Intrinsic DNA synthesis fidelity of xenotropic murine leukemia virus‐related virus reverse transcriptase

Abstract

Although recent reports have provided strong evidence to suggest that xenotropic murine leukemia virus-related virus (XMRV) is unlikely to be the causative agent of prostate cancer and chronic fatigue syndrome, this recombinant retrovirus can nonetheless infect human cells in vitro and induce a chronic infection in macaques. In the present study, we determined the accuracy of DNA synthesis of the reverse transcriptases (RTs) of XMRV and Moloney murine leukemia virus (MoMLV) using a combination of pre-steady-state kinetics of nucleotide incorporation and an M13mp2-based forward mutation assay. The results obtained were compared with those previously reported for the HIV type 1 BH10 strain (HIV-1(BH10)) RT. MoMLV and XMRV RTs were 13.9 and 110 times less efficient [as determined by the catalytic rate constant of the nucleotide incorporation reaction ((pol))/equilibrium constant (K(d))] than the HIV-1(BH10) RT in incorporating correct nucleotides. Misinsertion and mispair extension kinetic studies demonstrated that MoMLV RT was more accurate than the HIV-1(BH10) RT. In comparison with the MoMLV RT, the XMRV RT showed decreased mispair extension fidelity and was less faithful when misincorporating C or A opposite A. However, the XMRV RT showed stronger selectivity against G in misinsertion fidelity assays. Forward mutation assays revealed that XMRV and MoMLV RTs had similar accuracy of DNA-dependent DNA synthesis, but were > 13 times more faithful than the HIV-1(BH10) enzyme. The mutational spectra of XMRV and MoMLV RTs were similar in having a relatively higher proportion of frameshifts and transversions compared with the HIV-1(BH10) RT. However, the XMRV polymerase was less prone to introduce large deletions and one-nucleotide insertions.