Human P45IA1 upstream regulatory sequences expressing the chloramphenicol acetyltransferase gene. Effect of Ha-MSV enhancer and comparison of transient with stable transformation assays.
Upstream sequences of the human P450IA1 gene were inserted into a promoterless expression vector (pSVO-cat) containing the chloramphenicol acetyltransferase (CAT) gene, with and without the Harvey murine sarcoma virus (Ha-MSV) core enhancer, and either plasmid was transfected into human breast carcinoma MCF-7 and MDA-231 and mouse hepatoma Hepa-1 cell lines. In most instances constitutive and inducible CAT activities in the transient CAT expression assay were similar (within 3-fold) to those in the stable transformation CAT assay (selection of G418-resistant colonies following co-transfection with pSV2-neo). In the case of Ha-MSV-containing constructs stably integrated in the two human breast cancer lines, however, CAT expression was more than two orders of magnitude greater than that transiently expressed in these cells. Since the major difference between these two assays is plasmid copy number, these data suggest the presence of limiting amounts of tissue-specific positive-control enhancer-binding factor(s) in the breast carcinoma cell lines.
Jaiswal
A K
AK
Laboratory of Development Pharmacology, National Institute of Child Health and Human Development, Bethesda, Maryland 20892.
Neuhold
L A
LA
Nebert
D W
DW
D000123:Acetyltransferases / Q000235:genetics*; D000818:Animals; D001943:Breast Neoplasms; D002460:Cell Line; D015500:Chloramphenicol O-Acetyltransferase; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics*; D004251:DNA Transposable Elements; D004742:Enhancer Elements, Genetic; D005260:Female; D005796:Genes; D005798:Genes, Bacterial; D005822:Genetic Vectors; D006251:Harvey murine sarcoma virus / Q000235:genetics; D006801:Humans; D008114:Liver Neoplasms, Experimental; D051379:Mice; D010957:Plasmids; D014162:Transfection
D004251:DNA Transposable Elements; D003577:Cytochrome P-450 Enzyme System; D000123:Acetyltransferases; D015500:Chloramphenicol O-Acetyltransferase