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Anti-Phosphotyrosine, clone 4G10 antibody


Antibody ID


Target Antigen

Phosphotyrosine clone 4G10 c elegans/worm, chemical, hamster, non-human primate, sheep, xenopus/amphibian, chicken/bird, other invertebrate, zebrafish/fish, porcine, amoeba/protozoa, bacteria/archaea, canine, donkey, horse, rabbit, rat, feline, goat, mollusc, plant, drosophila/arthropod, guinea pig, other mammalian, bovine, mouse, reptile, yeast/fungi, human, virus, all

Proper Citation

(Millipore Cat# 05-321, RRID:AB_309678)


monoclonal antibody


seller recommendations: IgG2; IgG2 Immunohistochemistry; Immunoprecipitation; Immunocytochemistry; Western Blot; IC, IH, IP, WB

Host Organism




Angiogenin/Ribonuclease 5 Is an EGFR Ligand and a Serum Biomarker for Erlotinib Sensitivity in Pancreatic Cancer.

  • Wang YN
  • Cancer Cell
  • 2018 Apr 9

Literature context:


Pancreatic ribonuclease (RNase) is a secreted enzyme critical for host defense. We discover an intrinsic RNase function, serving as a ligand for epidermal growth factor receptor (EGFR), a member of receptor tyrosine kinase (RTK), in pancreatic ductal adenocarcinoma (PDAC). The closely related bovine RNase A and human RNase 5 (angiogenin [ANG]) can trigger oncogenic transformation independently of their catalytic activities via direct association with EGFR. Notably, high plasma ANG level in PDAC patients is positively associated with response to EGFR inhibitor erlotinib treatment. These results identify a role of ANG as a serum biomarker that may be used to stratify patients for EGFR-targeted therapies, and offer insights into the ligand-receptor relationship between RNase and RTK families.

Funding information:
  • NCI NIH HHS - P30 CA016672()
  • NCI NIH HHS - R01 CA211615()
  • NCI NIH HHS - T32 CA186892()
  • NCI NIH HHS - U01 CA201777()
  • NIGMS NIH HHS - R01 GM098294(United States)

Mechanism of Allosteric Coupling into and through the Plasma Membrane by EGFR.

  • Sinclair JKL
  • Cell Chem Biol
  • 2018 Apr 21

Literature context:


Epidermal growth factor receptor (EGFR) interacts through its extracellular domain with seven different growth factors. These factors induce different structures within the cytoplasmic juxtamembrane (JM) segment of the dimeric receptor and propagate different growth factor-dependent signals to the cell interior. How this process occurs is unknown. Here we apply diverse experimental and computational tools to show that growth factor identity is encoded by the EGFR transmembrane (TM) helix into discrete helix dimer populations that differ in both cross-location and cross-angle. Helix dimers with smaller cross-angles at multiple cross locations are decoded to induce an EGF-type coiled coil in the adjacent JM, whereas helix dimers with larger cross-angles at fewer cross locations induce the TGF-α-type coiled coil. We propose an updated model for how conformational coupling across multiple EGFR domains results in growth factor-specific information transfer, and demonstrate that this model applies to both EGFR and the related receptor ErbB2.

Funding information:
  • NIAID NIH HHS - K22AI071011(United States)
  • NIGMS NIH HHS - R01 GM083257()
  • NIGMS NIH HHS - T32 GM008283()

Subdomain 2, Not the Transmembrane Domain, Determines the Dimerization Partner of Growth Hormone Receptor and Prolactin Receptor.

  • Liu Y
  • Endocrinology
  • 2017 Oct 1

Literature context:


Growth hormone receptor (GHR) and prolactin (PRL) receptor (PRLR) are homologous transmembrane class I cytokine receptors. In humans, GH interacts with GHR homodimers or PRLR homodimers and PRL interacts with only PRLR homodimers to promote signaling. In human breast cancer cells endogenously expressing both receptors, GHR and PRLR specifically coimmunoprecipitate. We previously devised a split luciferase complementation assay to study GHR and PRLR assemblages. In this technique, firefly luciferase is split into two fragments (N- and C-terminal fragments of the luciferase), each without enzyme activity and tethered to the tails of two receptors. The fragments restore luciferase activity when brought close to each other by the receptors. Real-time ligand-induced complementation changes reflect the arrangement of receptors and indicate that GHR/PRLR is arranged as a heteromultimer comprised of GHR-GHR homodimers and PRLR-PRLR homodimers. We now dissect determinants for GHR and PRLR homodimerization versus heteroassociation. GHR and PRLR have extracellular domains comprised of the ligand-binding N-terminal subdomain 1 and a membrane-proximal subdomain 2 (S2), which fosters receptor-receptor contact. Based on previous studies of S2 versus the transmembrane domain (TMD) in GHR dimerization, we constructed GHR(PRLRS2), GHR(PRLRS2-TMD), and GHR(PRLRTMD), replacing GHR's S2 alone, S2 plus TMD, and TMD alone with PRLR's counterpart. We tested by complementation the ability of these chimeras and GHR or PRLR to homodimerize or heteroassociate. Comparing various combinations, we found GHR(PRLRS2) and GHR(PRLRS2-TMD) behaved as PRLR, whereas GHR(PRLRTMD) behaved as GHR regarding their dimerization partners. We conclude that S2 of GHR and PRLR, rather than their TMDs, determines their dimerization partner.

The Plasticity of the Hsp90 Co-chaperone System.

  • Sahasrabudhe P
  • Mol. Cell
  • 2017 Sep 21

Literature context:


The Hsp90 system in the eukaryotic cytosol is characterized by a cohort of co-chaperones that bind to Hsp90 and affect its function. Although progress has been made regarding the underlying biochemical mechanisms, how co-chaperones influence Hsp90 client proteins in vivo has remained elusive. By investigating the effect of 12 Hsp90 co-chaperones on the activity of different client proteins in yeast, we find that deletion of co-chaperones can have a neutral or negative effect on client activity but can also lead to more active clients. Only a few co-chaperones are active on all clients studied. Closely related clients and even point mutants can depend on different co-chaperones. These effects are direct because differences in client-co-chaperone interactions can be reconstituted in vitro. Interestingly, some co-chaperones affect client conformation in vivo. Thus, co-chaperones adapt the Hsp90 cycle to the requirements of the client proteins, ensuring optimal activation.

A Phosphosite within the SH2 Domain of Lck Regulates Its Activation by CD45.

  • Courtney AH
  • Mol. Cell
  • 2017 Aug 3

Literature context:


The Src Family kinase Lck sets a critical threshold for T cell activation because it phosphorylates the TCR complex and the Zap70 kinase. How a T cell controls the abundance of active Lck molecules remains poorly understood. We have identified an unappreciated role for a phosphosite, Y192, within the Lck SH2 domain that profoundly affects the amount of active Lck in cells. Notably, mutation of Y192 blocks critical TCR-proximal signaling events and impairs thymocyte development in retrogenic mice. We determined that these defects are caused by hyperphosphorylation of the inhibitory C-terminal tail of Lck. Our findings reveal that modification of Y192 inhibits the ability of CD45 to associate with Lck in cells and dephosphorylate the C-terminal tail of Lck, which prevents its adoption of an active open conformation. These results suggest a negative feedback loop that responds to signaling events that tune active Lck amounts and TCR sensitivity.

Defining Human Tyrosine Kinase Phosphorylation Networks Using Yeast as an In Vivo Model Substrate.

  • Corwin T
  • Cell Syst
  • 2017 Aug 23

Literature context:


Systematic assessment of tyrosine kinase-substrate relationships is fundamental to a better understanding of cellular signaling and its profound alterations in human diseases such as cancer. In human cells, such assessments are confounded by complex signaling networks, feedback loops, conditional activity, and intra-kinase redundancy. Here we address this challenge by exploiting the yeast proteome as an in vivo model substrate. We individually expressed 16 human non-receptor tyrosine kinases (NRTKs) in Saccharomyces cerevisiae and identified 3,279 kinase-substrate relationships involving 1,351 yeast phosphotyrosine (pY) sites. Based on the yeast data without prior information, we generated a set of linear kinase motifs and assigned ∼1,300 known human pY sites to specific NRTKs. Furthermore, experimentally defined pY sites for each individual kinase were shown to cluster within the yeast interactome network irrespective of linear motif information. We therefore applied a network inference approach to predict kinase-substrate relationships for more than 3,500 human proteins, providing a resource to advance our understanding of kinase biology.

Funding information:
  • Intramural NIH HHS - P 22043(United States)

A Novel Small Molecule GDNF Receptor RET Agonist, BT13, Promotes Neurite Growth from Sensory Neurons in Vitro and Attenuates Experimental Neuropathy in the Rat.

  • Sidorova YA
  • Front Pharmacol
  • 2017 Jul 6

Literature context:


Neuropathic pain caused by nerve damage is a common and severe class of chronic pain. Disease-modifying clinical therapies are needed as current treatments typically provide only symptomatic relief; show varying clinical efficacy; and most have significant adverse effects. One approach is targeting either neurotrophic factors or their receptors that normalize sensory neuron function and stimulate regeneration after nerve damage. Two candidate targets are glial cell line-derived neurotrophic factor (GDNF) and artemin (ARTN), as these GDNF family ligands (GFLs) show efficacy in animal models of neuropathic pain (Boucher et al., 2000; Gardell et al., 2003; Wang et al., 2008, 2014). As these protein ligands have poor drug-like properties and are expensive to produce for clinical use, we screened 18,400 drug-like compounds to develop small molecules that act similarly to GFLs (GDNF mimetics). This screening identified BT13 as a compound that selectively targeted GFL receptor RET to activate downstream signaling cascades. BT13 was similar to NGF and ARTN in selectively promoting neurite outgrowth from the peptidergic class of adult sensory neurons in culture, but was opposite to ARTN in causing neurite elongation without affecting initiation. When administered after spinal nerve ligation in a rat model of neuropathic pain, 20 and 25 mg/kg of BT13 decreased mechanical hypersensitivity and normalized expression of sensory neuron markers in dorsal root ganglia. In control rats, BT13 had no effect on baseline mechanical or thermal sensitivity, motor coordination, or weight gain. Thus, small molecule BT13 selectively activates RET and offers opportunities for developing novel disease-modifying medications to treat neuropathic pain.

Amplification of F-Actin Disassembly and Cellular Repulsion by Growth Factor Signaling.

  • Yoon J
  • Dev. Cell
  • 2017 Jul 24

Literature context:


Extracellular cues that regulate cellular shape, motility, and navigation are generally classified as growth promoting (i.e., growth factors/chemoattractants and attractive guidance cues) or growth preventing (i.e., repellents and inhibitors). Yet, these designations are often based on complex assays and undefined signaling pathways and thus may misrepresent direct roles of specific cues. Here, we find that a recognized growth-promoting signaling pathway amplifies the F-actin disassembly and repulsive effects of a growth-preventing pathway. Focusing on Semaphorin/Plexin repulsion, we identified an interaction between the F-actin-disassembly enzyme Mical and the Abl tyrosine kinase. Biochemical assays revealed Abl phosphorylates Mical to directly amplify Mical Redox-mediated F-actin disassembly. Genetic assays revealed that Abl allows growth factors and Semaphorin/Plexin repellents to combinatorially increase Mical-mediated F-actin disassembly, cellular remodeling, and repulsive axon guidance. Similar roles for Mical in growth factor/Abl-related cancer cell behaviors further revealed contexts in which characterized positive effectors of growth/guidance stimulate such negative cellular effects as F-actin disassembly/repulsion.

Funding information:
  • NIMH NIH HHS - R01 MH085923()

A Tunable Brake for HECT Ubiquitin Ligases.

  • Chen Z
  • Mol. Cell
  • 2017 May 4

Literature context:


The HECT E3 ligases ubiquitinate numerous transcription factors and signaling molecules, and their activity must be tightly controlled to prevent cancer, immune disorders, and other diseases. In this study, we have found unexpectedly that peptide linkers tethering WW domains in several HECT family members are key regulatory elements of their catalytic activities. Biochemical, structural, and cellular analyses have revealed that the linkers can lock the HECT domain in an inactive conformation and block the proposed allosteric ubiquitin binding site. Such linker-mediated autoinhibition of the HECT domain can be relieved by linker post-translational modifications, but complete removal of the brake can induce hyperactive autoubiquitination and E3 self destruction. These results clarify the mechanisms of several HECT protein cancer associated mutations and provide a new framework for understanding how HECT ubiquitin ligases must be finely tuned to ensure normal cellular behavior.

Funding information:
  • NCI NIH HHS - P50 CA062924()
  • NCI NIH HHS - R01 CA074305()
  • NCI NIH HHS - R37 CA043460()
  • NIGMS NIH HHS - F32 GM120855()
  • NIGMS NIH HHS - R01 GM034933()
  • NIGMS NIH HHS - R01 GM109102()
  • NIGMS NIH HHS - R35 GM118177()

Phosphorylation of β-Tubulin by the Down Syndrome Kinase, Minibrain/DYRK1a, Regulates Microtubule Dynamics and Dendrite Morphogenesis.

  • Ori-McKenney KM
  • Neuron
  • 2016 May 4

Literature context:


Dendritic arborization patterns are consistent anatomical correlates of genetic disorders such as Down syndrome (DS) and autism spectrum disorders (ASDs). In a screen for abnormal dendrite development, we identified Minibrain (MNB)/DYRK1a, a kinase implicated in DS and ASDs, as a regulator of the microtubule cytoskeleton. We show that MNB is necessary to establish the length and cytoskeletal composition of terminal dendrites by controlling microtubule growth. Altering MNB levels disrupts dendrite morphology and perturbs neuronal electrophysiological activity, resulting in larval mechanosensation defects. Using in vivo and in vitro approaches, we uncover a molecular pathway whereby direct phosphorylation of β-tubulin by MNB inhibits tubulin polymerization, a function that is conserved for mammalian DYRK1a. Our results demonstrate that phosphoregulation of microtubule dynamics by MNB/DYRK1a is critical for dendritic patterning and neuronal function, revealing a previously unidentified mode of posttranslational microtubule regulation in neurons and uncovering a conserved pathway for a DS- and ASD-associated kinase.

Disruption of protein-tyrosine phosphatase 1B expression in the pancreas affects β-cell function.

  • Liu S
  • Endocrinology
  • 2014 Sep 25

Literature context:


Protein-tyrosine phosphatase 1B (PTP1B) is a physiological regulator of glucose homeostasis and energy balance. However, the role of PTP1B in pancreatic endocrine function remains largely unknown. To investigate the metabolic role of pancreatic PTP1B, we generated mice with pancreas PTP1B deletion (panc-PTP1B KO). Mice were fed regular chow or a high-fat diet, and metabolic parameters, insulin secretion and glucose tolerance were determined. On regular chow, panc-PTP1B KO and control mice exhibited comparable glucose tolerance whereas aged panc-PTP1B KO exhibited mild glucose intolerance. Furthermore, high-fat feeding promoted earlier impairment of glucose tolerance and attenuated glucose-stimulated insulin secretion in panc-PTP1B KO mice. The secretory defect in glucose-stimulated insulin secretion was recapitulated in primary islets ex vivo, suggesting that the effects were likely cell-autonomous. At the molecular level, PTP1B deficiency in vivo enhanced basal and glucose-stimulated tyrosyl phosphorylation of EphA5 in islets. Consistently, PTP1B overexpression in the glucose-responsive MIN6 β-cell line attenuated EphA5 tyrosyl phosphorylation, and substrate trapping identified EphA5 as a PTP1B substrate. In summary, these studies identify a novel role for PTP1B in pancreatic endocrine function.

Funding information:
  • NINDS NIH HHS - R01 NS084959(United States)

Effects of the antitumor drug OSI-906, a dual inhibitor of IGF-1 receptor and insulin receptor, on the glycemic control, β-cell functions, and β-cell proliferation in male mice.

  • Shirakawa J
  • Endocrinology
  • 2014 Jun 19

Literature context:


The IGF-1 receptor has become a therapeutic target for the treatment of cancer. The efficacy of OSI-906 (linstinib), a dual inhibitor of IGF-1 receptor and insulin receptor, for solid cancers has been examined in clinical trials. The effects of OSI-906, however, on the blood glucose levels and pancreatic β-cell functions have not yet been reported. We investigated the impact of OSI-906 on glycemic control, insulin secretion, β-cell mass, and β-cell proliferation in male mice. Oral administration of OSI-906 worsened glucose tolerance in a dose-dependent manner in the wild-type mice. OSI-906 at a dose equivalent to the clinical daily dose (7.5 mg/kg) transiently evoked glucose intolerance and hyperinsulinemia. Insulin receptor substrate (IRS)-2-deficient mice and mice with diet-induced obesity, both models of peripheral insulin resistance, exhibited more severe glucose intolerance after OSI-906 administration than glucokinase-haploinsufficient mice, a model of impaired insulin secretion. Phloridzin improved the hyperglycemia induced by OSI-906 in mice. In vitro, OSI-906 showed no effect on insulin secretion from isolated islets. After daily administration of OSI-906 for a week to mice, the β-cell mass and β-cell proliferation rate were significantly increased. The insulin signals in the β-cells were apparently unaffected in those mice. Taken together, the results suggest that OSI-906 could exacerbate diabetes, especially in patients with insulin resistance. On the other hand, the results suggest that the β-cell mass may expand in response to chemotherapy with this drug.

Funding information:
  • NIEHS NIH HHS - R01ES015145(United States)
  • NINDS NIH HHS - NS072202(United States)

Metformin inhibits androgen-induced IGF-IR up-regulation in prostate cancer cells by disrupting membrane-initiated androgen signaling.

  • Malaguarnera R
  • Endocrinology
  • 2014 Apr 24

Literature context:


We have previously demonstrated that, in prostate cancer cells, androgens up-regulate IGF-I receptor (IGF-IR) by inducing cAMP-response element-binding protein (CREB) activation and CREB-dependent IGF-IR gene transcription through androgen receptor (AR)-dependent membrane-initiated effects. This IGF-IR up-regulation is not blocked by classical antiandrogens and sensitizes cells to IGF-I-induced biological effects. Metformin exerts complex antitumoral functions in various models and may inhibit CREB activation in hepatocytes. We, therefore, evaluated whether metformin may affect androgen-dependent IGF-IR up-regulation. In the AR(+) LNCaP prostate cancer cells, we found that metformin inhibits androgen-induced CRE activity and IGF-IR gene transcription. CRE activity requires the formation of a CREB-CREB binding protein-CREB regulated transcription coactivator 2 (CRTC2) complex, which follows Ser133-CREB phosphorylation. Metformin inhibited Ser133-CREB phosphorylation and induced nuclear exclusion of CREB cofactor CRTC2, thus dissociating the CREB-CREB binding protein-CRTC2 complex and blocking its transcriptional activity. Similarly to metformin action, CRTC2 silencing inhibited IGF-IR promoter activity. Moreover, metformin blocked membrane-initiated signals of AR to the mammalian target of rapamycin/p70S6Kinase pathway by inhibiting AR phosphorylation and its association with c-Src. AMPK signals were also involved to some extent. By inhibiting androgen-dependent IGF-IR up-regulation, metformin reduced IGF-I-mediated proliferation of LNCaP cells. These results indicate that, in prostate cancer cells, metformin inhibits IGF-I-mediated biological effects by disrupting membrane-initiated AR action responsible for IGF-IR up-regulation and suggest that metformin could represent a useful adjunct to the classical antiandrogen therapy.

Funding information:
  • Intramural NIH HHS - ZIA ES102805(United States)

Evidence of cell-nonautonomous changes in dendrite and dendritic spine morphology in the met-signaling-deficient mouse forebrain.

  • Judson MC
  • J. Comp. Neurol.
  • 2010 Nov 1

Literature context:


Human genetic findings and murine neuroanatomical expression mapping have intersected to implicate Met receptor tyrosine kinase signaling in the development of forebrain circuits controlling social and emotional behaviors that are atypical in autism-spectrum disorders (ASD). To clarify roles for Met signaling during forebrain circuit development in vivo, we generated mutant mice (Emx1(Cre)/Met(fx/fx)) with an Emx1-Cre-driven deletion of signaling-competent Met in dorsal pallially derived forebrain neurons. Morphometric analyses of Lucifer yellow-injected pyramidal neurons in postnatal day 40 anterior cingulate cortex (ACC) revealed no statistically significant changes in total dendritic length but a selective reduction in apical arbor length distal to the soma in Emx1(Cre)/Met(fx/fx) neurons relative to wild type, consistent with a decrease in the total tissue volume sampled by individual arbors in the cortex. The effects on dendritic structure appear to be circuit-selective, insofar as basal arbor length was increased in Emx1(Cre)/Met(fx/fx) layer 2/3 neurons. Spine number was not altered on the Emx1(Cre)/Met(fx/fx) pyramidal cell populations studied, but spine head volume was significantly increased (∼20%). Cell-nonautonomous, circuit-level influences of Met signaling on dendritic development were confirmed by studies of medium spiny neurons (MSN), which do not express Met but receive Met-expressing corticostriatal afferents during development. Emx1(Cre)/Met(fx/fx) MSN exhibited robust increases in total arbor length (∼20%). As in the neocortex, average spine head volume was also increased (∼12%). These data demonstrate that a developmental loss of presynaptic Met receptor signaling can affect postsynaptic morphogenesis and suggest a mechanism whereby attenuated Met signaling could disrupt both local and long-range connectivity within circuits relevant to ASD.

Funding information:
  • NCRR NIH HHS - U54 RR022220(United States)