INSULIN-LIKE SIGNALING IN PARASITIC NEMATODE DEVELOPMENT

描述(由申请人提供)：寄生线虫感染了世界上相当大比例的人口，给人类疾病造成了巨大的损失。这些寄生虫以发育受阻的感染性幼虫(通常是第三阶段幼虫，L3I)的形式感染宿主，一旦进入宿主，就会恢复发育。借鉴秀丽隐杆线虫的大量相关发现，我们发现了明确的证据，即在寄生线虫Strongyloids stercoris感染之前和感染过程中，胰岛素样(ILS)和类固醇核激素受体(NHR)信号调节L3I的发育。在这一新的应用中，我们建议完成我们关于ILS对Sstercoris L3I发育的调控的研究，并开始新的研究，以保守的类固醇NHR信号通路作为这种寄生虫的治疗靶点。我们的两个特定目标中的第一个是询问胰岛素信号是否调节胸锁螺旋体L3I的形成和维持。具体地说，我们将确定胰岛素样受体激酶SS-DAF-2和PI3激酶SS-AGE-1是否阻断L3I的发育再激活并促进其在宿主中的发育再激活，以及胰岛素样多肽SS-ILP-6和SS-ILP-7是否分别促进L3I的发育再激活和抑制。我们的方法将包括分析由于表达基于这些分子的主导转基因结构而导致的表型。具体目标2将询问，通过SS-DAF-12 NHR的信号是否增强了司体珊瑚中ILS的调节作用，以及这种小分子受体是否可能成为基于自然产生的类固醇或其类似物的化疗靶点。我们建议鉴定SS-DAF-12的天然配体，并将它们与线虫的DAF-12配体、D4-和D7-Dafachronic酸(DA)在调节L3I抑制和重新激活方面的活性进行比较。我们将使用化学抑制剂来探索内源性NHR DAF-12在斯特氏链球菌中的功能，并使用沙土鼠感染模型来确定应用SS-DAF-12抑制剂或DA是否可以阻止L3I的发展，清除成虫感染和/或阻断暴发性自身感染，暴发性自身感染是人类弓形虫病的一种潜在致命并发症。这些实验将直接反映DAF-12 NHR作为寄生线虫化疗靶点的潜力。

DESCRIPTION (provided by applicant): Parasitic nematodes infect a significant proportion of the world's population and exact an enormous toll of human illness. These parasites infect their hosts as developmentally arrested infective larvae (usually third-stage larvae, (L3i) that resume development once they enter the host. Drawing upon extensive relevant findings in Caenorhabditis elegans, we have uncovered definitive evidence that insulin-like (ILS) and steroid-nuclear hormone receptor (NHR) signaling regulate L3i development before and during the infective process in the parasitic nematode Strongyloides stercoralis. In this renewal application, we propose to complete our studies on regulation of L3i development by ILS in S. stercoralis and initiate new research on the potential of a conserved steroid NHR signaling pathway as a therapeutic target in this parasite. The first of our two Specific Aims asks whether insulin signaling regulates formation and maintenance of L3i by S. stercoralis. Specifically we will determine whether the insulin-like receptor kinase Ss-DAF-2 and the PI3 kinase Ss-AGE-1 block arrest of L3i and promote their developmental reactivation in the host and whether insulin-like peptides Ss-ILP-6 and Ss-ILP-7 promote developmental reactivation and arrest of L3i, respectively. Our approach will involve analyzing phenotypes that result from expressing dominant transgene constructs based on these molecules in S. stercoralis. Specific Aim 2 will ask whether signaling through the Ss-DAF-12 NHR augments regulatory ILS effects in S. stercoralis and whether this small molecule receptor could be a target for chemotherapy based on naturally occurring steroids or their analogs. We propose to identify the natural ligands of Ss-DAF-12 and compare their activity in regulating L3i arrest and reactivation to the DAF-12 ligands from C. elegans, D4- and D7-dafachronic acids (DA). We will use chemical inhibitors to probe endogenous NHR DAF-12 function in S. stercoralis and use a gerbil model of infection to determine whether administered Ss-daf-12 inhibitors or DA can prevent development of L3i, clear adult worm infections and/or block fulminant autoinfection, a potentially fatal complication of human strongyloidiasis. These experiments will directly reflect the potential of DAF-12 NHR function as a chemotherapeutic target in parasitic nematodes.