Avian retroviral vectors derived from avian defective leukemia virus: role of the translational context of the inserted gene on efficiency of the vectors.
We have constructed retroviral vectors derived from the genome of avian erythroblastosis virus ES4 (AEV ES4). The neo selectable gene was substituted for the original v-erbA or v-erbB oncogenes of AEV, either in the same or in a different reading frames. Recombinant retrovirus were rescued and used to infect chicken embryo fibroblasts or quail QT6 cells. When the neo gene was inserted in the same reading frame as the original oncogene, we obtained (1) a high level of expression of the neo gene, (2) a balanced ration of both genomic and subgenomic RNAs, and (3) high titer recombinant viruses. Conversely, when the neo gene was inserted in a reading frame different from that of the original oncogene, we observed (1) a very low level of expression of the neo protein, (2) a predominance of the viral transcript used as translational template for the neo protein synthesis, and (3) low titer recombinant viruses. One of the vectors was used to transfer a human delta-globin gene into avian cells in culture without detectable rearrangement of this gene, but exhibited a deletion within the conserved noncoding region located between the two original oncogenes. Our data provide information for further construction of double expression vectors. Furthermore, three of the vectors would provide helpful tools to identify genetic elements of the virus genome involved in splicing regulation.
Benchaibi
M
M
Laboratoire de Biologie, Ecole Normale Supérieure de Lyon, France.
Mallet
F
F
Thoraval
P
P
Savatier
P
P
Xiao
J H
JH
Verdier
G
G
Samarut
J
J
Nigon
V
V
D001355:Alpharetrovirus / Q000235:genetics*; D000818:Animals; D001354:Avian Leukosis Virus / Q000235:genetics*; D001483:Base Sequence; D015152:Blotting, Northern; D002460:Cell Line; D002478:Cells, Cultured; D015342:DNA Probes; D004279:DNA, Viral / Q000235:genetics; D003673:Defective Viruses / Q000235:genetics*; D005814:Genes, Viral; D005819:Genetic Markers; D005822:Genetic Vectors; D019868:Kanamycin Kinase; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D018774:Oncogene Proteins v-erbA; D018739:Oncogene Proteins v-erbB; D009857:Oncogenes; D010770:Phosphotransferases / Q000235:genetics; D014176:Protein Biosynthesis; D011533:Proviruses / Q000235:genetics; D012367:RNA, Viral / Q000032:analysis; D015183:Restriction Mapping; D012191:Retroviridae Proteins / Q000235:genetics*; D014158:Transcription, Genetic; D014764:Viral Proteins / Q000235:genetics
D015342:DNA Probes; D004279:DNA, Viral; D005819:Genetic Markers; D018774:Oncogene Proteins v-erbA; D018739:Oncogene Proteins v-erbB; D012367:RNA, Viral; D012191:Retroviridae Proteins; D014764:Viral Proteins; D010770:Phosphotransferases; D019868:Kanamycin Kinase