Differences between normal and transformed murine fibroblasts in the expression of various promoter/enhancer-chloramphenicol acetyltransferase constructs.
High levels of poly(A)+ RNAs homologous to certain endogenous retrovirus-related DNA sequences are frequently seen in carcinogen-transformed rodent cells. To explore the underlying mechanism, transient expression assays were done to determine whether carcinogen- or radiation-transformed C3H 10T 1/2 cell lines differ from normal 10T 1/2 cells in terms of their ability to express the chloramphenicol acetyltransferase (CAT) gene when it is linked to various promoter/enhancer sequences, including two independently isolated intracisternal A particle (IAP) long terminal repeat sequences designated prcm and pMIA6. We found that with several constructs, CAT activity was always 3- to 10-fold higher in the transformed 10T 1/2 cell lines than in the normal 10T 1/2 cells. The prcm-CAT construct displayed the highest CAT activity in both the normal and transformed C3H 10T 1/2 cells. Studies with 32P-labeled prcm-CAT DNA and Southern blot analyses indicated that the differences in CAT activity between normal and transformed cells were not due to greater uptake or retention of the transfected DNA by the transformed cells. Competition studies provided evidence that factors required for the expression of the prcm-CAT construct are present in limited amounts in normal 10T 1/2 cells and in excess amounts in transformed 10T 1/2 cells. These putative factors may play a role in the increased expression of endogenous retrovirus-related sequences in the transformed cells.
Begemann
M
M
Comprehensive Cancer Center, Columbia University, New York, New York 10032.
Hsiao
W L
WL
Weinstein
I B
IB
D000818:Animals; D001358:Avian Sarcoma Viruses; D001564:Benzo(a)pyrene; D002461:Cell Line, Transformed / Q000528:radiation effects; D015500:Chloramphenicol O-Acetyltransferase; D004742:Enhancer Elements, Genetic; D005347:Fibroblasts / Q000166:cytology; D007426:Genes, Intracisternal A-Particle; D051379:Mice; D011401:Promoter Regions, Genetic; D012016:Reference Values; D012091:Repetitive Sequences, Nucleic Acid; D013539:Simian virus 40
D001564:Benzo(a)pyrene; D015500:Chloramphenicol O-Acetyltransferase