The basic objective of this research is to determine the roles of extra-mutational events, such as mitotic recombination, gene inactivation, chromosomal rearrangements, deletions, or chromosome segregation, in the expression of recessive mutations in cultured mammalian cells. Our ultimate goal is a clear understanding of the cellular processes and mechanisms involved in the generation and expression of heritable variation in mammalian somatic cells, and definition of the role of such processes in the initiation and promotion stages of carcinogenesis. Combining classical somatic cell genetic approaches with recombinant DNA technologies, we will study the effects of chromosomal rearrangements on gene expression and mutation or deletion. We will utilize DNA-mediated gene transfer to study how the expression and mutability of a gene are affected by its chromosomal environment, and to determine whether transferred Chinese hamster APRT gene sequences differ from the endogenous CHO APRT genes in their susceptibility to mutation, inactivation, or deletion. In addition, we propose to utilize a cell line that is heterozygous for four different linked genetic or cytogenetic markers to assay mitotic recombination, gene inactivation, chromosomal rearrangements, deletions, or chromosome segregation events leading to the expression of APRT- recessive mutant phenotypes in mutagen-treated or untreated control cell cultures. We will use this system to determine whether extra-mutational events are induced by known tumor initiators or promoters. These experiments should provide information concerning the cellular processes and mechanisms involved in tumor initiation and promotion.
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