Cloning of adenovirus type 12 E1 genes into a retroviral vector and their differential splicing in mouse cells.
Early region 1 (E1) of adenovirus type 12 (Ad12) genome is able to transform nonpermissive primary rodent cells in vivo and in vitro. To analyse the role of the E1a gene products alone or in connection with the 58-kDa protein encoded by E1b during oncogenic transformation, we have cloned genomic fragments of both subregions into the retroviral vector, pZIP-NeoSV(X)1. Both constructs are expressed in mouse 3T3 cells, but, in contrast to E1b, the amount of genomic retroviral RNA carrying E1a-specific sequences was low in transfected psi2 cells and not detectable in infected NIH3T3 cells. Nevertheless, we could demonstrate the integration of the complete E1a-carrying provirus into the NIH3T3 genome. However, after infection of primary mouse embryo fibroblasts, high retrovirus-mediated expression of E1a leads to the immortalization of these cells. In the derived cell line, only the 13S transcript and the unspliced form of E1a RNA could be demonstrated, but not the 12S transcript. These results demonstrate that the ratio of genomic vs. subgenomic retroviral RNAs of Ad12 E1-carrying vectors is dependent on the cloned insert and the cell system used.
Brockmann
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Institute of Molecular Biology (Cancer Research), University of Essen, F.R.G.
Schmidtmann
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Fürst
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Tries
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Esche
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D000256:Adenoviridae / Q000235:genetics*; D017456:Adenovirus Early Proteins; D000818:Animals; D015152:Blotting, Northern; D015139:Blotting, Southern; D002460:Cell Line; D003001:Cloning, Molecular; D004279:DNA, Viral / Q000235:genetics; D005814:Genes, Viral; D005822:Genetic Vectors; D051379:Mice; D009856:Oncogene Proteins, Viral / Q000235:genetics*; D016133:Polymerase Chain Reaction; D011533:Proviruses / Q000235:genetics; D012326:RNA Splicing; D012190:Retroviridae / Q000235:genetics; D014162:Transfection
D017456:Adenovirus Early Proteins; D004279:DNA, Viral; D009856:Oncogene Proteins, Viral