LEISHMANIA-MACROPHAGE CELLULAR INTERACTIONS IN VITRO

利什曼原虫-巨噬细胞体外相互作用

• 批准号: 3130195
• 负责人: Kwang Poo Chang
• 金额: $ 23.62万
• 依托单位: ROSALIND FRANKLIN UNIV OF MEDICINE & SCI
• 依托单位国家: 美国
• 财政年份: 1983
• 资助国家: 美国
• 起止时间: 1983-04-01 至 1994-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3130195
• 关键词: Leishmania major; chromatography; dolichol; electron microscopy; electrophoresis; genetic manipulation; genetic translation; glycoproteins; glycosylation; glycosyltransferase; hamsters; host organism interaction; laboratory mouse; leishmaniasis; liposomes; lysosomes; macrophage; messenger RNA; microorganism culture; microorganism immunology; molecular cloning; natural gene amplification; peptidases; phagocytes; protein purification; protein transport; protozoal antigen; tissue /cell culture; transposon /insertion element; virulence

Our long term objectives are to elucidate the questions of how Leishmania infect macrophages, and how these parasites subsequently differentiate, survive and multiply in these phagocytes. Understanding the biochemical and molecular mechanisms of such host-parasite cellular interactions will provide leads for developing more effective chemo and/or immuno- therapy and -prophylaxis for leishmaniasis. A Leishmania major surface glycoprotein (gp63) was purified from promastigotes and found to be a metallo-protease active at acidic pH. It appears to mediate the binding of Leishmania to macrophages and to protect parasites from degradation in phagolysosomes. Leishmania virulent phenotype is associated with the abundance of gp63 and tunicamycin-resistance, marked by DNA amplification and an increased level of tunicamycin-sensitive N- acetylglucosamine-1-phosphate transferase (NAGT) in the dolichol pathway. This is a continuation of above-mentioned work to study cell biology, biochemistry and molecular biology of gp63 as a virulent determinant and the regulation of its function by N-glycosylation in Leishmania mexicana spp. as follows: (1) Further characterization of gp63 as a metallo-protease for developing specific inhibitors; (2) Purification of gp63 from amastigotes for studying its protease activity; (3) Understanding the protective functions of gp63 in phagolysosomes by studying interactions of macrophages with gp63-coated liposomes; (4) Analyses of intracellular routing of gp63, the structure of its glycans and Leishmania NAGT to understand the regulatory role of N-glycosylation in the function of gp63 as a virulent determinant; (5) Characterization of NAGT and other potential virulent genes in amplified DNA of tunicamycin-resistant cells by molecular cloning in expression vectors. The results of these studies will help not only our understanding leishmanial virulence in intracellular parasitism of macrophages but also metallo-protease, glycoprotein and dolichol pathway in general.

我们的长期目标是阐明如何 利什曼原虫感染巨噬细胞，以及这些寄生虫如何 随后区分，生存和繁殖 吞噬细胞。 了解生化和分子 这种宿主 - 寄生虫相互作用的机制将 提供潜在客户，以开发更有效的化学疗法和/或免疫 - 利什曼病的疗法和 - 促囊肿。 利什曼原虫主要表面糖蛋白（GP63）从中纯化 前寄生物，发现在酸性上活跃于金属蛋白酶 ph。 它似乎调解了利什曼尼亚对 巨噬细胞并保护寄生虫免受降解 吞噬体。 Leishmania有毒表型与 以DNA为标志 扩增和对穿刺霉素敏感的N-的水平增加 乙酰葡萄糖1-磷酸转移酶（NAGT） 路径。 这是研究细胞的上述工作的延续 GP63的生物学，生物化学和分子生物学 决定因素及其功能通过n-糖基化对其功能 在墨西哥利什曼尼亚属。如下：（1）进一步 GP63作为用于开发的金属蛋白酶的表征 特定的抑制剂； （2）从amastigotes纯化GP63 用于研究其蛋白酶活性； （3）理解 通过研究 巨噬细胞与GP63涂层脂质体的相互作用； （4） GP63的细胞内路由的分析，其结构 Glycans和Leishmania Nagt了解 gp63功能中的N-糖基化作为毒 决定因素； （5）表征NAGT和其他潜力 在皮承霉素细胞的扩增DNA中的强毒基因通过 表达载体中的分子克隆。 这些研究的结果不仅将有助于我们的理解 巨噬细胞细胞内寄生虫中的利什人类毒力 但也有金属蛋白酶，糖蛋白和多利果途径 一般的。