Par1-substrates responsible for CagA-mediated pathogenesis of Helicobacter pylori

描述(申请人提供)：导致CagA介导的幽门螺杆菌胃炎和胃癌的Par1底物是世界上与癌症相关的死亡的第二大原因，它与幽门螺杆菌的感染有关，幽门螺杆菌是一种定植于胃粘膜的细菌。含有细胞毒素相关基因A(CagA)基因的幽门螺杆菌菌株比缺乏CagA基因的菌株毒力更强，对宿主的癌症风险要高得多。最近，这种毒素在小鼠身上被证明是一种癌基因。CAGA通过激活RAS-MAP-CAGA通路，诱导胃上皮细胞极性丧失，细胞迁移和细胞凋亡增加，促进异常生长信号。最近，我们和其他人已经确定丝氨酸/苏氨酸激酶Par1是与幽门螺杆菌病理相关的上皮极性缺陷的CagA靶点。目前面临的挑战是描绘Par1下游的CagA信号通路(S)。虽然PAR1是一个已知的极性决定因素，但目前还没有发现介导哺乳动物上皮极性的PAR1底物。在初步实验中，我们开发了一种无偏的Par1底物筛选，使我们能够识别63种可能的底物，其中大多数是新的，以及它们在上皮细胞中精确的Par1b磷酸化位点。我们的目标是评估所有经过验证的底物对CagA介导的上皮细胞形状和极性丧失的贡献。既然我们已经确定Par1b抑制上皮细胞的增殖并负向调节mTOR信号，我们也将评估Par1b底物是否与CagA对增殖的影响有关。在两层方法中，我们将首先描述Par1b的特征 CagA下游底物在肾上皮细胞系MDCK中的表达及其对原代人胃上皮细胞幽门螺杆菌感染的意义 公共卫生相关性：胃癌是世界上与癌症相关的死亡的第二大原因，也是第四种最常见的癌症，全世界每年约有930,000例新诊断病例。这种疾病与幽门螺杆菌感染有关，幽门螺杆菌是一种定植于胃粘膜的细菌。我们的项目旨在确定幽门螺杆菌导致胃上皮肿瘤发生的新的信号机制。

DESCRIPTION (provided by applicant): Par1-substrates responsible for CagA-mediated pathogenesis of Helicobacter pylori Gastritis and gastric carcinoma, the second leading cause of cancer-related deaths in the world, have been linked to infection with Helicobacter pylori, a bacterium that colonizes the gastric mucosa. H. pylori strains that harbor a gene called cytotoxin-associated gene A (CagA) are more virulent and present a much higher cancer risk for their host than strains that lack CagA and recently the toxin has been shown to function as an oncoene in mice. CagA induces loss of cell polarity, increased cell migration and apoptosis of gastric epithelial cells and promotes aberrant growth signals by activating the Ras-MAP-cascade. Recently, we and others have identified the serine/threonine kinase Par1 as a CagA-target responsible for epithelial polarity defects associated with H. pylori pathology. The challenge at hand now is to delineate the CagA signaling pathway(s) downstream of Par1. Although a known polarity determinant, Par1-substrates that mediate mammalian epithelial polarity have not yet been discerned. In preliminary experiments, we have developed an unbiased Par1 substrate screen that enabled us to identify 63 putative substrates, most of them novel, and their precise Par1b phosphorylation sites in epithelial cells. Our goal is to evaluate the contribution of all validated substrates to CagA-mediated loss of epithelial cell shape and polarity. Since we have determined that Par1b inhibits proliferation and negatively regulates mTOR signaling in epithelial cells, we will also evaluate whether Par1b substrates are relevant for the effects of CagA on proliferation. In a two-tiered approach we will first characterize Par1b substrates downstream of CagA in the kidney-derived epithelial model cell line MDCK and subsequently validate their significance for H. pylori infection of primary human gastric epithelia PUBLIC HEALTH RELEVANCE: Gastric carcinoma is the second leading cause of cancer-related deaths in the world and the fourth most common cancer with approximately 930,000 new cases diagnosed worldwide annually. The disease has been linked to infection with Helicobacter pylori, a bacterium that colonizes the gastric mucosa. Our project is designed to identify novel signaling- mechanisms by which H. pylori leads to tumorigenesis of the stomach epithelium.