FIDELITY OF THE RNA-DEPENDENT DNA-SYNTHESIS EXHIBITED BY THE REVERSE TRANSCRIPTASES OF HUMAN-IMMUNODEFICIENCY-VIRUS TYPE-1 AND TYPE-2 AND OF MURINE LEUKEMIA-VIRUS - MISPAIR EXTENSION FREQUENCIES

Human immunodeficiency viruses type 1 and 2 (HIV-1 and HIV-2, respectively) exhibit extensive genetic variations. It was postulated that much of this genetic variability stems from the low fidelity of the reverse transcription step. Both HIV reverse transcriptases (RTs) were shown to be particularly error-prone during the in vitro DNA-dependent DNA synthesis relative to other retroviral RTs. Extension of mismatched 3'-termini of the primer DNA was shown to be a major determinant in the infidelity of HIV RTs. However, reverse transcriptases generally exhibit dual template specificities. Therefore, we determined in the current study the fidelity of RNA-dependent DNA synthesis catalyzed in vitro by the RTs of HIV-1 and HIV-2 in comparison with that of murine leukemia virus (MLV) RT. Consequently, we examined the ability of these enzymes to extend preformed 3'-terminal A.A, A.C, and A.G mispairs by quantitating the amount and length of extended primers in a primer extension assay using ribosomal RNA as a template. The results demonstrate that the three RTs studied exhibit efficient extensions from 3'-terminal mispairs with a specificity of A.C > A.A > A.G. Nevertheless, the HIV RTs are qualitatively as well as quantitatively more error-prone than MLV RT. The mispair extension efficiency appears to be affected mainly by the increase of apparent K(m) values, rather than by the change in V(max) values. In all, it is apparent that polymerization errors produced during the RNA-dependent as well as DNA-dependent DNA synthesis by HIV-1 and HIV-2 RTs might be a major cause for the high mutation frequencies observed by these AIDS-causing viruses.