We have previously demonstrated that fibronectin (Fn) stimulates the proliferation of non-small cell lung carcinoma (NSCLC) cell growth through the induction of cyclooxygenase-2 (COX-2) and prostaglandin E2 secretion. Here, we demonstrate that NSCLC cells express mRNA and protein for the prostaglandin E2 receptor EP4 and that Fn enhances its stimulatory effect by inducing the expression of EP4, but not of EP1, EP2, and EP3 receptor subtypes. The effect of Fn on EP4 was inhibited by an antibody against alpha5beta1 integrin and by inhibitors of phosphoinositide 3-kinase (wortmannin) and extracellular signal-regulated kinase (PD98095), but not by inhibitors of protein kinase C (calphostin C), of protein kinase A (H-89), or of mammalian target of rapamycin (rapamycin). A COX-2 small interfering RNA was also inhibitory. Fn significantly increased AP-2 binding activity in the promoter of the EP4 gene, and AP-2 antisense oligonucleotides blocked Fn-induced EP4 expression. Using full-length and mutated EP4 promoter constructs, we found that Fn stimulation of EP4 gene expression was inhibited when one AP-2 site (-1000 bp) was mutated. Fn induced nuclear AP-2alpha protein expression through multiple signaling pathways. Our results indicate that Fn-induced NSCLC cell proliferation is mediated through EP4. Furthermore, they show that Fn induces EP4 expression through the activation of alpha5beta1-dependent signals that include induction of extracellular signal-regulated kinase and phosphoinositide 3-kinase pathways as well as expression of COX-2. These events lead to activation of the transcription factor AP-2alpha, which interacts with specific regions in the EP4 gene promoter, leading to transcription of the EP4 gene.