Constitutive ap‐1 dna binding and transactivating ability of malignant but not benign mouse epidermal cells

Abstract

The mouse epidermal cell line 308 contains an activated Ha-ras gene and forms benign papillomas when transplanted to the skin of athymic nude mice. A radiation-associated malignant variant of this cell line, 308-10Gy5, has been isolated and shown to form squamous cell carcinomas in nude mice. To further examine the molecular events involved in malignant conversion of 308-10Gy5, we assessed the activator protein-1 (AP-1) binding and transactivating ability of 308 and 308-10Gy5. In nuclear protein extracts of 308, AP-1 sequence-specific binding to an oligonucleotide containing a single high-affinity AP-1 binding site was induced by the tumor promoter 12-O-tetradecanoylphorbol-13-acetate, as determined by gel shift analysis. Nuclear extracts of 308-10Gy5 bound to the AP-1 oligonucleotide without treatment with tumor promoters. Not only was sequence-specific AP-1 DNA binding constitutively active in malignant versus benign tumor cells, but so was transactivation of a unique AP-1-responsive chloramphenicol acetyltransferase reporter construct, pTiCTaK. Constitutive transactivation of this AP-1-responsive reporter construct was observed in the malignant but not the benign tumor cells. Furthermore, steady-state transcript levels of the tumor-associated AP-1-responsive genes stromelysin, urokinase-type plasminogen activator, c-jun, and c-fos were higher in malignant 308-10Gy5 cells than in benign 308 cells. These results suggest that acquisition of constitutive AP-1 DNA binding and transactivation can result in sustained deregulation of gene expression. While malignant progression in keratinocytes is probably not due solely to the acquisition of constitutive cellular AP-1 activity, the effect of deregulated expression of AP-1-regulated genes, especially basement membrane-degrading enzymes, may be functionally related to malignant conversion.

Krp1, a novel kelch related protein that is involved in pseudopod elongation in transformed cells

“…Transformation of ®broblasts by oncogenes, such as raf and ras, have also been shown to be dependent on AP-1 activity (Johnson et al, 1996;Lloyd et al, 1991;Rapp et al, 1994;Suzuki et al, 1994). The role of AP-1 in transformation is not limited to ®broblasts, since mouse and human squamous cell carcinomas expressing the dominant negative Jun mutant have reduced tumourigenicity and in vitro invasiveness (Domann et al, 1994;Dong et al, 1997;Malliri et al, 1998).…”

Activation of p38 MAP kinase and ERK are required for ultraviolet-B induced c-fos gene expression in human keratinocytes

“…In vivo studies have shown that blocking AP-1 transactivation in malignant cell lines could inhibit their ability to form tumors upon subcutaneous injection into athymic nude mice (Domann et al, 1994). Furthermore, inactivation of AP-1 by aspirin and salicylates or retinoic acid correlated with inhibition of tumor promoter induced transformation and tumorgenesis Huang et al, 1997).…”

Tumor promoter-induced ornithine decarboxylase gene expression occurs independently of AP-1 activation

“…From studies utilizing reporter constructs measuring AP-1-dependent gene expression, the induction of AP-1 transactivation also has been shown to be essential for tumor promotion and progression in cell culture models (Dong et al, 1994(Dong et al, , 1997Li et al, 1998;Domann et al, 1994). In JB6 and human keratinocyte models, expression of dominant negative c-jun can block induced AP-1 activity and neoplastic transformation (Dong et al, 1994(Dong et al, , 1997Li et al, 1998;Domann et al, 1994).…”

“…In JB6 and human keratinocyte models, expression of dominant negative c-jun can block induced AP-1 activity and neoplastic transformation (Dong et al, 1994(Dong et al, , 1997Li et al, 1998;Domann et al, 1994). TPA-induced upregulation of the AP-1 transcription factor complex occurs prior to TPAinduced ODC gene expression.…”