AP-1 mediates stretch-induced expression of HB-EGF in bladder smooth muscle cells

Mechanical induction of growth factor synthesis may mediate adaptive respon ses of smooth muscle cells (SMC) to increases in physical load. We previous ly demonstrated that cyclic mechanical stretch induces expression of the SM C, fibroblast, and epithelial cell mitogen heparin-binding epidermal growth factor-like growth factor (HB-EGF) in bladder SMC, an observation that sug gests that this growth factor may be involved in compensatory bladder hyper trophy. In the present study we provide evidence that the activator protein -1 (AP-1) transcription factor plays a critical role in this mechanoinducti on process. Rat bladder SMC were transiently transfected with a series of 5 ' deletion mutants of a promoter-reporter construct containing 1.7 kb of th e mouse HB-EGF promoter that Tvas previously shown to be stretch responsive . The stretch-mediated increase in promoter activity was completely ablated with deletion of nucleotide positions -1301 to -881. Binding of AP-1, as e valuated by electrophoretic mobility shift assay, to a synthetic oligonucle otide containing an AP-1 binding site increased in response to stretch, and binding was inhibited by excess unlabeled DNA corresponding to nucleotides -993 to -973 from the HB-EGF promoter, a region that contains a previously recognized composite AP-1/Ets site. Stretch-induced promoter activity was significantly inhibited by site-directed mutagenesis of the AP-1 or Ets com ponents of this site. Consistent with the promoter and gel-shift studies, c urcumin, an inhibitor of AP-1 activation, suppressed the HB-EGF mRNA induct ion after stretch. Stretch also specifically increased mRNA levels for matr ix metalloproteinase (MMP)-1, the promoter of which contains a functional A P-1 element, but not for MMP-2, the promoter of which does not contain an A P-1 element. The stretch response of the MMP-1 gene was also completely inh ibited by curcumin. Collectively, these findings indicate that AP-1-mediate d transcription plays an important role in the regulation of gene expressio n in bladder muscle in response to mechanical forces.