Analysis of Host Defense Genes to Toxoplasma gondii

描述（由申请人提供）：刚地弓形虫是一种广泛存在的细胞内人类病原体，属于顶复门，包括人类病原体疟原虫（疟疾的病因）和隐孢子虫。虽然弓形虫正在成为细胞内寄生研究的模型系统；我们对宿主抗性基因如何控制弓形虫生长的理解落后于我们对弓形虫基因如何调节宿主细胞内病原体生长的理解。对细胞内宿主防御弓形虫的机制的清晰理解将为如何在感染患者中控制这类重要的人类病原体提供有用的见解。

DESCRIPTION (provided by the applicant): Toxoplasma gondii is a widespread intracellular human pathogen in the same phylum Apicomplexa that includes the human pathogens Plasmodium (the cause of malaria) and Cryptosporidium. Although Toxoplasma is emerging as a model system for the study of intracellular parasitism; our understanding of how host resistance genes control Toxoplasma growth has lagged behind our understanding of how Toxoplasma genes regulate pathogen growth within host cells. A clear understanding of the mechanisms responsible for intracellular host defenses to Toxoplasma will provide useful insights into how this important class of human pathogens can be controlled in infected patients. A major block impeding the functional analysis of how host-cell gene expression controls intracellular Toxoplasma growth is the tack of sensitivity of in vitro assays that measure intracellular parasite growth. To address this issue, a novel FACS-based assay has been developed to measure growth of GFP-tagged Toxoplasma in host cells. This FACS-based assay is significantly more sensitive than previous Toxoplasma growth assays, and can measure the inhibition of parasite growth mediated by individual retroviral-expressed genes in un-stimulated host cells. This new assay permits examination of host-cell responses to Toxoplasma in greater detail than previously possible, and will be used in Aim 1 to determine the mechanism of action of known host resistance genes, particularly the IGTP family of interferon-induced genes whose mechanism of action is unknown, despite having been implicated in the pathogenesis of multiple intracellular pathogens. In proof of principle studies, we have also established that known host resistance genes can be identified in functional screens designed to identify novel host resistance genes. Thus, in Aim 2, functional genetic screens will be conducted using retroviral cDNA libraries to identify host resistance genes that control Toxoplasma infection by (1) active resistance of growth of the virulent tachyzoite form of Toxoplasma, and by (2) passive inter-conversion of the virulent tachyzoite form to the dormant bradyzoite form of Toxoplasma.