title: The M184V mutation in HIV-1 reverse transcriptase reduces the restoration of wild-type replication by attenuated viruses.
abstract: OBJECTIVE
  To study the ability of HIV constructs containing the M184V substitution in reverse transcriptase (RT), which causes resistance to lamivudine, to evolve mutations that compensate for deletions within the HIV genome.
  
  
  METHODS
  Viruses containing deletions in non-coding regions of the viral genome were examined in tissue culture to see whether the additional presence of M184V delays the reestablishment of wild-type replication kinetics. Potential compensatory mutations were identified by sequencing, and site-directed mutagenesis was carried out to confirm the biological relevance of such substitutions. The rate of initiation of reverse transcription was measured using either recombinant wild-type RT or RT containing M184V.
  
  
  RESULTS
  M184V-containing viruses were unable to undergo compensatory mutagenesis to reestablish wild-type replication kinetics, whereas viruses that did not contain M184V were able to mutate extensively. This ability was demonstrated most extensively in viruses deleted of an "A-rich loop", located upstream of the primer-binding site, which is involved in initiation of reverse transcription. The rate of such initiation was severely diminished in virus containing the RT enzyme carrying the M184V substitution. This inhibitory effect was significantly enhanced in a biochemical system that included both the M184V mutant enzyme and a viral DNA template that contained the deletion in the A-rich loop.
  
  
  CONCLUSIONS
  These findings provide further biological and biochemical evidence that M184V-containing viruses are impaired in replication fitness. Viruses that had the A-rich-loop deleted were able to reestablish replication ability quickly in the wild-type RT, which provides further evidence for the plasticity of the HIV genome.
authors.affiliation: McGill University AIDS Centre, Lady Davis Institute-Jewish General Hospital, McGill University, 3755 chemin Côte-Sainte-Catherine, Montréal, Québec, Canada H3T 1E2.
authors.forename: Xin
authors.initials: X
authors.lastname: Wei
authors.forename: Chen
authors.initials: C
authors.lastname: Liang
authors.forename: Matthias
authors.initials: M
authors.lastname: Götte
authors.forename: Mark A
authors.initials: MA
authors.lastname: Wainberg
mesh_terms: D002478:Cells, Cultured; D004279:DNA, Viral / Q000378:metabolism; D017353:Gene Deletion; D015658:HIV Infections / Q000235:genetics / Q000821:virology; D054303:HIV Reverse Transcriptase / Q000235:genetics*; D015497:HIV-1 / Q000201:enzymology / Q000235:genetics*; D006801:Humans; D009154:Mutation / Q000235:genetics*; D021141:Nucleic Acid Amplification Techniques; D012367:RNA, Viral / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D017422:Sequence Analysis, DNA; D014157:Transcription Factors / Q000235:genetics; D014779:Virus Replication / Q000235:genetics*
chemical_list: D004279:DNA, Viral; D012367:RNA, Viral; D014157:Transcription Factors; D054303:HIV Reverse Transcriptase