A quantitative study of the recruitment potential of all intracellular tyrosine residues on EGFR, FGFR1 and IGF1R

Kaushansky, Alexis; Gordus, Andrew; Chang, Bryan H; Rush, A. John; MacBeath, Gavin

doi:10.1039/b801018h

Cited by 52 publications

(66 citation statements)

References 61 publications

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“…1D) at ∼0.52 s −1 . This result was surprising in two regards: First, the Grb2 SH2 domain binds to multiple EGFR p-Tyr sites in vitro, with varying affinities (12), which presumably arise partly from different dissociation rates. Heterogeneity is known to affect the shape of the hazard functions (16).…”

Section: Resultsmentioning

confidence: 85%

“…During the last decade, an expanding assembly of quantitative datasets has accumulated regarding tyrosine phosphorylation signaling, including the topological pattern of phosphorylation (7)(8)(9)(10), the temporal dynamics of phosphorylation (10,11), and characterization of the SH2/p-Tyr binding affinity (12,13). On the other hand, the in vivo assembly and turnover kinetics of the SH2 complexes at p-Tyr sites remain poorly understood, partly because existing methods measure mostly the ensemble properties of the system.…”

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confidence: 99%

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Fast rebinding increases dwell time of Src homology 2 (SH2)-containing proteins near the plasma membrane

Oh¹,

Ogiue-Ikeda

Jadwin

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

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Receptor tyrosine kinases (RTKs) control a host of biological functions by phosphorylating tyrosine residues of intracellular proteins upon extracellular ligand binding. The phosphotyrosines (p-Tyr) then recruit a subset of ∼100 Src homology 2 (SH2) domaincontaining proteins to the cell membrane. The in vivo kinetics of this process are not well understood. Here we use total internal reflection (TIR) microscopy and single-molecule imaging to monitor interactions between SH2 modules and p-Tyr sites near the cell membrane. We found that the dwell time of SH2 modules within the TIR illumination field is significantly longer than predictions based on chemical dissociation rate constants, suggesting that SH2 modules quickly rebind to nearby p-Tyr sites after dissociation. We also found that, consistent with the rebinding model, the effective diffusion constant is negatively correlated with the respective dwell time for different SH2 domains and the dwell time is positively correlated with the local density of RTK phosphorylation. These results suggest a mechanism whereby signal output can be regulated through the spatial organization of multiple binding sites, which will prompt reevaluation of many aspects of RTK signaling, such as signaling specificity, mechanisms of spatial control, and noise suppression.reaction | epidermal growth factor receptor | diffusion-limited dissociation T he Src homology 2 (SH2) domain (1) controls many cellular activities by binding specifically to tyrosine-phosphorylated peptides (2). The tyrosine phosphorylation signal often originates from the cell surface through activations of receptor tyrosine kinases (RTKs). Humans, for example, have 58 known RTKs (3), each of which could generate diverse phosphorylation patterns at their cytosolic tails as well as at other associated proteins. These phosphorylation patterns are "read" mainly by a repertoire of effector proteins containing the SH2 domain (2, 4, 5), via recruitment of these effector molecules to the phosphotyrosine (pTyr) sites. For example, the epidermal growth factor receptor (EGFR), a well-known model system for studying phosphotyrosine signaling, is activated by epidermal growth factor (EGF), a small protein ligand. The binding of EGF turns on the kinase activity of EGFR molecules, which then autophosphorylate themselves at more than 20 tyrosine sites on the cytoplasmic tail. The phosphotyrosines are thought to serve as docking sites for SH2-containing molecules, such as Grb2, which bind to these sites with varying affinity. Downstream signals are propagated by the SH2 proteins, which in many cases are enzymes whose substrates are localized to the plasma membrane. In other cases such as Grb2, the SH2 protein serves to recruit additional downstream effector proteins such as the Ras activator Sos. EGFR family members also play critical roles in cancer development and progression and are frequently disregulated in tumors (6).During the last decade, an expanding assembly of quantitative datasets has accumulated regarding tyrosine p...

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Section: Resultsmentioning

confidence: 85%

mentioning

confidence: 99%

Fast rebinding increases dwell time of Src homology 2 (SH2)-containing proteins near the plasma membrane

Oh¹,

Ogiue-Ikeda

Jadwin

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

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“…138,192 Substantial qualitative overlap in the recruitment profiles of IGF1R, FGFR1 and EGFR has recently been demonstrated. 139 Figure 3 shows that IGF1R and FGFR2b share common downstream signal transduction pathways which involve the action of androgens, GH, IGF-1 and FGFs. IGFR1 signaling upregulates androgen synthesis, the conversion of less potent to more potent androgens and AR activation.…”

Section: Interleukin-1α Network In Keratinocytesmentioning

confidence: 99%

“…137 Comparison of EGFR, FGFR1 and IGF1R downstream signaling pathways show, that for the most part, a similar repertoire of signaling proteins are recruited and activated by these tyrosine kinase receptors. 138,139 A cooperative interaction between EGFR, FGFR2b-and IGF1R-signaling in the pilosebaceous follicle has to be expected in mesenchymal-epithelial interactions for sebaceous gland development and homeostasis in the adult tissue.…”

Section: Fgfr2b-signaling In Acne Vulgarismentioning

confidence: 99%

Role of FGFR2-signaling in the pathogenesis of acne

Melnik

2009

Dermato-Endocrinology

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It is the purpose of this review to extend our understanding of the fibroblast growth factor (FGF) receptor-2b-signaling network in the pathogenesis of acne. A new concept of the role of FGFR2b-signaling in dermal-epithelial interaction for skin appendage formation, pilosebaceous follicle homeostasis, comedogenesis, sebaceous gland proliferation and lipogenesis is presented. The FGFR2-gain-of-function mutations in Apert syndrome and unilateral acneiform nevus are most helpful model diseases pointing the way to androgen-dependent dermalepithelial FGFR2-signaling in acne. Androgen-mediated upregulation of FGFR2b-signaling in acne-prone skin appears to be involved in the pathogenesis of acne vulgaris. In organotypic skin cultures, keratinocyte-derived interleukin-1α stimulated fibroblasts to secrete FGF7 which stimulated FGFR2b-mediated keratinocyte proliferation. Postnatal deletion of FGFR2b in mice resulted in severe sebaceous gland atrophy. The importance of FGFR2b in sebaceous gland physiology is further supported by the mode of action of anti-acne agents which have been proposed to attenuate FGFR2b-signaling. Downregulation of FGFR2b-signaling by isotretinoin explains its therapeutic effect in acne. Downregulation of FGFR2b-signaling during the first trimester of pregnancy disturbs branched morphogenesis and explains retinoid embryotoxicity. Insulin-like growth factor-1 (IGF-1), the mediator of growth hormone during puberty, intracts with androgen-dependent FGFR2b-signaling and links androgen-and FGF-mediated signal transduction important in sebaceous gland homeostasis. The search for a follicular defect in the dermalepithelial regulation of growth factor-signaling in acne-prone skin appears to be a most promising approach to clarify the pathogenesis of acne.

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“…When the ligand binds with its receptor, it induces autophosphorylation of tyrosine (Tyr) residues on the intracellular carboxyl terminal tail of the receptor (1,2). Activated-RTKs can associate with several adaptor proteins containing Src-homology 2 (SH2) domain, such as growth factor receptor bound protein 2 (Grb2), Src homologous and collagen (Shc), and phospholipase C Á (PLCÁ).…”

Section: Introductionmentioning

confidence: 99%

Basic study on SH2 domain of Grb2 as a molecular probe for detection of RTK activation

Saito¹,

Furukawa

Arano

et al. 2010

Int J Oncol

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Abstract. Epidermal growth factor receptor (EGFR) and other receptor tyrosine kinases (RTKs) are overexpressed and/or mutated in various cancers and their abnormal activation is implicated in carcinogenesis. We explored the possibility of generating an imaging probe for RTK activation using the SH2 domain of Grb2 for cancer characterization. For cell penetration, molecular analysis and radiolabeling, the SH2 domain was fused with TAT, flag and tyrosine residue, respectively (termed TSF). We analyzed TSF characteristics in cells such as cellular uptake, stability and localization. After uptake into EGFR-expressing cells, TSF was found to be binding to phosphorylated-EGFR, which increased by stimulation with EGF. TSF was co-localized with EGFR in EGFR-activated cells, while it was localized as dots in cytosol in EGFR-non-activated cells. Cellular retention time of TSF was significantly extended under EGFR activation with EGF stimuli and reduced under the treatment with a tyrosine kinase inhibitor, Tyrphostin AG1478. In conclusion, the SH2 domain of Grb2 has the potential to be used as a binding component of a probe to detect activated-RTK and to evaluate the effect of kinase inhibitors on RTK activation.

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A quantitative study of the recruitment potential of all intracellular tyrosine residues on EGFR, FGFR1 and IGF1R

Cited by 52 publications

References 61 publications

Fast rebinding increases dwell time of Src homology 2 (SH2)-containing proteins near the plasma membrane

Fast rebinding increases dwell time of Src homology 2 (SH2)-containing proteins near the plasma membrane

Role of FGFR2-signaling in the pathogenesis of acne

Basic study on SH2 domain of Grb2 as a molecular probe for detection of RTK activation

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