A common response element mediates differential effects of phorbol esters and forskolin on type‐1 plasminogen activator inhibitor gene expression in human breast carcinoma cells

We have characterized regulation of type‐1 plasminogen activator inhibitor (PAI‐1) gene expression by phorbol 12‐myristate 13‐acetate (PMA) and the cAMP‐inducing agent forskolin in the human breast carcinoma cell line MCF‐7. PMA caused a strong induction of PAI‐1, while forskolin suppressed the PMA response. Transfection experiments with fusion genes showed that sequences mediating PMA induction as well as forskolin suppression were present between base pairs –100 and –30 of the 57prime;‐flanking region of the PAI‐1 gene. The region was found to contain two Sp1 binding sites. A proximal sequence in the region, TGAGTTCA (P box), with sequence similarity to phorbol ester response elements (TRE) as well as to cAMP response elements (CRE), bound a low‐abundance, as yet unidentified nuclear protein in MCF‐7 cells. This sequence had a higher affinity to purified c‐jun homodimer than to c‐jun/c‐fos heterodimer in MCF‐7 nuclear extracts; it had no affinity to the proteins binding to CRE consensus sequences in these extracts. A distal TRE‐like sequence, TGAGTGG (D box), had a weak affinity to c‐jun/c‐fos heterodimer and c‐jun homodimer; binding of proteins to this sequence was facilitated by binding of proteins to the P box. Both the P box and the D box were necessary for PMA responsiveness, suggesting a cooperativity between the two binding sites. A mutation of the P box removing the CRE similarity abolished the forskolin suppression of the PMA response. We propose that the protein kinase C and the protein kinase A signal‐transduction pathways, with opposite effects on PAI‐1 gene expression, converge by modulating differently P‐box‐binding proteins. Copyright © 1994, Wiley Blackwell. All rights reserved