MUTAGENESIS BY HUMAN-IMMUNODEFICIENCY-VIRUS REVERSE-TRANSCRIPTASE - INCORPORATION OF O-6-METHYLDEOXYGUANOSINE TRIPHOSPHATE

The high frequency of incorporation of non-complementary nucleotides b y HIV-1 reverse transcriptase is likely to be a major factor in the ex ceptionally rapid accumulation of viral mutations during the course of AIDS infections. To investigate whether this high level of infidelity is also associated with the incorporation of nucleotide analogs, we a nalyzed O-6-methyldeoxyguanosine triphosphate and compared the incorpo ration of this analog by HIV-1 reverse transcriptase to that catalyzed by other DNA synthesizing enzymes. Our results indicate that O-6-meth yldeoxyguanosine triphosphate serves as a substrate for DNA synthesize d in vitro by HIV-1 RT on both DNA and RNA templates. The product DNA contains the modified purine; it is sensitive to the repair enzyme, O- 6-methylguanine methyltransferase, which specifically reacts with DNA containing methylated guanines at the O-6 position. Using a forward mu tation assay we demonstrated that the nucleotide analog incorporated b y HIV-1 RT is mutagenic. The mutations produced are single-base substi tutions opposite template thymidines and result in A:T --> G:C transit ions. The incorporation of a mutagenic nucleotide by HIV-1 RT highligh ts the possibility of increasing the rate of mutagenesis of HIV by the use of nucleotides that form non-complementary base pairs at high fre quency.