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 Shc independently of focal adhesion kinase (FAK) or through FAK acting on multiple target effectors, including Shc. We show that disruption of the actin cytoskeleton by cytochalasin D causes a complete inhibition of FAK but does not inhibit Shc 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 Shc, association of Shc 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 Shc 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 Shc and FAK pathways are activated independently and function in a parallel fashion. Although not necessary for signaling to ERK in primary fibroblasts, FAK may enhance and prolong integrin-mediated activation of ERK through p130(CAS), Crk, and Rap1 in cells expressing B-Raf.
Barberis
L
L
Cellular Biochemistry and Biophysics Program, Memorial Sloan-Kettering Cancer Center, New York, New York 10021, USA.
Wary
K K
KK
Fiucci
G
G
Liu
F
F
Hirsch
E
E
Brancaccio
M
M
Altruda
F
F
Tarone
G
G
Giancotti
F G
FG
D016475:3T3 Cells; D048868:Adaptor Proteins, Signal Transducing; D033942:Adaptor Proteins, Vesicular Transport; D000818:Animals; D050719:Crk-Associated Substrate Protein; D015638:Cytochalasin D / Q000378:metabolism; D004789:Enzyme Activation; D005347:Fibroblasts / Q000201:enzymology; D005353:Fibronectins / Q000378:metabolism; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; D016023:Integrins / Q000502:physiology*; D051379:Mice; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D010750:Phosphoproteins / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D011506:Proteins / Q000502:physiology*; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D050737:Proto-Oncogene Proteins c-crk; D050717:Retinoblastoma-Like Protein p130; D055765:Shc Signaling Adaptor Proteins; D015398:Signal Transduction; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D020827:rap1 GTP-Binding Proteins / Q000378:metabolism; D018909:src Homology Domains / Q000502:physiology*
D048868:Adaptor Proteins, Signal Transducing; D033942:Adaptor Proteins, Vesicular Transport; C493842:Bcar1 protein, mouse; D050719:Crk-Associated Substrate Protein; D005353:Fibronectins; D016023:Integrins; D010750:Phosphoproteins; D011506:Proteins; D011518:Proto-Oncogene Proteins; D050737:Proto-Oncogene Proteins c-crk; D050717:Retinoblastoma-Like Protein p130; D055765:Shc Signaling Adaptor Proteins; C528034:Shc1 protein, mouse; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D015638:Cytochalasin D; D011505:Protein-Tyrosine Kinases; D051417:Focal Adhesion Kinase 1; D051416:Focal Adhesion Protein-Tyrosine Kinases; C496051:Ptk2 protein, mouse; D020928:Mitogen-Activated Protein Kinases; D020827:rap1 GTP-Binding Proteins