Replication of avian leukosis viruses with mutations at the primer binding site: use of alternative tRNAs as primers.
We have tested whether avian leukosis viruses (ALVs) can use tRNAs other than tRNATrp to initiate reverse transcription. The primer binding site (PBS) of a wild-type ALV provirus, which is complementary to the 3' end of tRNA(Trp), was replaced with sequences homologous to the 3' ends of six different chicken tRNAs (tRN(APro), tRNA(Lys), tRNA(Met), tRNA(Ile), tRNA(Phe), and tRNA(Ser)). Transfection of these proviruses into chicken embryo fibroblasts resulted in the production of infectious viruses, all of which apparently used the tRNA specified by the mutated PBS to replicate. However, growth of these viruses resulted in reversion to the wild-type (tRNA(Trp)) PBS. Some of the viruses revert quite quickly, while others are more stable. The relative stability of a given PBS correlated with the concentration of the corresponding tRNA in the virion. We determined the percentage of viral RNA that had a tRNA bound to the PBS and found that the occupancy rate is lower in the mutants than in the wild-type virus. We conclude that many different tRNAs can be used as primers to initiate reverse transcription in ALV. However, ALVs that use tRNA(Trp) have a growth advantage over ALVs that use other tRNAs.
Whitcomb
J M
JM
ABL-Basic Research Program, NCI-Frederick Cancer Research and Development Center, Maryland 21702-1201, USA.
Ortiz-Conde
B A
BA
Hughes
S H
SH
D000818:Animals; D001354:Avian Leukosis Virus / Q000235:genetics / Q000502:physiology*; D001483:Base Sequence; D001665:Binding Sites; D002478:Cells, Cultured; D002642:Chick Embryo; D017931:DNA Primers; D005347:Fibroblasts; D005822:Genetic Vectors; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010957:Plasmids; D016133:Polymerase Chain Reaction; D011533:Proviruses / Q000235:genetics / Q000502:physiology; D012345:RNA, Transfer, Amino Acid-Specific / Q000378:metabolism*; D012364:RNA, Transfer, Trp / Q000378:metabolism; D012367:RNA, Viral / Q000096:biosynthesis*; D012194:RNA-Directed DNA Polymerase / Q000378:metabolism*; D015183:Restriction Mapping; D014158:Transcription, Genetic; D014162:Transfection; D014779:Virus Replication
D017931:DNA Primers; D012345:RNA, Transfer, Amino Acid-Specific; D012364:RNA, Transfer, Trp; D012367:RNA, Viral; D012194:RNA-Directed DNA Polymerase