated lymphoma. Motility is required for H. pylori to cross the gastric mucous layer and colonize the gastric epithelium, and H. pylori use a cluster of polar flagella to accomplish this. Regulation of flagellar gene expression in bacteria is complex and involves a transcriptional hierarchy that couples the expression of flagellar genes with assembly of the flagellum. The long-term objective of proposed research is to identify mechanisms used by H. pylori to regulate flagellar gene expression. The alternative sigma factor RpoN is needed for transcription of several H. pylori flagellar genes. Transcription of these RpoN-dependent genes also requires the FlgS/FlgR two-component system. Expression of H. pylori RpoN-dependent flagellar genes is linked to the flagellar protein export apparatus as mutations in genes encoding components of the export apparatus interfere with expression the RpoN regulon. The mechanism by which the export apparatus influences expression of the RpoN regulon is unknown. The goal of this project is to identify the molecular mechanism by which the export apparatus controls expression of the H. pylori RpoN flagellar regulon. The working hypothesis is that the export apparatus affects transcription of the H. pylori RpoN regulon by secreting an inhibitor of the RpoN regulon or influencing the activity of the sensor kinase FlgS through protein-protein interactions. This hypothesis will be tested objectively in three specific aims. The first specific aim will determine if a constitutively active form of FlgR bypasses export apparatus mutations to allow expression of RpoN-dependent reporter genes. In the second specific aim H. pylori strains with mutations that lock the export apparatus in a conformation that favors rod-/hook-type substrates will be examined to see how they influence expression of RpoN-dependent reporter genes. The third specific aim will determine if the novel protein HP1042 is an inhibitor of the RpoN regulon by disrupting the gene encoding this protein and determining how this mutation affects expression of RpoN-dependent reporter genes in H. pylori. The proposed work is relevant to NIH's mission of promoting public health in the United States as the results of the studies may lead to identification of new targets for the control or detection of H. pylori. This information is essential to begin to reduce the morbidity and mortality associated with H. pylori infections. PUBLIC HEALTH RELEVANCE: Helicobacter pylori is a causative agent of peptic ulcer disease and other gastric diseases, causing an estimated 1 million hospitalizations and 6,500 deaths each year in the United States. Results of the proposed studies may lead to the identification of new targets for the control or detection of H. pylori, which is important to begin to reduce the morbidity and mortality associated with this significant pathogen.