Cancer is due to misguided cell developmental processes such as cell growth, cell determination and differentiation. Unless we understand the basic molecular mechanisms of these processes, we cannot understand the developmental biology of neoplastic cells. To tackle these problems, advantage will be taken of the hereditary melanoma formation in certain fish. Genetic factors have been identified which govern the development of a distinct pigment cell, the M cell: the M gene, determines embryonic neural crest cells to differenctiate into M cells; a linked regulating gene, IR, controls this process, several non-linked R genes, n1R, control the proliferation and differentiation of the determined cells. By mutating or outcrossing R genes, one can create a whole spectrum of R genotype, each giving rise to developmentally mutant M cells forming benign melanomas at one end of the spectrum and malignant melanomas at the other end. The long-term goal of ths research is to analyze cell developmental processes by unravelling the molecular functions of the mentioned genetic factors and by studing at the cellular level, characteristics of normally and tumorously developing M cells. To achieve this goal, the research proposed two aims: (i) to isolate the M gene, thereby laying the basis for a molecular analysis of the determination process. The isolation will be done using the methodology of a recently described gene transfer system: total genomic DNA from M genotypes, when injected into the neural crest region of embryos, resulted in almost 10% of the fish displaying M cell colonies after 3 months. We will use this powerful system as a bioassay to sub-select from genomic and sub-genomic libraries for an M gene-specific clone. (ii) To establish an in vitro model system in the form of neural crest cells which give rise to normal M cells when R genes are unaltered, and to various abnormal (tumorous) M cells when respective R genes are mutated and/or outcrossed. While establishing the system, it is anticipated that initial insights will be gained into the function of R genes and thus into the principles of normal and cancerous cell development. One very important advantage of such a system: the development of cancerous cells in culture can be observed from the very beginning as the changes take place. This is an advantage over tumour cell lines where such changes have already taken place.
