Genetic dissection of the Toxoplasma gondii rhoptry kinome

弓形虫菱形激酶组的遗传解剖

• 批准号: 8267596
• 负责人: DAVID J BZIK
• 金额: $ 23.7万
• 依托单位: DARTMOUTH COLLEGE
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-07-01 至 2014-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8267596
• 关键词: AIDS-Related Opportunistic Infections; Acquired Immunodeficiency Syndrome; Acute; Alleles; Biological; Biological Assay; Biological Process; Biology; Brain; Cell Nucleus; Cells; Chronic; Complex; Cyst; Cytosol; Data; Defect; Development; Dissection; Encephalitis; Exhibits; Failure; Family; Family member; Firefly Luciferases; Future; Genes; Genetic; Genetic Models; Human; Image; Immunosuppression; In Vitro; Infection; Infection prevention; Knock-out; Knowledge; Life; Maintenance; Mammalian Cell; Methods; Modeling; Mus; Parasites; Patients; Pattern; Pharmaceutical Preparations; Phenotype; Phosphoric Monoester Hydrolases; Phosphotransferases; Proteins; Publishing; Recurrence; Reporter; Research; Research Design; Resistance; Rupture; Specific qualifier value; Staging; Sum; Tissues; Toxoplasma gondii; Toxoplasmosis; Transgenic Organisms; Vaccines; Vacuole; Validation; Virulence; Work; XRCC5 gene; functional genomics; improved; in vivo; innovation; killings; knockout gene; member; mutant; novel strategies; prevent; promoter; rhoptry; success; tool

DESCRIPTION (provided by applicant): Toxoplasma gondii is an important opportunistic infection of AIDS patients. Improved strategies and approaches are urgently needed to more effectively prevent and treat recurrent infections in AIDS. Following primary infection, a chronic life-long T. gondii infection characterized by the presence of persisting cysts containing slowly or nonreplicating bradyzoite parasite forms is established in CNS/brain. Reactivated infection due to cyst rupture during immune suppression causes a difficult to treat and life-threatening Toxoplasmic encephalitis in AIDS. Currently, no vaccine is approved for use in humans to prevent infection and drug treatments for acute infection are suboptimal. Eradicating pre-existing cysts and chronic infection is an excellent approach to control/prevent infection in AIDS. However, no current treatment is effective at eradicating pre-existing cysts and chronic infection. Lack of progress in targeting cyst stages can be attributed to major gaps in our knowledge of parasite biology underlying cyst development and maintenance of chronic infection. Recent progress in understanding the complex host-parasite interaction has revealed that many rhoptry bulb (ROP) proteins appear to be central players in host cell manipulation due to their secretion into the host cell at invasion and their subsequent localization to host cell cytosol/nucleus or their intimate association with the parasitophorous vacuole. Host cell manipulation by these secreted ROP proteins is likely to be critical to the success of parasite biology necessary for acute and chronic infection in vivo. The parasitophorous vacuole appears to be critical for replication of tachyzoites as well as being central to the development of the cyst wall and tissue cyst that characterize chronic infection. We hypothesize that a family of ~ 34 secreted ROP proteins possessing homology to kinases (the "ROP kinome") are outstanding candidates as potential targets to disrupt virulence, cyst development, or chronic infection. Here, we propose to delete each predicted gene member of the secreted ROP kinome in type II T. gondii, and to then ascertain any defect in acute virulence, in early cyst development, or in chronic infection. Using a directed functional genetic approach, this work will identify specific members of the secreted ROP kinome that participate in critical functions required for establishing or maintaining chronic infection in vivo. This project will identify and validate specific secreted ROP kinases as key targets for preventing acute or chronic infection. This project will also develop essential tools and methods required for the future undertaking of larger scale higher throughput knockout and functional genomic projects in type II T. gondii. Consequently, this innovative project has high overall impact by advancing genetic models for dissection of type II biology, by revealing new aspects of host-parasite interaction, and by exposing new targets to prevent or eradicate chronic infection.

描述（由申请人提供）： 弓形虫Gondii是艾滋病患者的重要机会感染。迫切需要改进的策略和方法，以更有效地预防和治疗艾滋病中的复发感染。原发性感染后，CNS/大脑中建立了持续存在的囊肿的慢性终生t. gondii感染，该感染的特征是持续存在的囊肿持续存在囊肿，该囊肿含有缓慢或不复制的Bradyzoite寄生虫形式。免疫抑制期间由于囊肿破裂引起的重新激活的感染导致艾滋病中难以治疗和威胁生命的毒质性脑炎。目前，未批准用于人类使用疫苗以防止感染，急性感染的药物治疗是次优的。根除预先存在的囊肿和慢性感染是控制/防止艾滋病感染的极好方法。但是，目前没有治疗可有效地消除预先存在的囊肿和慢性感染。靶向囊肿阶段缺乏进展可以归因于我们对寄生虫生物学基础囊肿开发和维持慢性感染的主要差距。了解复杂的宿主 - 寄生虫相互作用方面的最新进展表明，由于它们在入侵时分泌到宿主细胞中，并且随后将宿主的细胞质/核或与寄生虫液泡的亲密关联，许多Rhoptry Bulb（ROP）蛋白似乎是宿主细胞操作中的核心参与者。这些分泌的ROP蛋白的宿主细胞操纵对于体内急性和慢性感染所必需的寄生虫生物学的成功可能至关重要。寄生虫液泡似乎对于复制速氮族的复制至关重要，并且对于囊肿壁和表征慢性感染的组织囊肿的发展至关重要。我们假设一个〜34个分泌的ROP蛋白质家族，与激酶具有同源性（“ ROP Kinome”）是杰出的候选者，是破坏毒力，囊肿发育或慢性感染的潜在目标。在这里，我们建议在II型T. gondii中删除分泌的ROP Kinome的每个预测基因成员，然后确定急性毒力，早期囊肿发育或慢性感染中的任何缺陷。使用定向的功能遗传方法，这项工作将确定参与体内建立或维持慢性感染所需的关键功能的分泌ROP Kinome的特定成员。该项目将识别并验证特定的分泌ROP激酶是预防急性或慢性感染的关键靶标。该项目还将开发未来在II型T. gondii中进行更大规模吞吐量敲除和功能基因组项目所需的基本工具和方法。因此，通过揭示宿主 - 寄生虫相互作用的新方面以及暴露新目标以预防或消除慢性感染，这一创新项目通过推进了II型生物学的遗传模型，从而具有很大的总体影响。