For decades, spontaneous and induced mutant animals have yielded valuable insights into the roles of specific genes in development and disease. Recently, transgenic and knockout mice, together with large-scale mutagenesis screens of model organisms such as the fruitfly, Drosophila, and the nematode, C. elegans, have contributed detailed insights into the roles of specific genes in development. These intensive studies reveal a remarkable degree of conservation of genetic strategies for control of cell differentiation, cell proliferation, and cell death, as well as pattern formation during development throughout all animals. Advantages of zebrafish for study of fundamental vertebrate genetic principles and molecular developmental genetics include small adult size, relative hardiness in small aquaria, high fecundity, external fertilization and development with accessibility of all developmental stages, highly transparent eggshells, rapid embryonic development, short generation times, ease of embryonal cloning to allow gynogenesis or androgenesis, and applicability for large-scale, rapid mutant screens. Intense worldwide research in zebrafish molecular development, genomics, and mutagenesis during the past decade has complemented the previous strong database on biology, physiology, and development of zebrafish creating a powerful system in which to answer fundamental questions in biology and medicine. The investigators will develop lines of zebrafish highly sensitive to specific histological types of neoplasia occurring in specific tissues. To develop these lines, the investigators will evaluate carcinogen-induced neoplasia in lines of zebrafish with inactivating mutations in orthologs of human tumor suppressor genes. The investigators will screen offspring of fish mutagenized with N-ethyl-N-nitrosourea (ENU), using temperature mediated heteroduplex analysis (TMHA) and the mismatch cleavage enzyme CELI to detect point mutations in zebrafish orthologs of the human tumor suppressor genes P53, RB1, WT1, PTEN, or NF1. The investigators will treat zebrafish embryos or fry of these mutant lines by immersion exposure to N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) or 7,12-dimethylbenz[a]anthracene (DMBA), then will evaluate neoplasm incidences six and 12 months later. These fish will provide a rapid and relatively inexpensive whole-animal bioassay system to detect carcinogenic risks to man from new materials as well as from environmental contaminants. These lines of zebrafish will provide an efficient and inexpensive system for development of new treatment and prevention strategies for cancer, and will help identify new genes controlling growth and differentiation.
