MICROSOME-MEDIATED METHYLATION OF DNA BY N,N-DIMETHYLNITROSAMINE INVITRO

Incubation of 14C-labeled N,N-dimethylnitrosamine (DMN) with rat hepatic microsomes results in binding of 14C to exogenous calf thymus DNA and endogenous protein. This binding appears to be enzymatically mediated and dependent on NADPH. If this in vitro metabolism of DMN proceeds in a fashion similar to what occurs in vivo, it would be safe to assume that binding of 14C label to biological macromolecules in vitro is due to a methylation process. This remains to be shown by isolation and identification of specific methylated nucleotides from what may be methylated DNA. Binding of formaldehyde generated from DMN may also be occurring. Since no mechanism by which formaldehyde can be incorporated into nucleic acids as a methyl group covalently bound to the bases of nucleic acids is known, formaldehyde derived from 14C-DMN would not be responsible for the 14C label associated with these methylated bases. Isolation of methylated bases from 14C labeled DNA would verify this assumption. An NADPH dependent enzyme mediated transfer of 14C label from 14C-DMN to calf thymus DNA in vitro, which could represent a methylation process was shown. This alkylation of DNA may be the basis for the carcinogenic effect of DMN and, by implication, nitrites and other nitrosamine precursors.