Construction of adenoviral and retroviral vectors coexpressing the genes encoding the hepatitis B surface antigen and B7-1 protein.
Recombinant retroviral (re-Rv) and adenoviral (re-Ad) vectors for delivery of two foreign genes were constructed, using the internal ribosomal entry site (IRES) of encephalomyocarditis virus (EMCV) which mediates initiation of cap-independent translation. The first gene encoded the hepatitis B surface antigen (HBsAg) and the second encoded human or murine B7-1 molecule, a cell surface protein which is a costimulator for T cell activation. The EMCV IRES sequence was placed between the first and second coding sequences to form a dicistronic DNA fragment. In Rv vectors, the dicistronic fragment was inserted between the 5' long terminal repeat (LTR) and an internal promoter for the neomycin (neo) gene, so that the transcription initiated from the 5' LTR would generate a dicistronic mRNA for the HBsAg and B7-1 molecules. For Ad vectors, the dicistronic fragment was inserted between a cytomegalovirus promoter and a polyA signal to form a transcription cassette. This transcription cassette was inserted into the early region 1 of Ad5 genome to form a replication-defective re-Ad vector, or into early region 3 to form replication-competent vectors. Human cell line A549 infected with the re-Rv vectors or with the re-Ad vectors synthesized and secreted HBsAg at comparable levels, while the B7-1 molecules were detected at the surface of the infected cells, indicating both foreign genes carried by the Rv and Ad vectors were expressed efficiently.
Division of Infectious Diseases, Stanford University School of Medicine, CA 94305-5107, USA.
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D000256:Adenoviridae / Q000235:genetics*; D000818:Animals; D018122:B7-1 Antigen / Q000235:genetics* / Q000378:metabolism; D003587:Cytomegalovirus / Q000235:genetics; D004686:Encephalomyocarditis virus / Q000235:genetics*; D015967:Gene Expression Regulation, Viral; D005822:Genetic Vectors / Q000235:genetics*; D006514:Hepatitis B Surface Antigens / Q000235:genetics* / Q000378:metabolism; D006801:Humans; D051379:Mice; D008979:Moloney murine leukemia virus / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000235:genetics; D012190:Retroviridae / Q000235:genetics*; D014158:Transcription, Genetic; D014407:Tumor Cells, Cultured
D018122:B7-1 Antigen; D006514:Hepatitis B Surface Antigens