Adenovirus enhancement of polyethylenimine-mediated transfer of regulated genes in differentiated cells.
Efficient gene transfer is a prerequisite for analysing regulation of transfected promoters. We combined the DNA binding property of the cationic polymer polyethylenimine (PEI) and the potent endocytic activity of adenovirus in a PEI-DNA-adenovirus complex which provided efficient plasmid delivery in differentiated cultured cells. We transfected 3T3-F442A adipocytes, C2.7 myocytes and FAO hepatoma cells with a construct containing the simian virus 40 promoter fused to the chloramphenicol acetyltransferase (CAT) gene, using a combination of PEI and 200 p.f.u. per cell of replication-deficient type 5 adenovirus. Resulting CAT activities varied according to the cell type reaching about 0.6, 8 and 38 units/mg protein for respectively 3T3-F442A, FAO and C2.7 cells. Increases in transfection efficiencies were 140- to 300-fold when compared with those obtained with PEI alone. Then we tested physiologically regulated promoters: the phosphoenolpyruvate carboxykinase gene promoter in 3T3-F442A or FAO cells and the hexokinase II gene promoter in C2.7 myocytes. Gene expression was appropriately increased by clofibrate, dexamethasone and insulin for 3T3-F442A, FAO and C2.7 cells, respectively. Thus, the combination of PEI and adenovirus is a simple, efficient, inexpensive and versatile method of gene transfer which is applicable to several differentiated cells and provides a physiologically coherent transgene regulation. We name this method PEI-adenofection.
Meunier-Durmort
C
C
Centre de Recherche sur l'Endocrinologie Moléculaire et le Développement, CNRS UPR 9078, Meudon, France.
Grimal
H
H
Sachs
L M
LM
Demeneix
B A
BA
Forest
C
C
D000256:Adenoviridae; D017667:Adipocytes; D002460:Cell Line; D018014:Gene Transfer Techniques; D006801:Humans; D008099:Liver / Q000166:cytology; D009206:Myocardium / Q000166:cytology; D011094:Polyethyleneimine
D011094:Polyethyleneimine