Induction of Macrophage iNOS by Salmonella

描述（由申请人提供）：肠源性大肠杆菌的各种血清型 革兰氏阴性细菌病原体沙门氏菌负责许多 对公众健康有重大影响的疾病，包括急性肠胃炎， 以及伤寒。在感染过程中，这些生物体侵入 宿主的肠上皮细胞、树突细胞和巨噬细胞。在 这样，细菌将特定的效应蛋白引入宿主细胞， 通过一个特殊的分泌器官细胞骨架的变化， 由这些蛋白质诱导的细胞信号传导途径的激活促进了 细菌入侵，也引起宿主促炎性细胞因子的产生， 分子。阐明了效应蛋白是如何执行这些功能的 功能将有助于阐明发病机制，并可能提出新的 沙门氏菌相关疾病的治疗方法。初步 实验中，我发现效应子SopE 2，一种鸟嘌呤核苷酸， 哺乳动物Rho GTP酶的交换因子，是沙门氏菌所必需的 诱导型一氧化氮合酶（iNOS）的依赖性上调， 它负责控制一氧化氮（NO）的产生， 巨噬细胞NO的促炎和免疫调节作用有助于 抗微生物防御以及相关的组织损伤 感染了在进一步的研究中，我发现SopE 2激活了 转录因子NF-κ-B，无论是其本身，并在协同 与TRAF 6的相互作用，TRAF 6是一种通过成员参与信号传导的衔接分子 Toll样受体和TNF受体家族。提出的实验 在本申请中，将扩展这些初步观察以阐明 SopE 2及其同源物SopE的功能，通过（a）检查机制， SopE 2和SopE启动导致iNOS诱导的信号，特别是 Rho GTP酶在该过程中的作用（B）阐明SopE 2和SopE 激活NF-κ-B，并表征其对TRAF依赖性 信号，（c）鉴定所述细胞中的顺式作用转录调控元件， 应答沙门氏菌感染和应答Sop诱导信号iNOS启动子， 和（d）检查SopE 2和SopE在沙门氏菌诱导的iNOS中的作用， 原代巨噬细胞和树突细胞。这些研究的结果将揭示 了解SopE和SopE 2的新功能，也将有助于理解 调节iNOS表达的机制。

DESCRIPTION (provided by applicant): Various serotypes of the enteric Gram-negative bacterial pathogen Salmonella are responsible for a number of diseases of public health significance, including acute gastroenteritis, as well as typhoid fever. During the course of infection, these organisms invade intestinal epithelial cells, dendritic cells and macrophages of the host. In doing so, the bacteria introduce specific effector proteins into the host cells through a specialized secretory apparatus. The cytoskeletal changes and activation of cellular signaling pathways induced by these proteins facilitate bacterial invasion and also elicit the production of host pro-inflammatory molecules. Elucidating exactly how the effector proteins carry out these functions would help to clarify the pathogenesis of, and might suggest new approaches to treating, Salmonella -associated disease. In preliminary experiments, I have found that the effector SopE2, a guanine nucleotide exchange factor for mammalian Rho GTPases, is necessary for the Salmonella -dependent upregulation of inducible nitric oxide synthase (iNOS), the enzyme that is responsible for controlling the production of nitric oxide (NO) in macrophages. The pro-inflammatory and immunomodulatory effects of NO contribute to anti-microbial defense, as well as to the tissue damage that is associated with infection. In further studies, I have found that SopE2 activates the transcription factor NF-kappa-B, both on its own, and in a synergistic interaction with TRAF6, an adaptor molecule involved in signaling via members of the Toll-like receptor, and TNF receptor families. The experiments proposed in this application will extend these preliminary observations to elucidate the function of SopE2, and its homolog SopE, by (a) examining the mechanism by which SopE2 and SopE initiate signals leading to iNOS induction, particularly the role of the Rho GTPases in this process (b) elucidating how SopE2 and SopE activate NF-kappa-B, and characterizing their influence on TRAF-dependent signals, (c) identifying cis-acting transcriptional regulatory elements in the iNOS promoter that respond to Salmonella infection, and to Sop-induced signals, and (d) examining the role of SopE2 and SopE in iNOS induction by Salmonella in primary macrophages and dendritic cells. The results of these studies will shed light on a novel function of SopE and SopE2 and will also improve understanding of the mechanisms that regulate iNOS expression.