HIV-1, the causative agent of AIDS, exhibits high genomic variation. The high rate of variation permits rapid evolution of new forms of the virus that can evade the host immune response and hamper efforts for effective drug treatment and vaccine development. Three diferent replication steps operate during the life cycle of the virus: the viral reverse transcriptase (which transcribes the viral genomic ssRNA into dsDNA), cellular DNA polymerases (which replicate the integrated provirus), and RNA pol II (which regenerates the viral ssRNA). One source of HIV variation is the low fidelity of DNA synthesis by the viral reverse transcriptase. Measurements of the fidelity of the human replication complex indicate that replication of the human genome is highly accurate, contributing little to HIV diversity. However,there is little information bearing on whether transcription of the integrated viral genome by RNA pol II, to produce both viral mRNA and genomic RNA, contributes to HIV variation. In an attempt to answer this question we have initiated a study of the fidelity of RNA polymerases with the ultimate goal of measuring the fidelity of the human RNA pol II complex. We are currently developing the experimental strategy, which is addapted from the strategy for measuring the fidelity of reverse transcription by HIV-1 RT. To test the experimental strategy, we have initiated measurements of the fidelity of a simple transcription system, the RNA polymerase of bacteriophage T7.
