Microneme function in toxoplasma

描述（由申请人提供）：在细胞内病原体（包括寄生原生动物弓形虫）的生命周期中，离开宿主细胞是至关重要的一步。弓形虫可在怀孕期间引起严重疾病或免疫功能障碍，并在其他健康人群中引起使人衰弱的眼部病变。在细胞内复制后，寄生虫以包括疟原虫在内的其他寄生虫共有的方式从宿主细胞中破裂。疟原虫的裂解释放使其能够感染新的宿主细胞，但随之而来的损害也会引发炎症和发烧，这是弓形虫病和疟疾的特征。我们最近发现，弓形虫的快速出口和致命的急性疾病都依赖于一种成孔蛋白TgPLP1的及时分泌。我们发现TgPLP1从根尖微基因组释放，揭示了这些受调节的分泌细胞器的新作用。TgPLP1在输出过程中如何裂解宿主膜尚不清楚。由于缺乏这方面的知识，就无法采取战略性措施来消灭其活动并改变感染过程。我们的长期目标是了解关键微素蛋白在弓形虫感染中的作用。本资助期的目标是确定TgPLP1如何在出口时选择性和快速地裂解宿主细胞膜，而不会引起寄生虫的自我损伤。我们假设TgPLP1的孔形成是由其靶膜的脂质组成、环境因素和蛋白质的特定结构特征决定的。这一论断是基于初步数据，确定了选择性裂解的宿主脂质受体，pH和蛋白质水解在调节孔形成中的潜在作用，以及TgPLP1预测驱动膜结合和寡聚化的保守结构特征。具体目的是：(1)揭示宿主而非寄生虫膜选择性细胞溶解的机制；(2)揭示TgPLP1在输出时活跃而在入侵时不活跃；(3)确定快速孔隙形成的分子基础。通过对弓形虫生命周期的一个重要步骤提供更深入的机制理解，我们期望所提出的工作将创造新的机会来中断感染并可能改善疾病。

DESCRIPTION (provided by applicant): Egress from host cells is a crucial step in the life cycle of intracellular pathogens including the parasitic protozoan Toxoplasma gondii. T. gondii can cause severe disease during pregnancy or immune dysfunction, and debilitating ocular pathology in otherwise healthy people. After intracellular replication, the parasite ruptures from host cells in a manner shared by other parasites including Plasmodium. Lytic egress frees the parasite to infect new host cells but the ensuing damage also fuels inflammation and fever, hallmarks of toxoplasmosis and malaria. We recently showed that T. gondii rapid egress and fatal acute disease both depend on the timely secretion of a pore-forming protein, TgPLP1. Our finding that TgPLP1 is released from apical micronemes exposed a novel role for these regulated secretory organelles. How TgPLP1 lyses host membranes during egress is not known. The absence of such knowledge precludes strategic efforts to extinguish its activity and alter the course of infection. Our long-term goal is to understand the roles of key microneme proteins in T. gondii infection. The objective for this funding period is to determine how TgPLP1 selectively and rapidly lyses host cell membranes during egress without causing parasite self-damage. We hypothesize that TgPLP1 pore formation is dictated by the lipid composition of its target membranes, environmental factors, and specific structural features of the protein. This assertion is based on preliminary data identifying host lipid receptors targeted for selective lysis, potential roles for pH and proteolysis in regulating pore formation, and conserved structural features of TgPLP1 predicted to drive membrane binding and oligomerization. The specific aims are: (1) Reveal the mechanism for selective cytolysis of host but not parasite membranes; (2) Uncover how TgPLP1 is active during egress but not during invasion; and (3) Identify the molecular basis for rapid pore formation. By providing a deeper mechanistic understanding of an essential step in the T. gondii life cycle, we expect the proposed work will create novel opportunities to interrupt infection and potentially ameliorate disease.