Mutagenesis in cultured cells with altered poly (Adp-ribose) metabolism

Poly(ADP-ribose) is synthesized in response to DNA damage caused by either chemicals or radiation. The biological function of this polymer is not known. However, most of the evidence for role(s) of poly(ADP-ribose) is based on the inhibition of the enzyme poly(ADP-ribose) polymerase by different inhibitors. When the synthesis of poly(ADP-ribose) is inhibited following DNA damage, increases are observed in cytotoxicity, chromosomal aberrations, sister chromatid exchanges and malignant transformation. This investigation was conducted to study the effect of poly(ADP-ribose) metabolism on mutagenesis in C3H10T1/2 cells. Poly(ADP-ribose) metabolism was altered by the inhibitor, 3-methoxybenzamide (MBA) or by NAD-depletion. Ouabain-resistant mutations were induced by exposing cells to N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) in the presence and absence of poly(ADP-ribose) synthesis. Mutations at the Na('+)/K('+)ATPase locus were selected by growth in ouabain containing medium. In dividing cells treated with MNNG, limiting poly(ADP-ribose) synthesis by MBA or NAD-depletion resulted in significant decreases in mutation. Since inhibiting poly(ADP-ribose) synthesis is known to enhance the cytotoxicity of MNNG and cause cell cycle blocks, it was not clear whether the observed decrease in mutation frequency was the result of a block in the expression of mutations or whether poly(ADP-ribose) synthesis normally effects an SOS type of response to DNA damage. Therefore, analogous studies were conducted in non-dividing cells which eliminated cell cycling during the period of DNA repair and inhibition of poly(ADP-ribose) synthesis doubled the mutation rates. These data suggest that poly(ADP-ribose) may function either directly or indirectly in limiting events which lead to mutations at the Oua('r) locus.