Abstract

Treatment of DNA in vitro with the carcinogen N-acetoxy-2-fluorenylacetamide (N-acetoxy-2-FAA) drastically reduced the capacity of the DNA to serve as a template for RNA synthesis. Significant inactivation of template activity occurred within seconds after mixing the carcinogen with DNA. A given quantity of treated DNA could bind 10 times more RNA polymerase than the same quantity of control DNA. RNA synthesis directed by treated DNA ceased after 5 min of incubation, while RNA synthesis directed by control DNA continued for over 1 hr. Treated DNA was as effective as control DNA in supporting the exchange of [³²P]pyrophosphate into nucleoside triphosphates in the presence of RNA polymerase. These findings indicated that N-acetoxy-2-FAA inhibited RNA synthesis by preventing chain elongation, and that chain initiation was unaffected. Denaturation by heat or alkali decreased the template capacity of control DNA but increased the template capacity of treated DNA. In addition, treated DNA was more effective than control DNA in supporting the homopolymerization of adenylic acid by RNA polymerase. It therefore appeared likely that N-acetoxy-2-FAA produced regions of partial denaturation of DNA. Comparison of the effects of a series of N-acetoxyarylacetamides revealed that abolition of DNA template activity only occurred when the aryl moiety was greater than one ring in size and when the nitrogen atom was located para to the aromatic system.