Mutants of feline immunodeficiency virus resistant to 2',3'-dideoxy-2',3'-didehydrothymidine.
We selected mutants of feline immunodeficiency virus (FIV) that are resistant to 2',3'-dideoxy-2',3'-didehydrothymidine (d4T). Two mutants were selected in cultured cells with a stepwise increase in d4T concentration, resulting in mutants able to replicate in 100 microM d4T. These mutants were three- to sixfold more resistant to d4T than wild-type FIV. They were also cross-resistant to 3'-azido-3'-deoxythymidine (AZT), 3'-fluoro-2',3'-dideoxythymidine, 2',3'-dideoxycytidine, 2',3'-dideoxyinosine, and 9-(2-phosphonylmethoxyethyl)adenine, and they were highly resistant to phosphonoformic acid (PFA). Plaque-purified mutants were isolated from each of the mutant populations. The mutant phenotype was stable, because both of the plaque-purified mutants remained d4T resistant even after three passages in the absence of d4T. One of the plaque-purified mutants, designated D4R-3c, was further characterized. Compared with wild-type reverse transcriptase (RT), RT purified from D4R-3c was 3-fold resistant to inhibition by the 5'-triphosphate of d4T, 10-fold resistant to inhibition by the 5'-triphosphate of AZT, and 6-fold resistant to PFA. D4R-3c had a single point mutation in the RT-encoding region of the pol gene at position 2474, resulting in a Val to Ile mutation at codon 47 of the FIV RT. The role of this mutation in d4T resistance was confirmed by site-directed mutagenesis.
Zhu
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YQ
Division of Biological Sciences, University of Montana, Missoula 59812, USA.
Remington
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KM
North
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TW
D000818:Animals; D019380:Anti-HIV Agents / Q000494:pharmacology*; D002415:Cats; D002460:Cell Line; D017931:DNA Primers; D004279:DNA, Viral / Q000032:analysis / Q000187:drug effects / Q000302:isolation & purification; D004351:Drug Resistance; D016089:Immunodeficiency Virus, Feline / Q000187:drug effects* / Q000235:genetics* / Q000472:pathogenicity; D007700:Kinetics; D016297:Mutagenesis, Site-Directed; D015183:Restriction Mapping; D018894:Reverse Transcriptase Inhibitors / Q000494:pharmacology; D018119:Stavudine / Q000494:pharmacology*; D010948:Viral Plaque Assay
D019380:Anti-HIV Agents; D017931:DNA Primers; D004279:DNA, Viral; D018894:Reverse Transcriptase Inhibitors; D018119:Stavudine