Dot-like focal contacts in adherent eosinophils, their redistribution into peripheral belts, and correlated effects on cell migration and protected zone formation.
We have examined the organization of F-actin and focal contacts in eosinophils and the functional consequences correlated with their predistribution. In activated eosinophils adherent to laminin, F-actin is localized in large, uniformly distributed, dot-like structures. Co-localized with the F-actin are a variety of typical components of focal contacts including: the alpha 6 and beta 1 integrins subunits (which mediate eosinophil adhesion to laminin), vinculin, talin, pp125FAK, paxillin, and tyrosine phosphorylated proteins, but not the beta 2 integrin subunit which is not involved in eosinophil adhesion to laminin. In unactivated eosinophils on laminin (which do not adhere well) and in activated eosinophils on non-adhesive surfaces (human serum albumin, tenascin, or a mixture of tenascin and laminin), dot-like accumulations of F-actin do not form. When activated eosinophils are incubated on laminin in the presence of low concentrations of 1,1'-ethylidenebis(L-tryptophan), a remarkable rearrangement occurs. Both conventional and confocal microscopy suggest that the dot-like structures rearrange from a uniform distribution into thick, peripheral belts surrounding each cell with few dot-like structures internal to the belt. Functional consequences occur as a result of this rearrangement of focal contacts and the cytoskeleton. The belt of focal contacts acts as a size-dependent permeability barrier; fluorescent 8 kDa polyethylene glycol does not label the substrate underneath the cells whereas 3.5 kDa polyethylene glycol labels the substrate uniformly. The formation of belts of focal contacts also blocks the ability of eosinophils to migrate on laminin in response to a chemotactic gradient. While the mechanism of 1,1'-ethylidenebis(L-tryptophan) action remains to be determined, experiments indicate that the function of 1,1'-ethylidenebis(L-tryptophan) is distinct from the function of drugs that alter tyrosine kinase and serine/threonine kinase activities. Our observations reveal a novel mechanism for regulating cell behavior in invasive cells in which the rearrangement of focal contacts and the associated cytoskeleton may control cell migration and the sequestration of secreted cytotoxic and degradatory molecules. We propose that 1,1'-ethylidenebis(L-tryptophan) may mimic the function of an endogenous regulator of eosinophil morphology and behavior.
Tourkin
A
A
Division of Rheumatology and Immunology, Medical University of South Carolina, Charleston 29425, USA.
Bonner
M
M
Mantrova
E
E
LeRoy
E C
EC
Hoffman
S
S
D000199:Actins / Q000097:blood; D002448:Cell Adhesion; D002465:Cell Movement; D004804:Eosinophils / Q000166:cytology*; D006801:Humans; D007797:Laminin / Q000097:blood; D010590:Phalloidine / Q000378:metabolism
D000199:Actins; D007797:Laminin; D010590:Phalloidine