Nucleic Acids Research, 1991, Vol. 19, No. 7 1563-1569
© 1991
MOLECULAR BIOLOGY |
Cooperation between structural elements in hormonoregulated transcription from the mouse mammary tumor virus promoter
Unité de recherches sur les communications hormonales, INSERM U-33, Hopital du Kremlin Bicêtre, 78 rue du Général Leclerc 94275 Bicêtre Cedex, France
* To whom correspondence should be addressed
Received December 11, 1990. Revised March 4, 1991. Accepted March 4, 1991.
The mouse mammary tumor virus (MMTV) promoter is under the control of several types of regulatory agents. The proximal promoter within the long terminal repeat (LTR), from –200 to the CAP site and its regulation by steroid hormones have been extensively studied. However the precise role of sequences located upstream of this region remain unclear. We have constructed MMTV LTR deletion mutants coupled to the luciferase reporter gene and assayed their activities after transient transfection into transformed mammary epithelial cells (341) and Immortalized fibroblasts (NIH-3T3). In the absence of hormone, the MMTV promoter is almost silent, and deletions in the LTR have no significant effect on basal activity. In the presence of hormone, deletions spanning from the 5'-end to –455 have only slight effects on luciferase levels. In contrast, deletion of the region spanning from –450 to –201 leads to a dramatic decrease in transcription. A substantial decrease, more marked in 34i cells, is also clear when 90bp between –290 and –201 are deleted. At least one element cooperating positively with the glucocorticoid response element (GRE) is present between –223 and –201, as supported by the results of substitution mutation experiments. In 341 cell line, dexamethasone stimulates the MMTV LTR transcriptional activity to a level comparable to that of SV40. In contrast, in NIH-3T3 cells, MMTV promoter induciblllty is weak. This results from a glucocorticoid receptor content 10-fold lower in NIH-3T3 cells than in 341 cells. Transfection of a glucocorticoid receptor expression plasmid allows recovery of a high inducibility of the MMTV promoter. This was true with all the MMTV LTR mutants studied here and suggests that NIH-3T3 cells possess all the factors necessary to cooperate with the steroid hormone in order to achieve a high transcriptional activity.
+Present address: Institut Cochin de Génétique Moléculaire, INSERM U-152, Hopital Cochin, 27 rue du Faubourg Saint Jacques, 75014 Paris, France
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