Diminished efficiency of HIV-1 reverse transcriptase containing the K65R and M184V drug resistance mutations.
OBJECTIVE
To determine the underlying biochemical mechanisms responsible for the diminished viral replicative capacity associated with K65R/M184V-containing viruses.


METHODS
We studied the efficiency of (-)ssDNA synthesis by recombinant wild-type and mutated HIV-1 reverse transcriptases in cell-free assays. In addition, we determined susceptibility levels to nucleoside analog reverse transcriptase inhibitors (NRTIs) both in cell-free and cell culture assays.


RESULTS
We observed that the K65R/M184V mutations in reverse transcriptase caused reductions in the efficiency of initiation of (-)ssDNA synthesis by increasing pausing at positions +3 and +5 as well as diminished RNA usage. These findings were confirmed in cell culture data using MT-4 cells and cord blood mononuclear cells.


CONCLUSIONS
The simultaneous presence of K65R and M184V in reverse transcriptase has a negative impact with regard to the efficiency of initiation of (-)ssDNA synthesis and RNA usage, that exceeds the effect of either mutation on its own. These mechanisms, among others, are responsible for the diminished viral replicative capacity observed in tissue culture when K65R/M184V-containing viruses are studied.
Frankel
Fernando A
FA
McGill AIDS Centre, Lady Davis Institute-Jewish General Hospital, Montreal, Quebec, Canada.
Invernizzi
Cédric F
CF
Oliveira
Maureen
M
Wainberg
Mark A
MA
D002460:Cell Line; D017931:DNA Primers / Q000494:pharmacology; D004277:DNA, Single-Stranded / Q000096:biosynthesis / Q000187:drug effects; D004279:DNA, Viral / Q000096:biosynthesis / Q000187:drug effects; D024882:Drug Resistance, Viral; D015658:HIV Infections / Q000821:virology; D015497:HIV-1 / Q000187:drug effects* / Q000235:genetics / Q000254:growth & development; D006801:Humans; D009154:Mutation / Q000235:genetics; D010766:Phosphorylation; D012367:RNA, Viral / Q000187:drug effects; D018894:Reverse Transcriptase Inhibitors / Q000494:pharmacology*; D014779:Virus Replication / Q000187:drug effects*; D015215:Zidovudine / Q000494:pharmacology
D017931:DNA Primers; D004277:DNA, Single-Stranded; D004279:DNA, Viral; D012367:RNA, Viral; D018894:Reverse Transcriptase Inhibitors; D015215:Zidovudine