Many mutagens and carcinogens induce genomic rearrangements in mammalian cells. Often these rearrangements are lethal to the cell; however, rearrangements resulting in cell viability must be considered in assessing the consequences of mutagen and carcinogen action. Most mammalian mutagenesis assay systems lack the ability to detect genomic rearrangements. We study deletions and rearrangements using the Chinese hamster ovary (CHO) cell line, AS52, which carries a single functional copy of the bacterial gpt gene stably integrated into the CHO genome. AS52 cells are sensitive to induced mutation by a variety of clastogens and radiomimetic agents which are often classified as nonmutagens in other mutagenesis assays. This sensitivity is probably due to the location of the integrated gpt transgene. Mutations arising as a result of genomic rearrangements can be recovered in this line. The loss of functional gpt sequences can result from intrachromosomal deletion, mitotic recombination or gene conversion. We are refining the AS52 cell mutational assay system to distinguish the extent to which mutagens may induce each of the three stable (i.e., viable) genomic in situ hybridization techniques. In addition, we have initiated a project to clone the gpt site of insertion and to characterize flanking genomic sequences. These studies should result in the generation of gpt-linked chromosome specific probes that will allow one to assess frequencies of induced mitotic recombination or gene conversion events. Also, we have characterized a nonfunctional gpt gene carrying an insert of approximately 1 kb which is closely linked to the functional gpt gene in AS52 cells. This linked nonfunctional gpt sequence provides a marker for the rapid identification of mutants with deletions small enough to study in further detail . Two clastogenic agents currently under study are 5-azacytidine (5AC) and 1-beta-D-arabinofuranosylcytosine (ara-C). Neither 5AC nor ara-C are mutagenic at the hprt locus but both are potent mutagens at the gpt locus. Efforts are underway to define the spectrum of mutations induced by 5AC.
