Var gene regulation and antigenic variation in malaria

描述（由申请人提供）：疟疾仍然是发展中国家发病率和死亡率的主要原因之一。这种疾病是由原生动物寄生虫引起的，它们侵入并最终破坏宿主的循环红细胞，导致严重贫血和经常致命的脑疟疾综合征。这些寄生虫已经进化出了一种复杂的免疫逃避机制，在感染过程中，出现了抗原表型改变的寄生虫小亚群，从而避免了宿主的抗体反应。这一抗原变异过程是导致疾病持续存在的原因，也是在感染恶性疟原虫的个体中经常观察到的寄生虫病浪潮的原因。抗原表型的变异是多拷贝var基因家族个体成员之间表达开关的结果。这个家族由大约40-50个基因组成，这些基因编码主要的抗原决定因子，一种称为PIEMPI的蛋白质。任何寄生虫一次只表达一种变异基因，因此决定了被感染细胞的抗原类型。var基因表达的变化和由此产生的抗原变异似乎在转录水平上受到控制。这一建议的目的是确定分子机制，维持所有的，但一个var基因在转录沉默状态。待验证的假设是var启动子通过凝聚异色状态的组装而沉默，并且染色质修饰和重塑调节恶性疟原虫var基因的表达和抗原变异。实验设计利用了最近的发现，var启动子上游的元件与所有var基因中发现的保守内含子以合作方式起作用，以沉默除单个基因外的所有基因的转录。这种转录沉默状态可以在含有报告基因、var启动子和内含子的转染片段上组装。利用这一系统，将确定沉默和活跃var启动子的染色质结构和亚核定位。此外，还将鉴定var基因沉默和激活所必需的特定DNA元件，并阐明DNA元件作为沉默者或激活者的序列要求。

DESCRIPTION (provided by the applicant): Malaria remains one of the leading causes of morbidity and mortality in the developing world. The disease is caused by protozoan parasites that invade and ultimately destroy circulating red blood cells of their host, leading to severe anemia and the frequently lethal syndrome of cerebral malaria. These parasites have evolved a complex mechanism of immune evasion whereby, over the course of an infection, small sub-populations of parasites arise that have an altered antigenic phenotype, thus avoiding the antibody response of the host. This process of antigenic variation and is responsible for the persistent nature of the disease as well as the waves of parasitemia frequently observed in individuals infected by P. falciparum. The variation in antigenic phenotype is the result of switches in expression between individual members of the multicopy var gene family. This family consists of approximately 40-50 genes that encode the predominant antigenic determinant, a protein called PIEMPI. Only a single var gene is expressed at a time by any given parasite, thus determining the antigenic type of the infected cell. Changes in var gene expression and the resulting antigenic variation appear to be controlled at the level of transcription. The objective of this proposal is to determine the molecular mechanisms that maintain all but a single var gene in a transcriptionally silent state. The hypothesis to be tested is that var promoters are silenced through the assembly of a condensed, heterochromatic state and that chromatin modification and remodeling regulates expression of var genes and antigenic variation in Plasmodium falciparum. The experimental design exploits the recent discovery that elements upstream of var promoters act in a cooperative fashion with a conserved intron found in all var genes to silence transcription of all but a single gene. This transcriptionally silent state can be assembled on transfected episomes containing a reporter gene flanked by a var promoter and intron. Using this episomal system, the chromatin structure and subnuclear localization of silent and active var promoters will be determined. In addition, the specific DNA elements necessary for var gene silencing and activation will be identified, and the sequence requirements for DNA elements to functions as silencers or activators elucidated.