Transcriptional Regulators of the Malarial var Multigene Family

描述（由申请人提供）：本研究项目的目标是开发一种新方法，该方法结合染色质免疫沉淀、串联亲和纯化和质谱法，以鉴定编码疟疾毒力因子（恶性疟原虫红细胞膜蛋白1，PfEMP 1）的var多基因家族的可能激活子和/或阻遏子。在其红细胞内发育期间，疟原虫将表达转换为该变体表面蛋白的替代形式以逃避宿主免疫应答。每个恶性疟原虫的基因组包含大约60个分布在不同染色体上的var基因。然而，只有一个var基因是由每个寄生虫以互斥的方式表达。var基因的启动子区在这种调控中起着重要作用，据信这种机制可能涉及专门的阻遏物（PfVPR）和/或激活物（PfVPA）分子。目前建议的具体目标包括实验设计和方法，一些新开发的恶性疟原虫在我们的实验室，如串联亲和纯化和荧光原位杂交（FISH）。他们是：目标一。通过染色质免疫沉淀、串联亲和纯化和质谱法鉴定PfVPA和PfVPR对var基因表达的调节作用;启动恶性疟原虫PfVPA和PfVPR蛋白的功能分析。 公共卫生相关性：疟疾寄生虫侵入并最终破坏其宿主的循环红细胞，通常导致严重的临床疾病或死亡。在其红细胞内发育期间，疟原虫将表达切换为由var基因编码的变体表面蛋白PfEMP 1的替代形式，以逃避宿主免疫应答。这项研究合作的目标是将遗传学、分子生物学和免疫化学的联合收割机工具和技术以及两个实验室的专业知识结合起来，以解开疟疾感染期间抗原变异的分子决定因素。

DESCRIPTION (provided by applicant): The goal of this research project is to develop a new approach that combines chromatin immunoprecipitation, tandem affinity purification and mass spectrometry to identify possible activators and/or repressors of the var multigene family encoding the malaria virulence factor, Plasmodium falciparum erythrocyte membrane protein 1, PfEMP1. During their intraerythrocytic development, malaria parasites switch expression to alternative forms of this variant surface protein to evade the host immune response. The genome of each P. falciparum parasite contains approximately 60 var genes distributed on different chromosomes. However, only one var gene is expressed by each parasite in a mutually exclusive manner. The promoter region of var genes plays an important role in this regulation and it is believed that this mechanism might involve specialized repressor (PfVPR) and/or activator (PfVPA) molecules. The specific aims of the current proposal include experimental designs and methods, some newly developed in our laboratories for P. falciparum such as tandem affinity purification and fluorescent in situ hybridization (FISH). They are: Aim I. To identify PfVPA and PfVPR regulators of var gene expression by chromatin immunoprecipitation, tandem affinity purification and mass spectrometry; and Aim II. To initiate a functional analysis of the PfVPA and PfVPR proteins in P. falciparum. PUBLIC HEALTH RELEVANCE: Malaria parasites invade and ultimately destroy circulating red blood cells of their host, often leading to severe clinical illness or death. During their intraerythrocytic development, malaria parasites switch expression to alternative forms of a variant surface protein PfEMP1 encoded by the var genes to evade the host immune response. The goal of this research collaboration is to combine tools and techniques from genetics, molecular biology, and immunochemistry, and the expertise of two laboratories to unravel the molecular determinants of antigenic variation during malaria infection.