MT VET COBRE PROJECT 1: HOST-PATHOGEN COMMUNICATION IN TOXOPLASMA

这个子项目是许多研究子项目中利用 资源由NIH/NCRR资助的中心拨款提供。子项目和 调查员(PI)可能从NIH的另一个来源获得了主要资金， 并因此可以在其他清晰的条目中表示。列出的机构是 该中心不一定是调查人员的机构。 控制该寄生虫中缓殖子分化的分子机制完全未知。拉德克博士此前已经证明，I型弓形虫菌株对人工诱导的缓殖子分化和某些宿主细胞基因的过度表达具有天然抵抗力。拉德克博士的实验室使用(1)互补策略将基因从I型RH株转移到III型CTG寄生虫，以确定可能使I型株产生抗药性的基因；以及(2)当不同的宿主细胞基因在受感染的细胞中过度表达时，测试组蛋白甲基化是驱动诱导寄生虫发育的一个主要机制。他们已经将I型RH株的一个基因转移到III型株CTG寄生虫中，并恢复了从6300kBP开始映射到ChrVIII，在1200kbp映射到ChrXII的序列。目前的工作将集中于将候选耐药序列缩小到单个基因，这可能是控制缓殖体分化和宿主慢性感染发病的分子机制的基础。Radke博士最近发现，在HeLa细胞中，组蛋白3赖氨酸4、9和36处的新甲基化与CDA1的过度表达相一致，但这是否是CDA1的直接作用尚不清楚。确认(或不)由CDA1直接甲基化核心组蛋白残基的工作将使用定点突变来完成。与野生型蛋白相比，SET结构域中能够减少赖氨酸4、9、27或36位甲基化的部分残基的突变表明CDA1能够直接甲基化组蛋白残基。

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. Molecular mechanisms controlling bradyzoite differentiation in the parasite are completely unknown. Dr. Radke has shown previously that Toxoplasma Type I strains are naturally resistant to the artificially induced bradyzoite differentiation and to over-expression of some host cell genes. Dr. Radke's lab uses (1) a complementation strategy to transfer a gene from the Type I RH strain to a Type III strain CTG parasite to identify genes that may render Type I strains resistant to development; and (2) testing histone methylation as one primary mechanism driving induced parasite development when distinct host cell genes are over-expressed in the infected cell. They have transferred a gene from the Type I RH strain to a Type III strain CTG parasite and recovered sequence that maps to ChrVIII beginning at 6300 Kbp and ChrXII at 1200 Kbp. Present work will focus on narrowing the candidate resistance sequences to a single gene that may underlie the molecular mechanisms controlling bradyzoite differentiation and the pathogenesis of chronic infection in the host. Dr. Radke has recently shown that new methylation at lysines 4, 9, and 36 of histone 3 occur in concert with the over-expression of CDA1 in HeLa cells, but whether this is a direct action of CDA1 is unknown. Work to confirm (or not) direct methylation of core histone residues by CDA1 will be completed using site-directed mutagenesis. Mutation of select residues in the SET domain that are able to reduce methylation at lysine 4, 9, 27, or 36 relative to the wild-type protein would suggest CDA1 is able to directly methylate histone residues.