Laminin-10 mediates basal and EGF-stimulated motility of human colon carcinoma cells via alpha(3)beta(1) and alpha(6)beta(4) integrins

Signals from the epidermal growth factor (EGF) receptor and integrin-depend ent adhesion to laminin contribute to the progression and metastasis of col onic tumors. However, little is know about the mechanisms by which these si gnals cooperate. Recently, we have reported that the colon cancer cell line LIM1215 secretes and adhere to autocrine laminin-10 via multiple integrin receptors and that EGF stimulates spreading of these cells on the same subs trate. In this report, we investigate the effect of EGF and laminin-10 on c olon cancer cell migration in vitro. EGF stimulates migration of LIM1215 ce lls in a wound healing assay. The response to EGF is inhibited by anti-EGF receptor antibody 528, the EGF receptor kinase inhibitor AG-1478, or the MA P kinase kinase inhibitor PD98059 but not the PI3-K inhibitor wortmannin. U sing Transwell migration chambers, we demonstrate that laminin-10 but not c ollagen-I, collagen-IV, or a commercial preparation of human placental lami nin is a potent motility factor for LIM1215 cells. The migration response t o laminin-10 is increased upon stimulation of the cells with EGF and correl ates with the up-regulation of alpha (6)beta (4) integrin expression as mea sured by analysis of Triton X-100-soluble cellular extracts. The results fr om integrin inhibition experiments indicate that basal migration on laminin -10 is mediated by alpha (3)beta (1) but not alpha (2)beta (1) nor alpha (6 )beta (4) integrins. Alpha(3) blocking antibodies also inhibited EGF-stimul ated chemokinetic migration of LIM1215 cells on laminin-10. However, in con trast to unstimulated cells, alpha (6) or beta (4) integrin-blocking antibo dies inhibited the migration of EGF-stimulated cells by up to 50%. Taken to gether, these results support the cooperative role of EGF receptor and lami nin-10 on colon cancer cell motility and suggest a critical role for both t he alpha (3)beta (1) and the alpha (6)beta (4) integrins in this process. ( C) 2001 Academic Press.