Investigating Entamoeba histolytica colonic pathogenesis

描述（由申请人提供）：原生动物寄生虫溶组织内阿米巴每年导致大约5000万例侵袭性疾病。阿米巴感染最常见的表现是结肠疾病和肝脓肿。我们的目标是确定新的毒力决定因素在溶组织芽胞杆菌的目的是了解阿米巴发病机制的分子基础。我们之前已经开发并使用微阵列技术来鉴定那些表达仅限于毒性菌株和条件的基因。其中一个编码溶肠杆菌丝氨酸、苏氨酸、异亮氨酸富蛋白（EhSTIRP）的基因仅在毒力菌株和条件下表达，在非毒力菌株和条件下表达极低或不表达。初步数据表明，EhSTIRP下调的寄生虫体外粘附力和毒力下降。此外，在EhSTIRP中，下调的寄生虫降低了细胞骨架基因的表达。我们的假设是，EhSTIRP是溶组织芽胞杆菌的关键毒力决定因素，在寄生虫粘附和阿米巴发病相关的信号传导中具有潜在作用。我们拟进一步研究EhSTIRP在体外对溶组织芽胞杆菌毒力的贡献，确定EhSTIRP是否为体内毒力的必要条件，确定宿主入侵过程中EhSTIRP的定位，表征与寄生虫毒力相关的EhSTIRP结构域，以及鉴定与EhSTIRP相互作用的阿米巴蛋白。我们将使用遗传和生化方法来解剖基因功能，活细胞成像来确定阿米巴宿主细胞相互作用期间EhSTIRP的定位，微阵列分析来定义与EhSTIRP基因操作相关的寄生虫转录组变化。这些方法有望确定EhSTIRP在阿米巴发病机制中的作用，并剖析与EhSTIRP相关的遗传和生化网络，并调节溶组织内阿米巴的毒力。公共卫生相关性：溶组织内阿米巴是一种重要的病原体，在全球范围内对人类健康产生影响。主要表现为痢疾和肝脓肿。虽然大多数疾病发生在发展中国家，但这种寄生虫可以在水净化系统受到不利影响的任何地方引起感染。海啸或区域政治和社会基础设施动荡等事件可能导致疾病出现。我们对寄生虫致病的分子机制很感兴趣。

DESCRIPTION (provided by applicant): The protozoan parasite Entamoeba histolytica causes an estimated 50 million cases of invasive disease annually. The most common manifestations of amebic infection are colonic disease and liver abscesses. Our goal is to identify novel virulence determinants in E. histolytica with the aim of understanding the molecular basis of amebic pathogenesis. We have previously developed and used microarray technology to identify genes whose expression is restricted to virulent strains and conditions. One such gene, encoding an E. histolytica serine, threonine, isoleucine, rich protein (EhSTIRP) is expressed only in virulent strains and conditions and has very low to no expression in non-virulent strains and conditions. Preliminary data indicate that parasites in which EhSTIRP is downregulated have decreased adhesion and virulence in vitro. Additionally, in EhSTIRP downregulated parasites have reduced expression of cytoskeletal genes. Our hypothesis is that EhSTIRP is a key virulence determinant in E. histolytica with potential roles in parasite adhesion and signaling relevant to amebic pathogenesis. We propose to further characterize the contribution of EhSTIRP to E. histolytica virulence in vitro, determine whether EhSTIRP is necessary for virulence in vivo, identify the localization of EhSTIRP during host invasion, characterize domains of EhSTIRP with relevance to parasite virulence, and identify amebic proteins that interact with EhSTIRP. We will use genetic and biochemical approaches to dissect gene function, live cell imaging to determine EhSTIRP localization during amebic host cell interaction and microarray analysis to define parasite transcriptome changes associated with genetic manipulation of EhSTIRP. These approaches promise to identify the role of EhSTIRP in amebic pathogenesis and dissect the genetic and biochemical network that is associated with EhSTIRP and which regulates virulence in Entamoeba histolytica. PUBLIC HEALTH RELEVANCE: Entamoeba histolytica is an important pathogen and has an impact on human health on a global scale. The main disease manifestations are dysentery and liver abscesses. Although the majority of disease is in developing countries, this parasite can cause infections anywhere that water purification systems get adversely affected. Events such as the Tsunami or upheaval in the political and social infrastructure of a region can allow disease to emerge. We are interested in understanding the molecular mechanisms that the parasite uses to cause disease.