BUDR DEPENDENCE, MALIGNANCY, AND DIFFERENTIATION

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The long term goals are to elucidate the molecular mechanisms involved in regulating malignant transformation, differentiation, and chromosomal recombination in mammalian cells. These areas are central to an understanding of cancer, of normal development and birth defects, and of diseases associated with chromosomal breakage syndromes. The basic techniques are those of mammalian cell genetics and recombinant DNA. The specific aims are to: (1) determine the mechanism by which the thymdine (dT) analog 5-bromodeoxyuridine (BrdU) suppresses differentiation in Syrian hamster melanoma cells, and how deoxycytidine reverses the suppression of differentiation without affecting the amount of BrdU in DNA; these studies will involve matabolic tracer experiments and also a cloned dopa oxidase gene as a molecular probe; (2) carry out a genetic analysis of the ability of mutant cells to survive with all the dT residues in DNA replaced by BrdU, and clone the gene responsible for this phenotype; (3) analyze the mechanism by which BrdU induces sister chromatid exchanges (SCE), and determine whether a plasmid recombination system capable of detecting interstrand recombination can be used to study the induction of SCE; (4) analyze mutant melanoma cells which exhibit BrdU dependence and express the transformed phenotype only in the presence of BrdU, and determine whether BrdU dependence is due to a BrdU-activated oncogene; (5) establish a new system for oncogene detection, utilizing hybrid collagen promoter plasmids whose expression is suppressed in oncogene transformed cells; and (6) examine the expression of malignant transformation in hybrids between transformed and untransformed cells, utilizing the collagen promoter plasmids mentioned above, and determine if specific human chromosomes carry genes that suppress transformation in hybrid cells.