Toxoplasma gondii GRA protein function

描述(申请人提供)：致病真核生物弓形虫在免疫抑制的背景下引起特别严重的机会性感染。这些感染是由慢性/潜伏组织囊重新激活引起的，这些囊通常位于大脑皮质或中枢神经系统。复发感染往往是致命的，因为现有的治疗急性弓形虫感染的药物并不理想。流行病学研究还将慢性/潜伏的弓形虫感染与神经系统疾病(如精神分裂症)联系起来，这些成年人本来是健康的免疫能力强的成年人。因此，根除慢性/潜伏的弓形虫感染为预防免疫抑制引起的复发感染以及减少具有免疫能力的个人的潜在疾病负担提供了一种创新的方法。不幸的是，目前针对急性感染的现有治疗方法对确定慢性/潜伏感染的寄生虫组织囊肿期几乎没有影响。确定能够采用创新疗法消除慢性/潜伏组织囊肿的新药靶点将在这一领域产生重大影响。在我们对寄生虫生物学的知识中存在着一个主要的缺口，这是导致 以及维持组织囊肿。对于口腔感染性所需的组织囊肿生物学特征的理解也存在很大差距。GRA蛋白是与寄生性液泡和组织囊泡相关的重要蛋白质。GRA蛋白是在形成包囊的顶端复合体(弓形虫、新孢子虫、肉孢子虫)中唯一发现的，是可以选择性靶向的潜在的新药物靶点。我们假设，与寄生虫液泡的液泡内膜网络(IVN)相关的GRA蛋白提供了囊性发育和口腔感染性所需的关键生物学功能。我支持这一假设，我们最近报道了IVN的两个主要致密颗粒(GRA)蛋白，GRA4和GRA6，在体内正常的囊负荷的发展中是必需的。IVN相关蛋白在口腔感染和传播中的作用是假设的。具体目标1将检验IVN相关GRA蛋白(GRA2、GRA3、GRA4、GRA6、GRA7和GRA9)在体内分化、囊变和/或囊维持中所必需的假说。IVN组织GRA2蛋白的两亲性α螺旋的作用(S)将通过检查GRA2的突变版本来确定。具体目标2将检验IVN相关GRA蛋白是口腔感染性和传播性所必需的假设。总而言之，这些探索性的高影响目标将从功能上确定IVN相关的GRA蛋白在组织囊肿的发展和口腔传播中所起的重要作用。这些目标将确定和验证创新干预措施的新药物目标，以根除已经感染的个人的慢性/潜伏感染。因此，这一探索性的R21项目将通过提供重要的遗传工具和试剂，以及通过提供关于药物靶标发现的高影响生物学信息来指导未来研究的方向，从而在该领域产生持久的影响。

DESCRIPTION (provided by applicant): The pathogenic eukaryote Toxoplasma gondii causes particularly severe opportunistic infections in the context of immune suppression. These infections arise by the reactivation of chronic/latent tissue cysts, typically residing in the brai or CNS. Recrudescent infections are frequently fatal because the available drug treatments for acute T. gondii infection are suboptimal. Epidemiological studies have also linked chronic/latent T. gondii infections with neurological conditions such as schizophrenia in otherwise healthy immune competent adults. Thus, eradicating chronic/latent T. gondii infection provides an innovative approach to prevent recrudescent infections caused by immune suppression, as well as to reduce the potential of disease burden in immune competent individuals. Unfortunately, currently available treatments that target acute infection have little impact on the parasite tissu cyst stages that define chronic/latent infection. The identification of new drug targets that enabl innovative therapies to eliminate chronic/latent tissue cysts would have a major impact in the field. A major gap exists in our knowledge of parasite biology that is responsible for establishing and maintaining tissue cysts. There is also a major gap in understanding of the biological features of tissues cysts that are required for oral infectivity. GRA proteins are prominent proteins associated with the parasitophorous vacuole and the tissue cyst. GRA proteins are found uniquely in cyst-forming apicomplexans (Toxoplasma, Neospora, Sarcocystis) and represent potential novel drug targets that can be selectively targeted. We hypothesize GRA proteins that associate with the intravacuolar membranous network (IVN) of the parasitophorous vacuole provide critical biological functions required for cyst development and oral infectivity. I support of this hypothesis, we recently reported that two major dense granule (GRA) proteins of the IVN, GRA4 and GRA6, were required for development of normal cyst burdens in vivo. A role for IVN associated proteins in oral infectivity and transmission is hypothesized. Specific aim 1 will examine the hypothesis that IVN associated GRA proteins (GRA2, GRA3, GRA4, GRA6, GRA7, and GRA9) are required for differentiation, cyst development, and/or cyst maintenance in vivo. The role(s) of the IVN organizing amphipathic alpha helices of the GRA2 protein will be established by examining mutant versions of GRA2. Specific aim 2 will examine the hypotheses that IVN associated GRA proteins are required for oral infectivity and transmission. Collectively, these exploratory high impact aims will functionally identify essential roles that IVN associated GRA proteins play in the development of tissue cysts and in oral transmission. These aims will identify and validate new drug targets for innovative interventions to eradicate chronic/latent infection in already infected individuals. Consequently, this exploratory R21 project will have a lasting impact in the field by providing significant genetic tools and reagents, as well as by providing high impact biological information regarding drug target discovery that will guide the direction of future studies.