LEISHMANIA-MACROPHAGE CELLULAR INTERACTIONS IN VITRO

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 随后在这些细胞中分化、存活和繁殖， 吞噬细胞 了解生物化学和分子 这种宿主-寄生虫细胞相互作用的机制将 为开发更有效的化疗和/或免疫疗法提供线索。 治疗和预防利什曼病。 利什曼原虫主要表面糖蛋白（gp 63）是从 前鞭毛体，并发现是一种金属蛋白酶活性在酸性 博士 它似乎介导利什曼原虫与 巨噬细胞和保护寄生虫从降解， 吞噬溶酶体 利什曼原虫毒力表型与 DNA标记的gp 63和衣霉素抗性的丰度 扩增和衣霉素敏感的N- 乙酰葡糖胺-1-磷酸转移酶（NAGT）在长萜醇中 通路 这是上述研究细胞的工作的继续 gp 63毒力的生物学、生物化学和分子生物学 N-糖基化对其功能的调控 墨西哥利什曼原虫（Leishmania mexicana spp.）如下：（1）进一步 gp 63作为用于显影的金属蛋白酶的表征 特异性抑制剂;（2）无鞭毛体gp 63的纯化 研究其蛋白酶活性;（3）了解其蛋白酶活性。 gp 63在吞噬溶酶体中保护作用的研究 巨噬细胞与gp 63包被的脂质体的相互作用;（4） gp 63的胞内通路分析， 聚糖和利什曼原虫NAGT，以了解 gp 63毒力蛋白功能中的N-糖基化 （5）NAGT和其他潜在的表征 在衣霉素抗性细胞的扩增DNA中的毒力基因 表达载体中的分子克隆。 这些研究的结果不仅有助于我们理解 利什曼原虫在巨噬细胞内寄生中的毒力 还包括金属蛋白酶、糖蛋白和多萜醇途径， 将军

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.