GLOBAL ANALYSIS OF THE HOST CELL RESPONSE TO TOXOPLASMA GONDII INFECTION IN ASTR

这个子项目是许多研究子项目中的一个 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 弓形虫（Toxoplasma gondii）是中枢神经系统的主要机会致病菌，其在艾滋病患者和其他免疫功能低下的个体（如移植受者和接受化疗的癌症患者）中引起显著疾病。在这些个体中，感染是由于大脑中潜伏感染的重新激活，并导致严重且通常致命的坏死性脑炎。细胞因子在T.γ-干扰素（IFN-γ）是控制弓形虫复制的主要细胞因子。在大脑和其他组织中的弓形虫。在鼠星形胶质细胞和巨噬细胞中IFN γ抑制的机制部分依赖于IFN γ诱导的应答蛋白，GTP。自然宿主防御机制受到寄生虫的影响，已知寄生虫会显著改变宿主细胞的转录，并且有明确的证据表明寄生虫会主动干预影响宿主细胞凋亡和细胞因子反应的途径。这场拔河的生化结果影响了宿主细胞环境的建立，该环境要么支持要么不利于寄生虫的生长和发育。宿主细胞成分和寄生虫操纵宿主细胞环境的因素尚不清楚，但这些机制似乎在表达广泛生长和毒力表型的菌株之间存在广泛差异。星形胶质细胞是T.弓形虫是脑内重要的IFN γ激活效应细胞，介导对弓形虫的抗性。大脑中的弓形虫。因此，星形胶质细胞模型提供了一个机会，以了解宿主防御和寄生虫的生存机制，在临床相关的细胞类型。我们将采取全面的方法来表征在IFNy刺激提供保护的条件下，从体外暴露于弓形虫感染的动物中获得的原代星形胶质细胞中发生的宿主mRNA和蛋白质表达以及蛋白质翻译后修饰的变化。在这个项目中。我们将定义星形胶质细胞中的IFN γ反应，并探索这些宿主细胞的变化如何被不同遗传谱系和毒性表型的寄生虫改变。本研究的三个具体目标是：1）宿主细胞对T. IFN γ刺激的星形胶质细胞与未刺激的星形胶质细胞中的弓形虫感染，2）宿主细胞对T. IFN γ刺激的星形胶质细胞与未刺激的星形胶质细胞中的弓形虫感染和3）T.弓形虫存活突变体在星形胶质细胞中受到IFN γ刺激。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Toxoplasma gondii is a major opportunistic pathogen of the central nervous system, that causes significant disease in AIDS patients and other immunocompromised individuals, such as transplant recipients and cancer patients undergoing chemotherapy. In these individuals infection is due to reactivation of a latent infection in the brain and results in severe and often fatal necrotizing encephalitis. Cytokines play an important role in the regulation of T. gondii in the central nervous system and interferon-gamma (IFNv) is the main cytokine controlling replication of T. gondii in the brain and in other tissues. The mechanism of IFNy inhibition in murine astrocytes and macrophages is partially dependent upon the IFNv-induced response protein, IGTP. Natural host-defense mechanisms are influenced by the parasite, which is known to substantially alter host cell transcription, and there is clear evidence for active parasite intervention in pathways affecting host cell apoptosis and the cytokine response. The biochemical outcome of this tug-of-war influences the establishment of a host cell environment that either supports or is hostile to parasite growth and development. The host cell components and the factors by which the parasite manipulates the host cell environment are not understood, but these mechanisms appear to vary extensively between strains that express a wide range of growth and virulent phenotypes. Astrocytes are an important host cell for T. gondii in the brain and an important IFNy-activated effector cell, mediating resistance to T. gondii in the brain. As such, the astrocyte model provides an opportunity to understand host-defense and parasite survival mechanisms in a clinically relevant cell type. We will take a comprehensive approach to characterize the changes in host mRNA and protein expression and protein post-translational modification that occur in primary astrocytes obtained from animals that are exposed in vitro to Toxoplasma infection, under conditions where protection is afforded by IFNy stimulation in this project. We will define the IFNy response in astrocytes and explore how these host cell changes are altered by parasites of distinct genetic lineage and virulent phenotypes. The three specific aims of this proposal are: 1) Transcriptional analysis of the host cell response to T. gondii infection in IFNy stimulated vs. unstimulated astrocytes, 2) Proteomic analysis of the host cell response to T. gondii infection in IFNy stimulated vs. unstimulated astrocytes and 3) Development of a screen for T. gondii survival mutants subjected to IFNy stimulation in astrocytes.