INTEGRIN-MEDIATED TYROSINE PHOSPHORYLATION AND REDISTRIBUTION OF PAXILLIN DURING NEURONAL ADHESION

Integrins are important receptors for neuronal adhesion to laminin, wh ich is one of the best promoters of neurite outgrowth. The present stu dy was carried out to understand some of the intracellular mechanisms which allow integrin-mediated neurite extension on laminin. In chicken retinal neurons, integrin-mediated adhesion to laminin and antibody-i nduced integrin clustering caused an increase in tyrosine phosphorylat ion of paxillin and focal adhesion kinase, The kinetics of phosphoryla tion and dephosphorylation of these proteins were different in neurons plated on laminin, compared to neurons in which the receptors were cl ustered with anti-integrin antibodies. Analysis of sucrose velocity gr adients could not show any association of paxillin and focal adhesion kinase with the integrin receptors. On the other hand, by using digito nin and milder extraction conditions, we found an enrichment of the ty rosine-phosphorylated polypeptides in the cytoskeletal, digitonin-inso luble fraction, Furthermore, neuronal adhesion induced a dramatic incr ease in the fraction of tyrosine-phosphorylated paxillin recovered wit h the digitonin-insoluble fraction, suggesting redistribution of this protein following adhesion of neurons to laminin. Localization studies on the detergent-insoluble fraction showed codistribution of both pax illin and focal adhesion kinase with integrins. We also found that pax illin tyrosine phosphorylation, but not paxillin expression, is develo pmentally regulated in the retina. Our results show that integrin-medi ated neuronal adhesion leads to the accumulation of a pool of highly p hosphorylated proteins at adhesion sites. There they may be responsibl e for the reorganization of the cytoskeleton, which underlies the proc ess of neurite extension. (C) 1997 Academic Press.