Differential recruitment of α2β1 and α4β1 integrins to lipid rafts in Jurkat T lymphocytes exposed to collagen type IV and fibronectin

Collagen type IV (CnIV) and fibronectin (Fn) were used as ligands to study the distribution of alpha(2)beta(1) and alpha(4)beta(1) integrins in low-density, detergent-resistant microdomains (DRM) of Jurkat lymphocytes. CnIV-coated microspheres induced (optical trapping) the redistribution of GM(1)-associated fluorescence from the cell periphery to the area of contact. This was not observed in cells treated with beta-methyl cyclodextrin (MCD). Fn- or bovine serum albumin-coated microspheres did not modify the peripheral distribution of fluorescence. These observations were confirmed by confocal microscopy. Western blot analysis of cells exposed to surfaces coated with CnIV revealed that the alpha(2)-subunit was initially present at low levels in DRM, became strongly associated after 40 min, and returned to basal levels after 75 min. Fn induced a slight recruitment of the beta(1)-integrin alpha(4)-subunit in DRM after 5 and 10 min, followed by a return to basal levels. Neither CnIV nor Fn triggered significant changes in the distribution of the beta(1)-subunit in DRM. Fn- and CnIV-coated microspheres or surfaces coated with these ligands triggered a MCD-sensitive mobilization of Ca2+. MCD did not alter the state of the Ca2+ reserves. The differential distributions of the alpha(2)beta(1), and alpha(4)beta(1) integrins in DRM may provide one additional step in the regulation of outside-in signaling involving these integrins.