The beta 1-integrin cytosolic domain optimizes phospholipase A2-mediated arachidonic acid release required for NIH-3T3 cell spreading

Inhibition of PLA2 activity and rescue by addition of exogenous AA was used to demonstrate that AA production is essential for integrin-mediated NIH-3 T3 murine cell spreading. Both AA release and cell spreading after attachme nt to a FN substrate were inhibited by the PLA2 inhibitor mepacrine. AA rel ease was essential for signaling spreading since the inhibition of spreadin g induced by mepacrine was overcome by exogenous AA. Cells ectopically expr essing full-length chicken pl-integrins both released AA and spread fully o n a substrate of anti-chicken beta 1-integrin monoclonal antibody, and inhi bition of PLA2 by mepacrine suppressed both spreading and AA release. Exoge nous AA also reversed this mepacrine-induced inhibition of spreading. The r ole of the beta 1-integrin cytosolic domain in AA release was examined by c omparing responses of cells expressing full-length chicken beta 1-integrins versus cells expressing a deletion mutant chicken beta 1-integrin with a t runcated cytosolic domain. Cells expressing a truncated chicken beta 1-inte grin released significantly less AA and failed to spread on the anti-chicke n beta 1-integrin antibody substrate. Furthermore, clustering full-length r eceptors with soluble antibody stimulated greater AA release than clusterin g of receptors having truncated cytosolic domains. These data suggest the b eta 1-integrin cytosolic domain is required for optimal PLA2 activation to produce AA necessary for cell spreading. (C) 1999 Academic Press.