ROLE OF PFHO-1 IN P. FALCIPARUM INTRAERYTHROCYTIC DEVELOPMENT

描述(申请人提供)：控制恶性疟原虫红细胞内生长发育的探索还处于初级阶段。了解寄生虫进展的信号和效应器将增强我们对这种寄生虫如何在宿主细胞中生存的知识，并将发现新的化疗靶点。PfHO-1是一种功能未知的血红素加氧酶样蛋白，在红细胞内疟原虫中表达。我们发现PfHO-1具有与卟啉反应的DNA结合活性，并且优先与参与蛋白质合成的基因相关联。我们的假设是，PfHO-1对血红蛋白降解引起的血红素积累做出反应，对红细胞内恶性疟原虫的发展至关重要。这里提出的这项研究的目标是检验这一假说。我们将评估PfHO-1在红细胞内恶性疟原虫和大肠杆菌报告系统中对细胞内卟啉水平扰动的反应。我们将通过一种新的双阳性选择基因敲除方法和有条件地表达一种对卟啉无反应的蛋白来削弱PfHO-1的作用。这些研究将使我们能够确定PfHO-1是否在检测细胞卟啉水平和参与细胞内稳态的关键方面发挥重要作用。拟议的实验还将开发新的卟啉操纵技术，以及在反向基因工程寄生虫中选择有害表型的技术。

DESCRIPTION (provided by applicant): Exploration of the control of Plasmodium falciparum intraerythrocytic growth and development is in its infancy. Understanding the signals and effectors of parasite progression will enhance our knowledge of how this parasite survives in its host cell and will uncover new targets for chemotherapy. PfHO-1 is a heme oxygenase-like protein of unknown function expressed in intraerythrocytic malaria parasites. We have found that PfHO-1 has a porphyrin-responsive DNA- binding activity and associates preferentially with genes that are involved in protein synthesis. Our hypothesis is that PfHO-1 responds to heme accumulation from hemoglobin degradation and is crucial for intra-erythrocytic P. falciparum development. The goal of the study proposed here is to test this hypothesis. We will assess the response of PfHO-1 to perturbation of intracellular porphyrin levels in intraerythrocytic P. falciparum and in an E. coli reporter system. We will impair PfHO-1 action by a new double-positive selection gene knockout approach and by conditional expression of a porphyrin-unresponsive version of the protein. These studies will allow us to establish whether PfHO-1 plays an important role in sensing cellular porphyrin levels and participating in critical aspects f cellular homeostasis. The proposed experiments will also develop new technologies for porphyrin manipulation and for selection of deleterious phenotypes in reverse genetically engineered parasites.