MOLECULAR DISSECTION OF P. FALCIPARUM SEXUAL DEVELOPMENT

疟疾寄生虫每年造成3亿至5亿人感染和200万至300万人死亡。疟疾在脊椎动物宿主之间的传播涉及按蚊媒介中预定的性发育周期。性阶段对于疟疾传播是绝对必要的。恶性疟原虫性别分化和发育的分子机制仍然很大程度上不清楚。在最初进入有性周期后，疟原虫配子体经历了一个似乎是多步骤的生长和发育过程，可能涉及几个基因产物。这项建议的目的是研究两种蛋白(Pfg27和Pfs16)的功能参与，这两种蛋白在配子体发生早期大量表达。我们最近通过同源重组破坏了Pfg27基因，导致转化后的寄生虫失去了有性表型。这些研究首次表明，Pfg27是一种在配子体第一和第二阶段表达的蛋白质，对配子体发育至关重要。对Pfs16干扰的研究将进一步阐明恶性疟原虫的性别分化过程。对稳定转化的寄生虫的研究也将为评估转录调控机制、启动子的有性阶段特异性和中断基因的互补提供机会。将使用免疫化学和分子(酵母双杂交)相结合的方法来研究涉及Pfg27和Pfs16以及其他细胞蛋白的蛋白质-蛋白质相互作用。这些研究可能确定新的蛋白作为相互作用的功能伙伴，从而促进我们对Pfg27和Pfs16在有传播能力的寄生虫的性别分化和发育中的生物学作用的理解。因此，这些研究将提供一种新的分子遗传学方法来剖析恶性疟原虫的性发育机制。这些信息可导致合理发展免疫和/或化疗武器库，以防止恶性疟原虫的传播。

Malaria parasites are responsible for 300-500 million infections and 2-3 million deaths annually. Transmission of malaria between vertebrate hosts involves an obligate sexual developmental cycle in the anopheline mosquito vector. The sexual stages are absolutely essential for malaria transmission. The molecular mechanisms underlying sexual differentiation and development in Plasmodium falciparum remain largely unknown. After the initial commitment to the sexual cycle, Plasmodium gametocytes undergo what appears to be a multi-step growth and development process, possibly involving several gene products. The goal of this proposal is to investigate the functional involvement of two proteins (Pfg27 and Pfs16), abundantly expressed early during gametocytogenesis. We have recently disrupted the gene for Pfg27 by homologous recombination, resulting in the loss of sexual phenotype in the transformed parasites. These studies for the first time have shown that Pfg27, a protein expressed in stage I and II gametocytes is critical for gametocyte development. Studies on Pfs16 disruption will further elucidate sexual differentiation process in P. falciparum. Investigations on stably transformed parasites will also offer an opportunity to evaluate transcriptional control mechanisms, sexual stage- specificity of promoters and complementation of disrupted genes. A combination of immunochemical and molecular (yeast-two-hybrid) approaches will be employed to investigate protein-protein interactions involving Pfg27 and Pfs16 and other cellular proteins. These studies could possibly identify novel proteins as interacting functional partners, thus advancing our understanding of the biological roles of Pfg27 and Pfs16 in the sexual differentiation and development of transmission competent parasites. The proposed studies will thus offer a novel molecular genetic approach to dissect mechanisms underlying sexual development in P. falciparum. Such information could lead to rational development of immunological and/or chemotherapeutic arsenal to prevent transmission of P. falciparum malaria.