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 protei n 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. b ut does not inhibit She signaling and activation of ERK. We have then gener ated primary fibroblasts carrying a targeted deletion of the segment of bet a (1) subunit cytoplasmic domain required for activation of FAK Analysis of these cells indicates that FAK is not necessary for efficient tyrosine pho sphorylation 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 fa ctor stimulation. To examine if FAK could contribute to the activation of E RK in a cell type-specific manner through the Rap1/B-Raf pathway, we have u sed 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, introduc tion of B-Raf enhances and sustains integrin-mediated activation of ERK in wild-type primary fibroblasts but not in those carrying the targeted deleti on of the beta (1) cytoplasmic domain. Thus, the She and FAK pathways are a ctivated independently and function in a parallel fashion. Although not nec essary for signaling to ERK in primary fibroblasts, FAH may enhance and pro long integrin-mediated activation of ERK through p130(CAS), Crk, and Rap1 i n cells expressing B-Raf.