Protein kinase C-dependent mobilization of the α6β4 integrin from hemidesmosomes and its association with actin-rich cell protrusions drive the chemotactic migration of carcinoma cells

We explored the hypothesis that the chemo tactic migration of carcinoma cells that assemble hemidesmosomes involves the activation of a signaling pathway that releases the alpha 6 beta 4 integrin from these stable adhesion complexes and promotes its association with F-actin in cell protrusions enabling it to function in migration. Squamous carcinoma-derived A431 cells were used because they express alpha 6 beta 4 and migrate in response to EGF stimulation. Using function-blocking antibodies we show that the alpha 6 beta 4 integrin participates in EGF-stimulated chemotaxis and is required for lamellae formation on laminin-1, At concentrations of EGF that stimulate A431 chemotaxis (similar to 1 ng/ml), the alpha 6 beta 4 integrin is mobilized from hemidesmosomes as evidenced by indirect immunofluorescence microscopy using mAbs specific for this integrin and hemidesmosomal components and its loss from a cytokeratin fraction obtained by detergent extraction. EGF stimulation also increased the formation of lamellipodia and membrane ruffles that contained alpha 6 beta 4 in association with F-actin. Importantly, we demonstrate that this mobilization of alpha 6 beta 4 from hemidesmosomes and its redistribution to cell protrusions occurs by a mechanism that involves activation of protein kinase C-alpha and that it is associated with the phosphorylation of the beta 4 integrin subunit on serine residues. Thus, the chemotactic migration of A431 cells on laminin-1 requires not only the formation of F-actin-rich cell protrusions that mediate alpha 6 beta 4-dependent cell movement but also the disruption of alpha 6 beta 4-containing hemidesmosomes by protein kinase C.