Functional Plasticity in the Helicobacter pylori Type IV Secretion System

项目总结/摘要：修订 幽门螺杆菌主要在感染携带细胞毒素相关基因致病岛（cagPAI）的菌株的个体中引起临床疾病。cagPAI编码IV型分泌系统（T4 SS），其是将CagA癌蛋白注射到上皮细胞中和诱导促炎细胞因子白细胞介素-8（IL-8）所需的。CagY是H. pylori T4 SS具有不寻常的序列结构，其中大量的直接DNA重复被预测会引起重排，而重排总是预测框内插入或缺失。我们已经在小鼠和非人灵长类动物模型中证明，免疫驱动的CagY重排的宿主选择足以导致H.幽门螺杆菌T4 SS。我们假设CagY作为一种分子变阻器，改变了T4 SS的功能，并“调谐”宿主的炎症反应，以最大限度地提高持续感染。我们提出了三个具体目标来检验这一假设。在目标1中，我们将确定CagY中重组改变H.幽门螺杆菌T4 SS。表达赋予功能性或非功能性T4 SS的变体CagY蛋白的同基因菌株将用于确定CagY通过β1整联蛋白结合和发信号的能力，表征细菌膜中CagY的拓扑结构，检查CagY与其他必需派蛋白之间的相互作用，并确定DNA重复序列是否是T4 SS功能所必需的。在目标2中，我们将描述宿主免疫在选择CagY介导的H. pylori T4 SS的基因敲除和转基因小鼠模型。目的3的目标是使用高度相关的恒河猴模型和来自慢性感染患者的菌株更好地理解CagY和T4 SS功能的生理作用。这些实验的完成将表征一种新的策略，通过该策略细菌分泌系统改变宿主免疫应答，并确定T4 SS和宿主免疫交叉的机制。这些实验还将增强我们对派与感染临床结果之间关系的理解，并导致对慢性感染和炎症之间关系的更广泛理解。

PROJECT SUMMARY/ABSTRACT: revised Helicobacter pylori causes clinical disease primarily in those individuals infected with a strain that carries the cytotoxin associated gene pathogenicity island (cagPAI). The cagPAI encodes a type IV secretion system (T4SS) that is required for injection of the CagA oncoprotein into epithelial cells and induction of the proinflammatory cytokine, interleukin-8 (IL-8). CagY is an essential component of the H. pylori T4SS that has an unusual sequence structure, in which an extraordinary number of direct DNA repeats is predicted to cause rearrangements that invariably predict in-frame insertions or deletions. We have demonstrated in murine and non-human primate models that immune-driven host selection of rearrangements in CagY is sufficient to cause gain or loss of function in the H. pylori T4SS. We hypothesize that CagY functions as a sort of molecular rheostat that alters the function of the T4SS and “tunes” the host inflammatory response so as to maximize persistent infection. We propose three specific aims to test this hypothesis. In Aim 1 we will determine the mechanism by which recombination in CagY alters the function of the H. pylori T4SS. Isogenic strains expressing variant CagY proteins that confer a functional or non-functional T4SS will be used to determine the capacity of CagY to bind and signal through β1 integrins, characterize the topology of CagY in the bacterial membrane, examine the interaction between CagY and other essential PAI proteins, and determine if the DNA repeats are essential for T4SS function. In Aim 2 we will characterize the role of host immunity in selection for CagY-mediated modulation of function in the H. pylori T4SS using knockout and transgenic mouse models. The goal of Aim 3 is to better understand the physiological role of CagY and T4SS function using the highly relevant rhesus macaque model and strains from chronically infected patients. Completion of these experiments will characterize a novel strategy by which a bacterial secretion system alters the host immune response, and identify the mechanisms where the T4SS and host immunity intersect. These experiments will also enhance our understanding of the relationship between the PAI and the clinical outcome of infection, and lead to a broader understanding of the relationship between chronic infection and inflammation.