Expression of a functional influenza viral cap-recognizing protein by using a bovine papilloma virus vector.
The gene for the influenza viral PB2 protein, which recognizes and binds the 5'-terminal cap 1 structures (m7GpppNm) on eukaryotic mRNAs, was inserted into a bovine papilloma virus vector under the control of a mouse metallothionein I (MT-I) promoter. After transfection of this vector into mouse NIH 3T3 cells, a cell line, cPB2, was obtained that produces PB2-specific mRNA and authentic PB2 protein. Induction of the MT-I promoter with CdCl2 causes about a 10-fold increase in PB2 mRNA and protein levels. The expressed PB2 protein is functional, as it relieves the block in viral mRNA synthesis exhibited by a temperature-sensitive viral mutant containing a cap-binding defect in the PB2 protein. This demonstrates complementation of a function of a negative-strand RNA virus by a gene product expressed in a cell line from recombinant DNA.
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D000818:Animals; D001426:Bacterial Proteins; D052688:Bovine papillomavirus 1 / Q000235:genetics*; D002460:Cell Line; D004262:DNA Restriction Enzymes / Q000378:metabolism; D015247:Deoxyribonuclease HindIII; D015252:Deoxyribonucleases, Type II Site-Specific; D004591:Electrophoresis, Polyacrylamide Gel; D005786:Gene Expression Regulation; D009980:Influenza A virus / Q000235:genetics*; D051379:Mice; D027383:Papillomaviridae / Q000235:genetics*; D012315:RNA Caps / Q000378:metabolism*; D014162:Transfection; D014764:Viral Proteins / Q000235:genetics* / Q000378:metabolism
D001426:Bacterial Proteins; D012315:RNA Caps; D014764:Viral Proteins; D004262:DNA Restriction Enzymes; D015247:Deoxyribonuclease HindIII; C024404:BglII endonuclease; D015252:Deoxyribonucleases, Type II Site-Specific