MECHANISMS OF MALARIA SEXUAL STAGE GENE EXPRESSION

恶性疟原虫(简称恶性疟原虫)的生活史以明显的形态变化为特征。这些变化与寄生虫表面独特蛋白质的阶段特异性表达有关。针对受精卵和卵子主要表面蛋白的抗体Pfs25和Pfs28分别阻止了蚊子中肠寄生虫的形态发育。这表明这些蛋白的表达对性阶段的发育至关重要。这一信息已被用于制定传播阻断疫苗。然而，由于对寄生虫高度富含AT的基因间隔区进行测序的技术困难，以及无法对推测的基因调控元件进行功能性测试，对这些关键蛋白的有性阶段特异性表达的分子机制的定义一直受到阻碍。疟疾基因组计划的成功和疟疾寄生虫转基因技术的发展现在使人们能够描绘特定有性阶段基因表达的分子机制。了解恶性疟原虫有性期基因表达的基本机制可以为阻断疾病传播提供新的策略或方法。我们利用鸡疟原虫有性期的基因导入系统作为一个模型系统来确定进化上密切相关的恶性疟原虫的基因调控元件。对Pfs28，Pgs28的鸡疫霉同源物的研究表明，Pgs28蛋白的表达需要一个独特的富含T的元件，并结合pgs28 3‘UTR中的一个真核多聚腺苷化共识信号。在其他与发育相关的真核基因中，富含T的3‘基因侧翼元件的功能支持了Pfs25和Pfs28的协同表达是由3’基因侧翼调控元件介导的假说。本项目的具体目标是：1)确定富含T的元件调节Pgs28表达的分子机制；2)确定Pfs25和Pfs28协调表达的分子机制；3)确定3‘基因侧翼元件在介导性阶段特异性基因表达中的作用。

The life cycle of Plasmodium falciparum (P. falciparum) is characterized by pronounced morphologic changes. These changes are associated with the stage-specific expression of unique proteins on the parasite's surface. Antibodies to the major surface proteins of fertilized zygotes and ookinetes, Pfs25 and Pfs28 respectively, block the morphologic development of the parasite in the mosquito midgut. This indicates that the expression of these proteins is critical to the development of the sexual stages. This information has been utilized in the formulation of transmission blocking vaccines. Definition of the molecular mechanisms responsible for the sexual stage-specific expression of these critical proteins, however, has been hampered by technical difficulties in sequencing the parasite's highly AT- rich intergenic regions and the inability to functionally test putative gene regulatory elements. The success of the malaria genome project and the development of transfection techniques for the malaria parasite now permit the delineation of the molecular mechanisms of sexual stage-specific gene expression. Understanding the basic mechanisms of P. falciparum sexual stage gene expression could suggest novel strategies or methods to block disease transmission. We have utilized a gene transfection system of the sexual stages of the chicken malaria Plasmodium gallinaceum (P. gallinaceum) as a model system for defining the gene regulatory elements of the evolutionarily closely related P. falciparum parasite. Studies of the P. gallinaceum homologue of Pfs28, Pgs28, reveal that a unique T-rich element in conjunction with a eukaryotic polyadenylation consensus signal in the pgs28 3' UTR are required for Pgs28 protein expression. The function of T-rich 3' gene flanking elements in other eukaryotic genes associated with development supports the hypothesis that the coordinated expression of Pfs25 and Pfs28 is mediated by 3' gene flanking regulatory elements. The specific aims of this project are: 1) to determine the molecular mechanism by which the T-rich element regulates Pgs28 expression; 2) to define the molecular mechanisms responsible for the coordinated expression of Pfs25 and Pfs28; 3) to determine the contribution of 3' gene flanking elements in mediating sexual stage-specific gene expression.