IN-VITRO FOLATE-DEFICIENCY INDUCES DEOXYNUCLEOTIDE POOL IMBALANCE, APOPTOSIS, AND MUTAGENESIS IN CHINESE-HAMSTER OVARY CELLS

The genetic and epigenetic effects of nutritional folate deficiency we re studied in two Chinese hamster ovary (CHO) cell Lines. The CHO-ABS cell line (hemizygous at the aprt locus) and CHO-UV5 (DNA repair-defic ient mutant of AA8) were cultured in Ham's F-12 medium or in custom-pr epared Ham's F-12 medium lacking foIic acid, thymidine, and hypoxanthi ne. Cells cultured acutely in the folate deficient medium exhibited in itial growth arrest, followed by massive cell death and DNA fragmentat ion into nucleosomal multimers characteristic of apoptosis. Although p rolonged culture in the folate deficient medium was cytostatic and let hal to the majority cells, minor subpopulations in both cell lines fai led to initiate cell death, exhibited phenotypic abnormalities, and ad apted a selective growth advantage under marginal folate conditions. T hese ''resistant'' clones exhibited major alterations in deoxynucleoti de pools associated with an increase in mutant frequency at the aprt l ocus as detected by resistance to cytotoxicity in 8-azaadenosine. The mutation frequency in the DNA repair-deficient CHO-UV5 cells was simil ar to 100-fold greater than that in the parental AA8 clones, underscor ing the importance of DNA repair under conditions of folate deficiency and nucleotide pool imbalance. The enhanced mutation frequency in the DNA repair-competent folate-deficient CHO-BAS cells suggests that DNA repair activity is less effective under folate-deficient conditions. These results add to the accumulating clinical and experimental eviden ce relating chronic folate deficiency to genomic instability and carci nogenesis.