Distinct roles of the adaptor protein Shc and focal adhesion kinase in integrin signaling to ERK

It has been proposed that integrins activate ERK through the adaptor protein She independently of focal adhesion kinase (FAK) or through FAK acting on multiple target effecters, including She. We show that disrupt ion of the actin cytoskeleton by cytochalasin D causes a complete inhibition of FAK. but does not inhibit She signaling and activation of ERK. We have then generated primary fibroblasts carrying a targeted deletion of the segment of beta (1) subunit cytoplasmic domain required for activation of FAK Analysis of these cells indicates that FAK is not necessary for efficient tyrosine phosphorylation of She, association of She with Grb2, and activation of ERK in response to matrix adhesion. In addition, integrin-mediated activation of FAK does not appear to be required for signaling to ERK following growth factor stimulation. To examine if FAK could contribute to the activation of ERK in a cell type-specific manner through the Rap1/B-Raf pathway, we have used Swiss-3T3 cells, which in contrast to primary fibroblasts express B-Raf. Dominant negative studies indicate that She mediates the early phase and peak, whereas FAK, p130(CAS), Crk, and Rap1 contribute to the late phase of integrin-dependent activation of ERK in these cells. In addition, introduction of B-Raf enhances and sustains integrin-mediated activation of ERK in wild-type primary fibroblasts but not in those carrying the targeted deletion of the beta (1) cytoplasmic domain. Thus, the She and FAK pathways are activated independently and function in a parallel fashion. Although not necessary for signaling to ERK in primary fibroblasts, FAH may enhance and prolong integrin-mediated activation of ERK through p130(CAS), Crk, and Rap1 in cells expressing B-Raf.