Mechanisms of Giardia Lamblia Cyst Wall Formation

贾第鞭毛虫囊壁形成机制

• 批准号: 6431333
• 负责人: FRANCES D. GILLIN
• 金额: $ 33.7万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN DIEGO
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-09-28 至 2005-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6431333
• 关键词: Giardia; cell components; cell membrane; cell wall; communicable disease transmission; disulfide bond; endocytosis; enzyme activity; epitope mapping; host organism interaction; intracellular parasitism; membrane biogenesis; protein biosynthesis; protein disulfide isomerase; protein folding; protein transport; proteomics; radionuclide double label; secretory protein; vesicle /vacuole

DESCRIPTION: (provided by the applicant): The cyst wall of Giardia lamblia is critical to its success as a parasite. This primitive extracellular matrix is responsible for giardial survival in the environment and for its ability to infect new hosts. Based on our extensive studies of giardial encystation, we have formulated a four-stage model of cyst wall protein (CWP) and cyst maturation. The goal of this revised proposal is to validate and expand that model for a more comprehensive understanding of this essential differentiation. We will focus in depth on CWP processing, transport, and assembly. Our central hypothesis is that assembly of the cyst wall requires co-coordinated activities within several intracellular compartments, but final maturation occurs on the cell surface. Specific Aim 1 is to evaluate the role of the novel giardial lysosomal/endosomal peripheral vesicle (PV) compartment in processing of CWPs. We will test the hypothesis that passage through the PV is necessary for CWP maturation. Alternatively, CWP may traffic through the PV in an endocytic pathway to recover CWP that has been released prematurely. We will use tetracycline-regulated expression of epitope-tagged CWPs and cell surface labeling to trace CWP. These studies will elucidate the role of CWP traffic through the PV in achieving mature cyst wall architecture and function. In Specific Aim 2, we will test the hypothesis that CWPs undergo sequential stages of S-S bond formation and re-arrangement catalyzed by unusual giardial protein disulfide isomerases, leading to formation of resistant fibrils on the cell surface. Specific Aim 3 is to test the hypothesis that iso-peptide bond formation is needed for differentiation into cysts and to identify the proteins cross linked, using specific labels and inhibitors. This revised proposal will reveal basic insights into formation, function, and evolution of an ancient eukaryotic extracellular matrix, using integrated biochemical, and cell biological, and molecular approaches. Since cystic stages are key to transmission of many parasites, cyst wall formation is a target for interruption of the life cycle.

描述：（由申请人提供）：贾第鞭毛虫的囊壁是 对其作为寄生虫的成功至关重要。这种原始的细胞外基质是 负责贾第虫在环境中的生存及其能力 感染新宿主。基于我们对贾第鞭毛虫包囊的广泛研究，我们 制定了囊肿壁蛋白（CWP）和囊肿的四阶段模型 成熟。此修订提案的目标是验证和扩展 模型，以便更全面地理解这种本质区别。 我们将深入关注 CWP 加工、运输和组装。我们的中央 假设囊肿壁的组装需要协调活动 在几个细胞内区室中，但最终成熟发生在 细胞表面。具体目标 1 是评估新型贾第虫的作用 CWP 处理中的溶酶体/内体外周囊泡 (PV) 区室。 我们将检验以下假设：通过 PV 对于 CWP 是必要的 成熟。或者，CWP 可以通过内吞方式通过 PV。 恢复已过早释放的 CWP 的途径。我们将使用 四环素调节表位标记的 CWP 和细胞表面的表达 标记以追踪 CWP。这些研究将阐明 CWP 流量的作用 通过PV实现成熟的囊肿壁结构和功能。在 具体目标 2，我们将检验 CWP 经历连续阶段的假设 异常贾第虫蛋白催化的 S-S 键形成和重排 二硫键异构酶，导致细胞上形成抗性原纤维 表面。具体目标 3 是检验异肽键的假设 分化为囊肿和鉴定蛋白质需要形成 使用特定的标记和抑制剂进行交联。 该修订后的提案将揭示对构成、功能和功能的基本见解 古代真核细胞外基质的进化，使用集成 生物化学、细胞生物学和分子方法。自囊性阶段以来 是许多寄生虫传播的关键，包囊壁的形成是其目标 生命周期的中断。