Generation and Rapid Mapping of Low-Penetrance Disease Alleles in Zebrafish

描述（由申请人提供）：这项工作的长期目标是通过发现致病突变来改善人类健康。许多人类疾病，包括癌症，最终都是由特定基因的突变引起的。这些基因的发现是改善疾病检测和诊断的关键一步，也是设计基于分子的靶向治疗的关键一步。然而，疾病基因的发现受到在人群中工作的困难和许多致病等位基因的低外显率的阻碍。为了改善疾病基因的发现，我们正在使用斑马鱼，这是一种人类疾病的优秀模型，包括癌症、微生物发病机制和出生缺陷。以前，我们使用正向遗传筛选来识别导致斑马鱼癌症易感性增加的基因突变。我们和其他人也表明，特定人类疾病等位基因的转基因表达使斑马鱼易患相关的人类疾病。这些“易感菌株”通常需要“第二次攻击”——在特定位点发生突变——才能表现出疾病，因此可能是识别关键疾病——修饰人类疾病基因的宝贵资源。由于疾病的低外显率和长潜伏期，以及缺乏用于人类最常见癌症上皮癌的斑马鱼模型，使用易感菌株进行基因发现的进展受到阻碍。我们已经成功地利用单倍型定位鉴定了一种在斑马鱼中引起睾丸肿瘤的低外显率、成人发病的疾病基因。在这里，我们建议建立一个强大的，用户友好的单倍型映射面板，作为整个斑马鱼群落的资源。利用该小组和斑马鱼分离酶癌症易感性菌株，我们将确定负责上皮癌变的新基因突变。单倍型定位方法和这些上皮癌株的可用性将显著扩大斑马鱼用于人类疾病研究的能力。

DESCRIPTION (provided by applicant): The long-term goal of this work is to improve human health by enabling the discovery of disease-causing mutations. Many human diseases, including cancer, are ultimately caused by mutations in specific genes. Discovery of these genes is a critical step in improving detection and diagnosis of disease, as well as for the design of molecularly-based, targeted therapies. However, disease gene discovery is hampered by the difficulties of working with human populations and by the low penetrance of many disease-causing alleles. To improve disease gene discovery, we are using the zebrafish, an excellent model of human diseases including cancer, microbial pathogenesis and birth defects. Previously, we used forward genetic screens to identify gene mutations that cause increased cancer susceptibility in zebrafish. We and others have also shown that transgenic expression of specific human disease alleles makes zebrafish susceptible to the relevant human disease. These "susceptibility strains" generally require a "second hit"-a mutation at a specific locus-to manifest disease, and thus could be an invaluable resource for identifying critical disease- modifying genes in human disease. Progress in using the susceptibility strains for gene discovery is hampered by the low penetrance and long latency of disease, and by the lack of zebrafish models for epithelial cancers, the most common cancers in humans. We have successfully used haplotype mapping to identify a low-penetrance, adult onset disease gene causing testicular tumors in zebrafish. Here we propose to establish a robust, user-friendly haplotype mapping panel as a resource for the entire zebrafish community. Taking advantage of this panel and the zebrafish separase cancer-susceptibility strain, we will identify novel gene mutations responsible for epithelial carcinogenesis. The availability of haplotype mapping methods and these epithelial cancer strains will significantly expand the power of the zebrafish for human disease research. PUBLIC HEALTH RELEVANCE: More than half a million people die of cancer each year in the US, and better treatments are needed. To discover the genes mutated in cancers and allow the development of more effective therapies, we are using the genetic vertebrate model system, the zebrafish. In this proposal we describe methods to significantly increase the efficiency of cancer gene discovery in zebrafish and the generation of zebrafish that accurately model human cancers.