Multiple Promoter Elements Are Required for the Stimulatory Effect of Insulin on Human Collagenase-1 Gene Transcription

Abstract

Several of the complications seen in patients with both type I and type II diabetes mellitus are associated with alterations in the expression of matrix metalloproteinases. To identify the cis-acting elements that mediate the stimulatory effect of insulin on collagenase-1 (matrix metalloproteinase-1) gene transcription a series of collagenase-chloramphenicol acetyltransferase (CAT) fusion genes were transiently transfected into HeLa cells. Multiple promoter elements, including an Ets and activator protein-1 (AP-1) motif, were required for the effect of insulin. The AP-1 motif appears to be a target for insulin signaling because it is sufficient to mediate an effect of insulin on the expression of a heterologous fusion gene, whereas the data suggest that the Ets motif acts to enhance the effect of insulin mediated through the AP-1 motif. Multiple promoter elements were also required for the stimulatory effect of phorbol esters on collagenase-CAT gene transcription, and the AP-1 motif was also a target for phorbol ester signaling. However, the cis-acting elements required for the effects of insulin and phorbol esters were not identical. Moreover, phorbol esters were a much more potent inducer of collagenase-CAT gene transcription than insulin, a difference that may be explained by selective effects of insulin and phorbol esters on AP-1 expression.

Insulin-mediated activation of activator protein-1 through the mitogen-activated protein kinase pathway stimulates collagenase-1 gene transcription in the MES 13 mesangial cell line

“…Moreover, it is unclear whether this change in collagenase expression represents a direct effect of insulin as opposed to an indirect effect mediated through changes in glucose concentration. Nevertheless, a direct stimulatory effect of insulin on collagenase fusion gene transcription has been observed in NIH 3T3 (Medema et al 1991), CHO.T (Rutter et al 1995) and HeLa cells (Chapman et al 1999).…”

Transcriptional repression of MMP-1 by p21SNFT and reduced in vitro invasiveness of hepatocarcinoma cells

“…HeLa cells were transiently cotransfected with chloramphenicol acetyltransferase (CAT) constructs driven by the element in triplicate, and with p21 SNFT expression construct as indicated ( Figure 3a). As previously demonstrated (Chapman et al, 1999), in the absence of p21 SNFT (0 mg), the Ets mutant construct promotes approximately 20-25% of the CAT activity produced by the wild-type element, while the AP-1-binding region mutant is inactive. In response to p21 SNFT , the wild-type element is repressed up to 75-80% and the Ets-binding region mutant is repressed to basal levels.…”

“…The pEGFP-SNFT, pQE30Fosdb, pQE32Jundb (Bower et al, 2002), pCI/SNFT, GST-SNFT (Iacobelli et al, 2000), his-Ets-1 DBD (Nikolajczyk et al, 1996), and Ets/AP-1-CAT wildtype and mutant constructs (Chapman et al, 1999) have been described previously. The MMP-1 promoter luciferase construct was generated by cloning the human MMP-1 promoter region (À610 to þ 61) from the pUC9pCllase II genomic cosmid (Angel et al, 1987) into the pGL3basic luciferase vector (Promega Corp., Madison, WI, USA) multiple cloning site.…”

“…While the activity of MMP-1 is regulated at several levels, control of MMP-1 transcription at the proximal 600 bp region of the promoter is significant and highly characterized (Angel et al, 1987;Gutman and Wasylyk, 1990;Jonat et al, 1992;Borden and Heller, 1997;Westermarck et al, 1997;Chapman et al, 1999;Nagese and Woessner, 1999;Westermark and Kahari, 1999). A major enhancer element of the promoter is the À88 Ets/AP-1 composite element.…”

“…This element consists of a consensus AP-1 site, first identified as a 12-O-tetradecanoyl-phorbol-13-acetate (TPA) responsive element (TRE) (Angel et al, 1987), adjacent to a nonconsensus Ets-binding element. Ets and AP-1 proteins have been shown to bind and transactivate transcription from this element alone and together (Angel et al, 1987;Gutman and Wasylyk, 1990;Buttice et al, 1996;Chapman et al, 1999;Ozaki et al, 2002).…”