Two defective forms of reverse transcriptase can complement to restore retroviral infectivity.
Retroviral DNA synthesis requires both the DNA polymerase and the RNaseH activities of reverse transcriptase (RT). To test whether two defective RTs--one carrying a mutation in the RNaseH domain and the other with a mutation in DNA polymerase--could work together to complete viral DNA synthesis, we generated phenotypically mixed virions of Moloney murine leukemia virus (M-MuLV) that contained two kinds of mutant RTs. One RNaseH catalytic site mutant complemented both tested DNA polymerase mutants and small amounts of intact viral DNA were generated. This demonstrates that retroviral DNA synthesis can be completed--albeit inefficiently--when DNA polymerase and RNaseH activities are provided by separate RT molecules. Other RNaseH mutants failed to complement, suggesting that some aspects of the RNaseH domain are essential to RT's DNA polymerase function. Phenotypically mixed virions were also used to demonstrate that RT and integrase (IN) can be provided by separate polyprotein precursors and complete the early stages of retroviral replication.
Telesnitsky
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Department of Biochemistry and Molecular Biophysics, Columbia University College of Physicians and Surgeons, New York, NY 10032.
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D016475:3T3 Cells; D000818:Animals; D004254:DNA Nucleotidyltransferases / Q000235:genetics; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism; D005816:Genetic Complementation Test; D019426:Integrases; D051379:Mice; D008979:Moloney murine leukemia virus / Q000201:enzymology / Q000235:genetics* / Q000472:pathogenicity; D009154:Mutation; D010641:Phenotype; D011533:Proviruses / Q000201:enzymology / Q000235:genetics; D012194:RNA-Directed DNA Polymerase / Q000235:genetics* / Q000378:metabolism; D051381:Rats; D012191:Retroviridae Proteins / Q000235:genetics; D016914:Ribonuclease H / Q000378:metabolism; D014771:Virion / Q000201:enzymology / Q000235:genetics; D014774:Virulence; D016662:Virus Integration
D012191:Retroviridae Proteins; D004254:DNA Nucleotidyltransferases; D019426:Integrases; D012194:RNA-Directed DNA Polymerase; D004259:DNA-Directed DNA Polymerase; D016914:Ribonuclease H