Signals from the
epidermal growth factor (
EGF) receptor and
integrin-dependent adhesion to
laminin contribute to the progression and
metastasis of colonic
tumors. However, little is know about the mechanisms by which these signals 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 substrate. In this report, we investigate the effect of
EGF and
laminin-10 on
colon cancer cell migration in vitro.
EGF stimulates migration of LIM1215 cells 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
MAP kinase kinase inhibitor
PD98059 but not the PI3-K inhibitor
wortmannin. Using Transwell migration chambers, we demonstrate that
laminin-10 but not
collagen-I,
collagen-IV, or a commercial preparation of human placental
laminin is a potent motility factor for LIM1215 cells. The migration response to
laminin-10 is increased upon stimulation of the cells with
EGF and correlates with the up-regulation of
alpha(6)beta(4) integrin expression as measured by analysis of Triton X-100-soluble cellular extracts. The results from
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-stimulated chemokinetic migration of LIM1215 cells on
laminin-10. However, in contrast to unstimulated cells, alpha(6) or beta(4)
integrin-
blocking antibodies inhibited the migration of
EGF-stimulated cells by up to 50%. Taken together, these results support the cooperative role of
EGF receptor and
laminin-10 on
colon cancer cell motility and suggest a critical role for both the alpha(3)beta(1) and the alpha(6)beta(4)
integrins in this process.