Molecular Studies of Immunoparasitology in Snails

描述(申请人提供)：淡水蜗牛在人类血吸虫曼氏血吸虫的传播中起着不可或缺的作用。增加我们对这种蜗牛体内防御的基本机制的了解，可以帮助开发控制血吸虫病的新方法。血吸虫病目前仍在全世界感染2亿人。关于血吸虫和它们的钉螺宿主之间的分子相互作用仍有许多未知之处，而光滑杆菌-曼氏杆菌模型系统是迄今为止评估潜在的抗性和亲和性机制的最佳模型。这项拟议的研究建立在之前的研究基础上，这些研究表明蜗牛血蛋白FREPs(纤维蛋白原相关蛋白)在抗血吸虫防御中具有功能作用。他们进一步受到我们的发现的启发，这些发现表明，光滑假单胞菌通过点突变和重组产生了令人惊讶的多样性的FREP基因，这一发现与其他实验室的工作一起，引发了对无脊椎动物产生多样化防御反应能力的全面重新考虑。在这项申请中，我们开发了一种通用的方法来评估假定的免疫效应器(FREP是主要的，但不是唯一的，可以使用这种方法进行测试的效应器)在影响曼氏葡萄球菌在光滑芽孢杆菌中成功发育的能力方面的功能。在这样做的时候，重要的是要从参与抵抗感染的角度和作为亲和性的一般决定因素来检查候选效应分子。具体地说，我们将进一步开发RNA干扰(RNAi)和重组蛋白/抗体方法，使用体内和体外方法来研究FREPs在影响不相容宿主中寄生虫发展方面的功能作用。我们最近确定的其他候选基因，即革兰氏阴性细菌结合蛋白(GNBP)和肽聚糖识别蛋白(PGRPs)，也可以用于研究，这是控制Toll/IMD上游通路的关键分子的蜗牛同源物，从而进一步拓宽了我们对钉螺免疫的理解。最后，我们将尝试破坏曼氏链球菌对相容蜗牛的成功寄生。实现这种破坏的能力将是一项重要的进步，因为兼容性确保了血吸虫病在流行地区的持续传播。这项研究将使我们能够对FREP和其他候选分子的功能获得有价值的见解，更重要的是，开发一个普遍适用的系统，以评估任何蜗牛候选分子在影响曼氏螺旋体与其钉螺宿主相遇的结果中所起的作用。

DESCRIPTION (provided by applicant): The freshwater snail Biomphalaria glabrata plays an indispensible role in transmission of the human blood fluke, Schistosoma mansoni. Increasing our understanding of the fundamental mechanisms of internal defense of this snail could help develop novel methods of control of schistosomiasis which still infects 200 million people world-wide. Much remains unknown regarding the molecular interplay between schistosomes and their snail hosts, and the B. glabrata - S. mansoni model system is by far the best for assessing underlying mechanisms of resistance and compatibility. The proposed investigations are built on previous studies suggestive of a functional role of snail blood proteins called FREPs (fibrinogen-related proteins) in anti-schistosome defenses. They are further inspired by our discoveries showing that B. glabrata generates a surprising diversity of FREP genes through point mutation and recombination, a discovery that along with work in other labs has provoked a general reconsideration of the capacity of invertebrates to generate diversified defense responses. In this application, which has been extensively revised in response to the previous review, we develop a general approach for assessing the functionality of putative immune effectors (FREPs are the main, but not the sole, effectors available to be tested using this approach) in influencing the ability of S. mansoni to develop successfully in B. glabrata. In so doing, it is important to examine candidate effector molecules both from the perspective of their involvement in resistance to infection, and as general determinants of compatibility. Specifically, we will further develop RNA interference (RNAi) and recombinant protein/antibody approaches to examine the functional role of FREPs in affecting the development of parasites in incompatible hosts, using both in vivo and in vitro approaches. Other candidates we have recently identified, namely gram-negative bacteria binding protein (GNBP) and peptidoglycan recognition proteins (PGRPs), the snail homologs of key molecules controlling upstream Toll/Imd pathways, are also available for study, thus further broadening our understanding of snail immunity. Finally, we will attempt to disrupt the successful parasitization of compatible snails by S. mansoni. The ability to effect such disruption would be an important advance because compatibility ensures the continued transmission of schistosomiasis in endemic areas. This study will enable us to gain valuable insights into the functionality of FREPs and other candidates, and even more importantly, to develop a system of general applicability for evaluating the role of any snail candidate molecules in influencing the outcome of an encounter between S. mansoni and its snail host.