THE INTEGRIN ALPHA-6-BETA-4 FUNCTIONS IN CARCINOMA CELL-MIGRATION ON LAMININ-1 BY MEDIATING THE FORMATION AND STABILIZATION OF ACTIN-CONTAINING MOTILITY STRUCTURES

Functional studies on the alpha 6 beta 4 integrin have focused primari ly on its role in the organization of hemidesmosomes, stable adhesive structures that associate with the intermediate filament cytoskeleton. In this study, we examined the function of the alpha 6 beta 4 integri n in clone A cells, a colon carcinoma cell line that expresses alpha 6 beta 4 but no alpha 6 beta 1 integrin and exhibits dynamic adhesion a nd motility on laminin-1. Time-lapse videomicroscopy of clone A cells on laminin-1 revealed that their migration is characterized by filopod ial extension and stabilization followed by lamellae that extend in th e direction of stabilized filopodia. A function-blocking mAb specific for the alpha 6 beta 4 integrin inhibited clone A migration on laminin -1. This mAb also inhibited filopodial formation and stabilization and lamella formation. Indirect immunofluorescence microscopy revealed th at the alpha 6 beta 4 integrin is localized as discrete clusters in fi lopodia, lamellae, and retraction fibers. Although beta 1 integrins we re also localized in the same structures, a spatial separation of thes e two integrin populations was evident. In filopodia and lamellae, a s triking colocalization of the alpha 6 beta 4 integrin and F-actin was seen. An association between alpha 6 beta 4 and F-actin is supported b y the fact that alpha 6 beta 4 integrin and actin were released from c lone A cells by treatment with the F-actin-severing protein gelsolin a nd that alpha 6 beta 4 immunostaining at the marginal edges of clone A cells on laminin-1 was resistant to solubilization with Triton X-100. Cytokeratins were not observed in filopodia and lamellipodia. Moreove r, alpha 6 beta 4 was extracted from these marginal edges with a Tween -40/deoxycholate buffer that solubilizes the actin cytoskeleton but no t cytokeratins. Three other carcinoma cell lines (MIP-101, CCL-228, an d MDA-MB-231) exhibited alpha 6 beta 4 colocalized with actin in filop odia and lamellae. Formation of lamellae in these cells was inhibited with an alpha 6-specific antibody. Together, these results indicate th at the alpha 6 beta 4 integrin functions in carcinoma migration on lam inin-1 through its ability to promote the formation and stabilization of actin-containing motility structures.