Optimizing gene expression in BPV-transformed cells: effects of cell type on enhancer/promoter interaction.
We have compared several combinations of enhancers and promoters in expressing the chloramphenicol acetyl transferase gene in transient assays, in mouse C127, the most widely used host cell for the bovine papilloma virus (BPV) expression vector. Of the various combinations tested, the unit comprised of the SV40 enhancer and adenovirus type 2 major late promoter (MLP) was the most active in BPV transformed C127 cells. We further demonstrate that untransformed and BPV transformed C127 cells respond differently to the various enhancer/promoter combinations tested. Moving the SV40 enhancer closer to the cap site of a complete MLP (from -414 to -107) reduced potentiation to less than half in BPV transformed cells. The level of potentiation with enhancer at either site was similar in human HeLa cells. In BPV transformed C127 cells, the SV40 enhancer and the MLP (at the -414 site) supports 4-5 fold greater levels of expression than the murine sarcoma virus (MSV) enhancer/mouse metallothionein (MT) promoter which has previously been extremely effective in BPV vectors. Our findings provide a basis for the improvement of the BPV vector system in supporting increased levels of expression of proteins of important therapeutic application.
Hurwitz
D R
DR
Rorer Biotechnology Inc., Molecular Biology Development Division, Springfield, VA 22151.
Hodges
R
R
Drohan
W
W
Sarver
N
N
D000123:Acetyltransferases / Q000235:genetics; D000818:Animals; D001483:Base Sequence; D052688:Bovine papillomavirus 1 / Q000235:genetics*; D002460:Cell Line; D002471:Cell Transformation, Neoplastic; D015500:Chloramphenicol O-Acetyltransferase; D003587:Cytomegalovirus / Q000235:genetics; D004742:Enhancer Elements, Genetic; D005796:Genes; D005814:Genes, Viral; D005822:Genetic Vectors; D027383:Papillomaviridae / Q000235:genetics*; D010957:Plasmids; D011401:Promoter Regions, Genetic; D013539:Simian virus 40 / Q000235:genetics; D014158:Transcription, Genetic; D014162:Transfection
D000123:Acetyltransferases; D015500:Chloramphenicol O-Acetyltransferase