Cyclin-mediated control of Toxoplasma development

细胞周期蛋白介导的弓形虫发育控制

• 批准号: 10161722
• 负责人: Elena Suvorova
• 金额: $ 37.38万
• 依托单位: UNIVERSITY OF SOUTH FLORIDA
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-05-08 至 2025-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10161722
• 关键词: Acquired Immunodeficiency Syndrome; Acute; Affect; Animal Model; Animals; Architecture; Auxins; Biochemical; Biological; Biological Assay; Biology; Cell Cycle; Cell division; Cells; Chronic; Chronic Disease; Clinical; Complex; Cyclins; Cyst; Development; Differentiation and Growth; Disease; Disease Outbreaks; Disease Progression; Drug Targeting; Ensure; Equilibrium; Eukaryota; Evaluation; Foundations; Frequencies; G1 Phase; Gene Expression; Genetic; Genetic Models; Genome; Goals; Human; Immune; Immune response; Immune system; Immunity; Immunocompromised Host; In Vitro; Individual; Infection; Kentucky; Life; Link; Measures; Mechanics; Mediating; Modeling; Monitor; Mothers; Mus; Organ Transplantation; Parasites; Parasitic infection; Patients; Pharmaceutical Preparations; Phosphotransferases; Population; Post-Translational Protein Processing; Pregnancy; Proteomics; Readiness; Regulation; Rest; Role; Site; Specialist; Testing; Time; Toxoplasma; Toxoplasma gondii; Toxoplasmosis; Universities; Woman; base; chemotherapy; differential expression; effective therapy; experience; experimental study; human pathogen; immune function; in vitro activity; in vivo; mouse model; mutant; novel; prenatal; prevent; protein expression; success; transmission process

Project Abstract Toxoplasma gondii is an important human pathogen that causes severe disease in immunocompromised individuals, such as those undergoing chemotherapy, organ transplantation, and AIDS patients. It also afflicts women who become infected for the first time during pregnancy. A healthy immune system or current drug regiment controls the replication of the tachyzoites associated with clinical toxoplasmosis. However, there are no effective therapies to eliminate the chronic stages associated with encysted bradyzoites and, importantly, to prevent the cyst reactivation. To find new avenues for combating the chronic and reactivated toxoplasmosis, we focus on the mechanisms of the tachyzoite and bradyzoite interconversions that are poorly understood. The critical difference between a tachyzoite and a bradyzoite is the rate of parasite replication and the cell cycle architecture. A tachyzoite divides fast and has a relatively short G1 period. In contrast, a bradyzoite rarely divides and spends a progressively longer time in the G1 phase. The time parasite spends in the G1 period is regulated by the RESTRICTION checkpoint (R-point) that in T. gondii lacks conventional regulators. The current application is based on the central hypothesis that the atypical TgCrk2 kinase and P-type cyclins define the novel G1 checkpoint that governs transitions between acute and chronic toxoplasmosis. In our preliminary studies, we showed a differential expression of three P-cyclins in the fast- (RH) and slow-dividing (ME49) T. gondii strains, which also differ in their ability to differentiate. We also showed that all three P-cyclins interact with G1 kinase TgCrk2 in vivo. We believe that P-cyclins differentially regulate levels of TgCrk2 activity, therefore, facilitate or block R-point passage. This dictates the parasite’s choice to either replicate as a tachyzoite or to convert into a resting bradyzoite. To prove our hypothesis, we will define the mechanism of the R-point regulation by TgCrk2 kinase and cyclins TgCycP2, and TgCycP3 in the tachyzoite and bradyzoite development in vitro (Aim 1); in the natural progression of the disease using mouse model (Aim 2); and determine function of the novel R-point components TgCables1 and TgRch1 (Aim 3). Our proposal is built on a strong foundation of advanced genetics, which is bolstered by our extensive experience in studying cyclin/Crk regulators of the T. gondii tachyzoite cell cycle. Using a panel of strains with novel auxin-dependent conditional protein expression of P-cyclins, we will test whether differences in regulation of TgCrk2/P-cyclin complexes explain the dynamics of the bradyzoite differentiation and cyst reactivation. Altogether, our experiments will break new ground in understanding the mechanics and regulation of the developmental switch that regulates the progression of the disease.

项目摘要 弓形虫是一种重要的人类病原体，在免疫低下的情况下会引起严重的疾病。 个人，如那些正在接受化疗、器官移植和艾滋病患者。它还折磨着 在怀孕期间首次感染的妇女。健康的免疫系统或当前的药物 Regiment控制与临床弓形虫病相关的速殖子的复制。然而，有一些 没有有效的治疗方法来消除与包囊慢殖子相关的慢性阶段，更重要的是， 防止囊肿复活。为了寻找新的途径对抗慢性和复发的弓形虫病，我们 把重点放在速殖子和缓殖子相互转换的机制上，这些机制还知之甚少。这个 速殖子和缓殖子之间的关键区别是寄生虫的复制速度和细胞周期 建筑。速殖子分裂快，具有相对较短的G1周期。相比之下，缓殖子很少分裂 并且在G1阶段花费逐渐较长的时间。寄生虫在G1期的时间是受调节的 通过弓形虫缺乏常规调节器的限制检查点(R点)。海流 应用基于中心假设，即非典型的TgCrk2激酶和P型细胞周期蛋白定义 新的G1检查点，控制急性和慢性弓形虫病之间的转换。在我们的预赛中 研究表明，三种P-细胞周期蛋白在快分裂(RH)和慢分裂(ME49)T细胞中的表达存在差异。 它们的分化能力也不同。我们还表明，所有三个P-细胞周期蛋白都相互作用 在体内与G1激酶TgCrk2结合。我们认为，P-Cyclins对TgCrk2活性的调节具有差异性，因此， 促进或阻止R点通道。这决定了寄生虫的选择，要么作为速殖子复制，要么 变成一个静止的缓虫体。为了证明我们的假设，我们将定义R点调节的机制 TgCrk2蛋白和细胞周期蛋白TgCycP2、TgCycP3在速殖子和缓殖子发育中的作用 体外(目标1)；在疾病的自然发展过程中使用小鼠模型(目标2)；以及确定 新的R点组件TgCables1和TgRch1(目标3)。我们的建议建立在一个坚实的基础上 先进的遗传学，这是由我们在研究T. 弓形虫速殖子细胞周期。使用一组具有新的生长素依赖条件蛋白表达的菌株 对于P-细胞周期蛋白，我们将测试TgCrk2/P-细胞周期蛋白复合体调控的差异是否解释了 缓殖子分化和包囊重新激活。总之，我们的实验将在 理解发育开关的机制和调节，该开关调节 疾病。