Identification and characterization of type III secretion system (T3SS) effectors in Edwardsiella tarda

水产养殖在加拿大是一个价值10亿美元的行业，在许多农村和沿海社区提供了大约1.5万个全职高薪工作。2011年，加拿大是全球第五大海产品出口国。水产养殖业是加拿大经济稳定和强劲的重要贡献者；它还为加拿大人口提供食品安全和健康食品。不幸的是，鱼类疾病是对水产养殖业的可持续性和生存能力的主要挑战。这一领域的研究不仅是为了支持当地工业，也是为了支持世界各地的其他企业。了解病原体如何感染和生活在鱼的细胞和组织中，以及它们如何在整条鱼中传播，有助于制定适当的控制措施和其他生物技术应用。细菌将蛋白质分泌到周围环境或宿主细胞中，以使它们获得竞争优势或促进进入宿主并在宿主内部生存。至少有七种不同类型的细菌蛋白质分泌系统，被命名为I型到VII型。这些系统将致病蛋白质释放到周围环境中，直接进入宿主细胞，甚至是在同一环境中发现的其他细菌。这些蛋白靶向特定的宿主蛋白并改变其生物功能以获得对宿主的控制。在世界上研究细菌分泌系统的不同实验室中，我们的实验室在迟发爱德华杆菌的III型和VI型分泌系统（T3SS和T6SS）方面都有相当的专业知识。缓步绦虫是淡水和海鱼中常见的重要致病菌，在水环境中分布广泛。迟发E.的疾病特性在很大程度上是由于它们的T3SS和T6SS。姗姗来迟的基因组已被破译。提出的研究旨在鉴定和表征迟达E. T3SS致病蛋白。平均有20-30种致病细菌蛋白从T3SS分泌。E. tarda编码约3700种蛋白质，其中1000多种被归类为潜在的新蛋白质。我们的第一个目标是从1000多个新蛋白中鉴定出T3SS致病蛋白。我们将使用生物信息学的方法来预测基于其分泌信号的T3SS致病蛋白。将训练一个计算程序来预测T3SS致病蛋白。一旦我们有了一个足够的程序，它将被用来筛选1000多个新的迟达E.蛋白质，以筛选出大约50个候选蛋白。这些候选者将使用我们既定的T3SS交付确认协议进行验证。第二个目的是鉴定鉴定的T3SS致病蛋白的功能。我们希望找出与这些细菌蛋白相互作用的目标宿主蛋白，这些细菌蛋白在宿主细胞中破坏或模仿的生理功能，细菌蛋白在细胞和分子水平上的确切作用，以及它们如何共同和单独地促进鱼类的致病过程。凭借我在细菌分泌系统方面的专业知识和强大的多学科团队，我相信我们将完成本提案中概述的目标。潜在的生物技术应用包括开发预防鱼类疾病的疫苗和新疗法。这项研究不仅将促进加拿大的知识型经济，而且将具有全球重要性。

Aquaculture is a one-billion-dollars business in Canada and it provides about 15,000 full time well-paid jobs in many rural and coastal communities. In 2011, Canada was the fifth largest seafood exporter globally. The aquaculture industry is a significant contributor to the stable and strong Canadian economy; it also provides food security as well as healthy food for the Canadian population. Unfortunately, fish diseases are major challenges to the sustainability and viability of the aquaculture industry. Research in this area is urgently needed not only to support the local industry but also other enterprises around the world. Understanding how pathogens infect and live inside fish cells and tissues, and how they spread throughout the whole fish can lead to the development of adequate control measures and other biotechnological applications. Bacteria secrete proteins into the surrounding environment or host cells to allow them to gain a competitive advantage or to facilitate entry into and survival inside the hosts. At least seven different types of bacterial protein secretion systems, named type I to VII, exist. These systems release disease-causing proteins into their surroundings, directly into host cells, or even to other bacteria found in the same environment. These proteins target specific host proteins and alter their biological functions to gain control of the host. Among the different labs in the world studying bacterial secretion systems, our lab has considerable expertise on both type III and type VI secretion systems (T3SS and T6SS) found in the bacterium Edwardsiella tarda. E. tarda is abundant in water environments and is a common and important pathogen of freshwater and marine fish. The disease properties of E. tarda are due in large part to their T3SS and T6SS.The genome of E. tarda has been decoded. The proposed study seeks to identify and characterize T3SS disease-causing proteins in E. tarda. On average there are 20-30 disease-causing bacterial proteins secreted from a T3SS. E. tarda encodes about 3,700 proteins and 1,000+ of them are classified as potentially new proteins. Our first aim is to identify the T3SS disease-causing proteins from the 1,000+ new proteins. We will use a bioinformatics approach to predict T3SS disease-causing proteins based on their secretion signals. A computing program will be trained for the prediction of T3SS disease-causing proteins. Once we have an adequate program, it will be used to screen the 1,000+ new proteins of E. tarda to shortlist the candidates to about 50. These candidates will be validated using our established protocols for the confirmation of T3SS delivery. The second aim is to characterize the functions of the identified T3SS disease-causing proteins. We would like to find out the targeted host proteins that interact with these bacterial proteins, the physiological functions that these bacterial proteins disrupt or mimic in the host cells, the precise roles of the bacterial proteins at the cellular and molecular levels, and how they collectively and individually contribute to the disease-causing process in fish. With my expertise on bacterial secretion systems and a strong multidisciplinary team, I am confident that we will complete the objectives outlined in this proposal. Potential biotechnological applications include the development of vaccines and novel therapeutics to prevent diseases in fish. This research will not only promote a knowledge-based economy in Canada, but will also have global importance.