Enhanced expression of matrix metalloproteinase- 9 ( MMP-9) is associated with human lung tumor invasion and/ or metastasis. We have demonstrated that fibronectin (FN), a matrix glycoprotein, stimulates human non-small cell lung carcinoma (NSCLC) cell proliferation. The current study examines the effect of FN on MMP-9 expression in NSCLC cells. We show that FN increases MMP-9 protein, mRNA expression, and gelatinolytic activity in NSCLC cells. The integrin alpha 5 beta 1 mediated the effects of FN because alpha 5 small interfering RNA blocked FN-stimulated MMP- 9 protein expression, and also abrogated FN-induced phosphorylation of ERK and phosphatidylinositol 3-kinase ( PI3K) signals. The inhibitor of ERK, PD98095, and of PI3K, wortmannin, but not that of protein kinase A, H89, of Rho kinase, Y- 27632, of mTOR, rapamycin, or of JNK, SP600125, prevented FN- induced MMP-9 gelatinolytic activity and gene expression. FN enhanced MMP-9 gene promoter activity; however, there was no response to FN in DNA constructs with an AP-1 site mutation. FN increased AP-1 DNA binding activity, and this was abrogated by cyclic AMP response element decoy oligonucleotides, which also diminished FN- induced MMP-9 promoter activity. FN increased the expression of the AP- 1 subunit c- Fos protein, but not in the presence of PD98095 and wortmannin. The AP-1 inhibitor, nordihydroguaiaretic acid, and a c- Fos small interfering RNA eliminated the effect of FN on MMP-9 expression. This study indicates that FN, by binding to the integrin alpha 5 beta 1 receptor, stimulates the expression of MMP- 9 through increased AP- 1/ DNA binding and c- Fos protein expression via ERK and PI3K signaling pathways. The data unveils a novel mechanism by which FN could promote NSCLC cell invasion and metastasis.
