scholarly journals Nck-2, a Novel Src Homology2/3-containing Adaptor Protein That Interacts with the LIM-only Protein PINCH and Components of Growth Factor Receptor Kinase-signaling Pathways

1998 ◽  
Vol 9 (12) ◽  
pp. 3367-3382 ◽  
Author(s):  
Yizeng Tu ◽  
Fugang Li ◽  
Chuanyue Wu
Keyword(s):  
Growth Factor ◽  
Signaling Pathways ◽  
Growth Factor Receptor ◽  
Adaptor Protein ◽  
Sh3 Domain ◽  
Sh2 Domain ◽  
Kinase Signaling ◽  
Egf Receptors ◽  
Growth Factor Receptor Signaling ◽  
Receptor Kinase Signaling

Many of the protein–protein interactions that are essential for eukaryotic intracellular signal transduction are mediated by protein binding modules including SH2, SH3, and LIM domains. Nck is a SH3- and SH2-containing adaptor protein implicated in coordinating various signaling pathways, including those of growth factor receptors and cell adhesion receptors. We report here the identification, cloning, and characterization of a widely expressed, Nck-related adaptor protein termed Nck-2. Nck-2 comprises primarily three N-terminal SH3 domains and one C-terminal SH2 domain. We show that Nck-2 interacts with PINCH, a LIM-only protein implicated in integrin-linked kinase signaling. The PINCH-Nck-2 interaction is mediated by the fourth LIM domain of PINCH and the third SH3 domain of Nck-2. Furthermore, we show that Nck-2 is capable of recognizing several key components of growth factor receptor kinase-signaling pathways including EGF receptors, PDGF receptor-β, and IRS-1. The association of Nck-2 with EGF receptors was regulated by EGF stimulation and involved largely the SH2 domain of Nck-2, although the SH3 domains of Nck-2 also contributed to the complex formation. The association of Nck-2 with PDGF receptor-β was dependent on PDGF activation and was mediated solely by the SH2 domain of Nck-2. Additionally, we have detected a stable association between Nck-2 and IRS-1 that was mediated primarily via the second and third SH3 domain of Nck-2. Thus, Nck-2 associates with PINCH and components of different growth factor receptor-signaling pathways via distinct mechanisms. Finally, we provide evidence indicating that a fraction of the Nck-2 and/or Nck-1 proteins are associated with the cytoskeleton. These results identify a novel Nck-related SH2- and SH3-domain–containing protein and suggest that it may function as an adaptor protein connecting the growth factor receptor-signaling pathways with the integrin-signaling pathways.

scholarly journals Grb2 controls phosphorylation of FGFR2 by inhibiting receptor kinase and Shp2 phosphatase activity

2013 ◽  
Vol 200 (4) ◽  
pp. 493-504 ◽  
Author(s):  
Zamal Ahmed ◽  
Chi-Chuan Lin ◽  
Kin M. Suen ◽  
Fernando A. Melo ◽  
James A Levitt ◽  
...  
Keyword(s):  
Growth Factor ◽  
Phosphatase Activity ◽  
Tyrosine Phosphatase ◽  
Growth Factor Receptor ◽  
Adaptor Protein ◽  
Sh2 Domain ◽  
Receptor Kinase ◽  
Downstream Signaling ◽  
State Growth ◽  
Kinase Phosphorylation

Constitutive receptor tyrosine kinase phosphorylation requires regulation of kinase and phosphatase activity to prevent aberrant signal transduction. A dynamic mechanism is described here in which the adaptor protein, growth factor receptor–bound protein 2 (Grb2), controls fibroblast growth factor receptor 2 (FGFR2) signaling by regulating receptor kinase and SH2 domain–containing protein tyrosine phosphatase 2 (Shp2) phosphatase activity in the absence of extracellular stimulation. FGFR2 cycles between its kinase-active, partially phosphorylated, nonsignaling state and its Shp2-dephosphorylated state. Concurrently, Shp2 cycles between its FGFR2-phosphorylated and dephosphorylated forms. Both reciprocal activities of FGFR2 and Shp2 were inhibited by binding of Grb2 to the receptor. Phosphorylation of Grb2 by FGFR2 abrogated its binding to the receptor, resulting in up-regulation of both FGFR2’s kinase and Shp2’s phosphatase activity. Dephosphorylation of Grb2 by Shp2 rescued the FGFR2–Grb2 complex. This cycling of enzymatic activity results in a homeostatic, signaling-incompetent state. Growth factor binding perturbs this background cycling, promoting increased FGFR2 phosphorylation and kinase activity, Grb2 dissociation, and downstream signaling. Grb2 therefore exerts constitutive control over the mutually dependent activities of FGFR2 and Shp2.

Indirect recruitment of the signalling adaptor Shc to the fibroblast growth factor receptor 2 (FGFR2)

2008 ◽  
Vol 416 (2) ◽  
pp. 189-199 ◽  
Author(s):  
Annika C. Schüller ◽  
Zamal Ahmed ◽  
James A. Levitt ◽  
Kin M. Suen ◽  
Klaus Suhling ◽  
...  
Keyword(s):  
Growth Factor ◽  
Fibroblast Growth Factor Receptor ◽  
Growth Factor Receptor ◽  
Adaptor Protein ◽  
Sh2 Domain ◽  
Signalling Pathways ◽  
Fibroblast Growth ◽  
Src Homology ◽  
Receptor Family ◽  
Ptb Domain

The adaptor protein Shc (Src homology and collagen-containing protein) plays an important role in the activation of signalling pathways downstream of RTKs (receptor tyrosine kinases) regulating diverse cellular functions, such as differentiation, adhesion, migration and mitogenesis. Despite being phosphorylated downstream of members of the FGFR (fibroblast growth factor receptor) family, a direct interaction of Shc with this receptor family has not been described to date. Various studies have suggested potential binding sites for the Shc PTB domain (phosphotyrosine-binding domain) and/or the SH2 (Src homology 2) domain on FGFR1, but no interaction of full-length Shc with these sites has been reported in vivo. In the present study, we investigated the importance of the SH2 domain and the PTB domain in recruitment of Shc to FGFR2(IIIc) to characterize the interaction of these two proteins. Confocal microscopy revealed extensive co-localization of Shc with FGFR2. The PTB domain was identified as the critical component of Shc which mediates membrane localization. Results from FLIM (fluorescence lifetime imaging microscopy) revealed that the interaction between Shc and FGFR2 is indirect, suggesting that the adaptor protein forms part of a signalling complex containing the receptor. We identified the non-RTK Src as a protein which potentially mediates the formation of such a ternary complex. Although an interaction between Src and Shc has been described previously, in the present study we implicate the Shc SH2 domain as a novel mediator of this association. The recruitment of Shc to FGFR2 via an indirect mechanism provides new insight into the regulation of protein assembly and activation of various signalling pathways downstream of this RTK.

scholarly journals Convergence of integrin and growth factor receptor signaling pathways within the focal adhesion complex.

1995 ◽  
Vol 6 (10) ◽  
pp. 1349-1365 ◽  
Author(s):  
G E Plopper ◽  
H P McNamee ◽  
L E Dike ◽  
K Bojanowski ◽  
D E Ingber
Keyword(s):  
Growth Factor ◽  
Signaling Pathways ◽  
Phospholipase C ◽  
Focal Adhesion ◽  
Growth Factor Receptor ◽  
Growth Factor Receptors ◽  
Chemical Signaling ◽  
Focal Adhesion Complex ◽  
Growth Factor Receptor Signaling ◽  
Adhesion Complex

Extracellular matrix controls capillary endothelial cell sensitivity to soluble mitogens by binding integrin receptors and thereby activating a chemical signaling response that rapidly integrates with growth factor-induced signaling mechanisms. Here we report that in addition to integrins, growth factor receptors and multiple molecules that transduce signals conveyed by both types of receptors are immobilized on the cytoskeleton (CSK) and spatially integrated within the focal adhesion complex (FAC) at the site of integrin binding. FACs were rapidly induced in round cells and physically isolated from the remainder of the CSK after detergent-extraction using magnetic microbeads coated with fibronectin or a synthetic RGD-containing peptide. Immunofluorescence microscopy revealed that multiple signaling molecules (e.g., pp60c-src, pp125FAK, phosphatidylinositol-3-kinase, phospholipase C-gamma, and Na+/H+ antiporter) involved in both integrin and growth factor receptor signaling pathways became associated with the CSK framework of the FAC within 15 min after binding to beads coated with integrin ligands. Recruitment of tyrosine kinases to the FAC was also accompanied by a local increase in tyrosine phosphorylation, as indicated by enhanced phosphotyrosine staining at the site of integrin binding. In contrast, neither recruitment of signaling molecules nor increased phosphotyrosine staining was observed when cells bound to beads coated with a control ligand (acetylated low density lipoprotein) that ligates transmembrane scavenger receptors, but does not induce FAC formation. Western blot analysis confirmed that FACs isolated using RGD-beads were enriched for pp60c-src, pp125FAK, phospholipase C-gamma, and the Na+/H+ antiporter when compared with intact CSK or basal cell surface preparations that retained lipid bilayer. Isolated FACs were also greatly enriched for the high affinity fibroblast growth factor receptor flg. Most importantly, isolated FACs continued to exhibit multiple chemical signaling activities in vitro, including protein tyrosine kinase activities (pp60c-src and pp125FAK) as well as the ability to undergo multiple sequential steps in the inositol lipid synthesis cascade. These data suggest that many of the chemical signaling events that are induced by integrins and growth factor receptors in capillary cells may effectively function in a "solid-state" on insoluble CSK scaffolds within the FAC and that the FAC may represent a major site for signal integration between these two regulatory pathways. Future investigations into the biochemical and biophysical basis of signal transduction may be facilitated by this method, which results in isolation of FACs that retain the CSK framework as well as multiple associated chemical signaling activities.

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