CELL AND DEVELOPMENTAL BIOLOGY OF TRYPANOSOMATID PARASITES

利什曼原虫是专性胞内原生动物 人类的病原体。在受感染的患者体内，各种不同种类的 这种微生物栖息并破坏皮肤或皮肤内的巨噬细胞 内脏(即脾、肝和骨髓)。因此，他们 导致溃烂、无法愈合、毁容的皮肤损害 (例如墨西哥乳杆菌)或退化的，通常是致命的内脏 疾病(例如，多诺氏乳杆菌)。这些疾病折磨着1200多万人。 世界各地热带和新热带地区的患者。我们的研究是 旨在确定病理生理学中涉及的机制 这种有机体的。在这方面，基本细胞、分子和 利什曼原虫及相关锥虫的发育生物学 原生动物的研究是为了识别和表征 寄生虫分子对这些生物的生存至关重要 人类病原体。这些寄生虫是如何生存的，获得 营养物质，在它们的昆虫媒介内繁殖和分化 哺乳动物宿主是理解基本问题的核心 这些有机体的寄生性质和进化适应。 由于这些寄生虫与它们的宿主直接互动，因此了解 其表面膜的组成和功能似乎 必不可少的。为此，独特的寄生虫表面膜酶 和调节蛋白的鉴定和生化分析 以确定它们在生物生存中的功能作用为特征的 这些有机体。此外，编码这种蛋白质的基因是 第一次被孤立和表征，朝着定义 它们在寄生虫生长过程中的表达和调控， 差异化发展。在过去一年里，我们的研究 已经阐明了几个基因结构和染色体位点。 不同成员独特的锥虫表面膜 3？-L核酸酶/核酸酶家族 光亮克里迪亚线虫。这些基因的各种突变表达结构 基因被制造并用来识别这些基因的特定区域 负责它们的酶活性和它们的 适当地定位于寄生虫细胞表面。其中的几个 突变的表达式-构造也被证明对 实验室规模的生产和提纯这些通常 膜结合的可溶形式的寄生虫酶。类似 对独特的利什曼伴侣的研究正在进行中 蛋白质、LD-钙网蛋白和表膜酸性磷酸酶。 此外，涉及这些基因的定向删除的实验 而功能零突变寄生虫的产生在 进步。这些突变寄生虫的生存能力将在 SITE。这类研究的结果应该证明， 事实上，由这些基因编码的蛋白质对于 这些病原体在其昆虫媒介和/或 哺乳动物的宿主。因此，目前的研究旨在确定 这些生物体的内在需求，并确定它们的关键 大自然。我们最近和正在进行的研究的结果继续 提供相关信息以了解独特的 这些生物体的病理生理学。此外，这些研究还包括 对于证明这种寄生虫是否 酶和调节蛋白是设计的合乎逻辑的目标 2)新的化疗药物的开发 诊断工具。它们也与确定是否 它们作为针对这些人类病原体的潜在疫苗是有用的。

Leishmania are obligate intracellular protozoan pathogens of humans. Within infected patients, various species of this organism inhabit and destroy macrophages within the skin or internal organs (i.e., spleen, liver and bone marrow). Thus, they cause either ulcerative, non-healing, disfiguring cutaneous lesions (e.g., L. mexicana) or degenerative and most often fatal visceral disease (e.g., L. donovani ). These diseases afflict over 12 million patients in the Tropics and Neo-tropics worldwide. Our studies are aimed at defining the mechanisms involved in the pathophysiology of this organism. In that regard, the basic cell, molecular and developmental biology of Leishmania and related trypanosomatid protozoa are investigated toward identifying and characterizing parasite molecules which are essential for the survival of these human pathogens. How these parasites are able to survive, access nutrients, multiply and differentiate within their insect vector and mammalian hosts are questions central to understanding the basic parasitic nature and evolutionary adaptations of these organisms. Since these parasites interact directly with their hosts, knowledge of the composition and functions of their surface membranes seems essential. To that end, unique parasite surface membrane enzymes and regulatory proteins are identified and biochemically characterized to determine their functional roles in the survival of these organisms. Further, the genes encoding such proteins are being isolated and characterized, for the first time, toward defining their expression and regulation during course of parasite growth, differentiation and development. During the past year, our studies have elucidated the gene structure and chromosomal loci of several different members of the unique trypanosomatid surface membrane 3?-nucleotidase/ nuclease family from L .donovani, L. mexicana and Crithidia luciliae. Various mutated-expression constructs of these genes were made and used to identify the specific regions of these proteins responsible for their enzymatic activities and for their proper targeting to the parasite cell surface. Several of these mutated expression-constructs have also proven useful for the laboratory-scale production and purification of these normally membrane-bound parasite enzymes in a soluble form. Similar studies are in progress with the unique leishmanial chaperone protein, Ld-calreticulin and surface membrane acid phosphatase. Further, experiments involving the targeted deletion of these genes and the production of functionally null mutant parasites are in progress. The viability of such mutant parasites will be tested in situ. Results of such studies should demonstrate whether the proteins encoded by these genes are, in fact, essential for the survival of these pathogens within their insect-vector and/or mammalian hosts. Thus, the current studies are aimed at defining the intrinsic needs of these organisms and determining their critical nature. Results of our recent and ongoing studies continue to provide pertinent information toward understanding the unique pathophysiology of these organisms. In addition, these studies are of practical relevance toward demonstrating whether such parasite enzymes and regulatory proteins are logical targets for 1) the design of new chemotherapeutic drugs and 2) the development of new diagnostic tools. They are also relevant for determining whether they are useful as potential vaccines against these human pathogens.