GIARDIA LAMBLIA CYSTEINE PROTEASES: TRAFFICKING, LOCALIZATION, AND FUNCTION

贾第鞭毛虫半胱氨酸蛋白酶：运输、定位和功能

• 批准号: 8169751
• 负责人: James H. McKerrow
• 金额: $ 0.18万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-09-12 至 2011-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8169751
• 关键词: Cellular biology; Computer Retrieval of Information on Scientific Projects Database; Confocal Microscopy; Cysteine Protease; Endocytosis; Endoplasmic Reticulum; Endosomes; Family; Funding; Genes; Giardia; Giardia lamblia; Golgi Apparatus; Grant; Institution; Label; Location; Mass Spectrum Analysis; Membrane; Mitochondria; Modeling; Nutrient; Organelles; Peripheral; Protein Export Pathway; Proteins; Reporter; Research; Research Personnel; Resources; Secretory Vesicles; Sequence Analysis; Source; Structure; System; United States National Institutes of Health; base; fast protein liquid chromatography; member; novel; peroxisome; protein transport; trafficking

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. Giardia is an important model for cell biology because it represents the most basal of eukaryotic lineages based on sequence analysis of conserved genes. Giardia trophozoites lack many typical eukaryotic organelles such as mitochondria, peroxisomes, and protein trafficking compartments such as a classical Golgi apparatus and secretory granules. Instead, Giardia employs a simple endomembrane system to export proteins to distinct intracellular locations. We have discovered a novel protein compartment in Giardia that appears to serve as ER and endosome. Using reporter-constructs, confocal microscopy, and ultrastructural analysis, we have shown that Giardia cysteine proteases localize to a tubulovesicular network with an ER-like structure peripheral to the perinuclear membrane. Labeled proteins are rapidly endocytosed into this compartment and degraded by these Giardia cysteine proteases. It therefore appears that Giardia contains a transitional endomembranous structure that may pre-date compartmentalization of endocytic/lysosomal functions and the endoplasmic reticulum. Giardia cysteine proteases may function to break down endocytosed nutrients. However, it remains unclear which members of the Giardia cysteine protease family are responsible for this activity. Cysteine protease activity can be isolated and purified from Giardia lysate using FPLC. This lysate fraction can then be submitted for mass spectrometry analysis to determine the gene products responsible for this novel activity.

该副本是利用众多研究子项目之一 由NIH/NCRR资助的中心赠款提供的资源。子弹和 调查员（PI）可能已经从其他NIH来源获得了主要资金， 因此可以在其他清晰的条目中代表。列出的机构是 对于中心，这不一定是调查员的机构。 贾第鞭毛虫是细胞生物学的重要模型，因为它代表了基于保守基因序列分析的真核谱系的最基础。 贾第鞭毛虫缺乏许多典型的真核细胞器，例如线粒体，过氧化物酶体和蛋白质运输室，例如经典的高尔基体和分泌颗粒。 取而代之的是，贾迪亚采用简单的内膜系统将蛋白质导出到不同细胞内的位置。 我们已经发现了贾第氏菌的一种新型蛋白质室，似乎用作ER和内体。 使用报告基因构建，共聚焦显微镜和超微结构分析，我们表明，贾第鞭毛虫的半胱氨酸蛋白酶定位于核叶膜膜上具有ER样结构的微管网络。 标记的蛋白质迅速内吞成该区室，并被这些贾第鞭毛虫的半胱氨酸蛋白酶降解。 因此，贾第鞭毛虫似乎包含一种过渡性内膜结构，该结构可能会预先对内吞/溶酶体功能和内质网的隔室化。 贾第鞭毛虫的半胱氨酸蛋白酶可能作用可分解内吞营养素。 但是，目前尚不清楚贾第鞭毛虫的半胱氨酸蛋白酶家族的哪些成员负责这项活动。 半胱氨酸蛋白酶活性可以使用FPLC分离并从贾第鞭毛虫裂解物中纯化。 然后可以将此裂解液部分提交进行质谱分析，以确定负责这种新活性的基因产物。