MUTATIONS AFFECTING THE CYTOPLASMIC DOMAIN OF THE MOLONEY MURINE LEUKEMIA-VIRUS ENVELOPE PROTEIN - RAPID REVERSION DURING REPLICATION

Five premature termination mutations and five missense mutations were introduced into the portion of cloned Moloney murine leukemia virus (M -MuLV) DNA encoding the Env cytoplasmic domain. All of the mutant DNAs gave rise to replication-competent virus after transfection of NIH/3T 3 cells, but several of the mutant DNAs scored as replication-defectiv e when introduced into Rat2 cells. Cell lines stably expressing the mu tant DNAs all released virion particles, and in all but one case infec tious virus were generated. These viable mutants were all found to hav e reverted to the wild-type sequence. To generate fully mutant virus s tocks, the mutant DNAs were introduced transiently into COS cells, whi ch are resistant to infection with MuLV, thus prohibiting reversion by error-prone mechanisms involving reverse transcription. Virions harve sted from the COS cells were confirmed as mutant by analyzing both vir ion proteins and the viral DNA they generated, and were then tested fo r infectivity in NIH/3T3 cells. The mutant viruses were infectious, bu t still rapidly gave rise to revertants. We conclude that the mutation s within the cytoplasmic domain do not provide an absolute block to vi rus replication, but that the mutants replicate more slowly than the w ild-type and quickly give rise to revertants with selective advantage for replication.