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Mark Henkemeyer - One of the best experts on this subject based on the ideXlab platform.
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amygdala ephb2 signaling regulates glutamatergic neuron maturation and innate fear
The Journal of Neuroscience, 2016Co-Authors: Xiao Na Zhu, Mark Henkemeyer, Xian Dong Liu, Hanyi Zhuang, Jingyu YangAbstract:The amygdala serves as emotional center to mediate innate fear behaviors that are reflected through neuronal responses to environmental aversive cues. However, the molecular mechanism underlying the initial neuron responses is poorly understood. In this study, we monitored the innate defensive responses to aversive stimuli of either elevated plus maze or predator odor in juvenile mice and found that glutamatergic neurons were activated in amygdala. Loss of EphB2, a receptor tyrosine kinase expressed in amygdala neurons, suppressed the reactions and led to defects in spine morphogenesis and fear behaviors. We further found a coupling of spinogenesis with these threat cues induced neuron activation in developing amygdala that was controlled by EphB2. A constitutively active form of EphB2 was sufficient to rescue the behavioral and morphological defects caused by ablation of Ephrin-B3, a brain-enriched ligand to EphB2. These data suggest that kinase-dependent EphB2 intracellular signaling plays a major role for innate fear responses during the critical developing period, in which spinogenesis in amygdala glutamatergic neurons was involved. SIGNIFICANCE STATEMENT Generation of innate fear responses to threat as an evolutionally conserved brain feature relies on development of functional neural circuit in amygdala, but the molecular mechanism remains largely unknown. We here identify that EphB2 receptor tyrosine kinase, which is specifically expressed in glutamatergic neurons, is required for the innate fear responses in the neonatal brain. We further reveal that EphB2 mediates coordination of spinogenesis and neuron activation in amygdala during the critical period for the innate fear. EphB2 catalytic activity plays a major role for the behavior upon EphB–Ephrin-B3 binding and transnucleus neuronal connections. Our work thus indicates an essential synaptic molecular signaling within amygdala that controls synapse development and helps bring about innate fear emotions in the postnatal developing brain.
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a novel feedback mechanism by Ephrin b1 b2 in t cell activation involves a concentration dependent switch from costimulation to inhibition
European Journal of Immunology, 2012Co-Authors: Hiroki Kawano, Mark Henkemeyer, Yoshio Katayama, Kentaro Minagawa, Manabu Shimoyama, Toshimitsu MatsuiAbstract:Bidirectional signals via Eph receptors/Ephrins have been recognized as major forms of contact-dependent cell communications such as cell attraction and repulsion. T cells express EphBs, and their ligands, the Ephrin-Bs, have been known as costimulatory molecules for T-cell proliferation. Recently, another remarkable feature of Ephrin-As has emerged in the form of a concentration-dependent transition from promotion to inhibition in axon growth. Here we examined whether this modification plays a role in Ephrin-B costimulation in murine primary T cells. Low doses of Ephrin-B1 and Ephrin-B2 costimulated T-cell proliferation induced by anti-CD3, but high concentrations strongly inhibited it. In contrast, Ephrin-B3 showed a steadily increasing stimulatory effect. This modulation was virtually preserved in T cells from mice simultaneously lacking four genes, EphB1, EphB2, EphB3, and EphB6. High concentrations of Ephrin-B1/B2, but not Ephrin-B3, inhibited the anti-CD3-induced phosphorylation of Lck and its downstream signals such as Erk and Akt. Additionally, high doses of any Ephrin-Bs could phosphorylate EphB4. However, only Ephrin-B1/B2 but not Ephrin-B3 recruited SHP1, a phosphatase to suppress the phosphorylation of Lck. These data suggest that EphB4 signaling could engage in negative feedback to TCR signals. T-cell activation may be finely adjusted by the combination and concentration of Ephrin-Bs expressed in the immunological microenvironment.
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a dual shaping mechanism for postsynaptic Ephrin B3 as a receptor that sculpts dendrites and synapses
Nature Neuroscience, 2011Co-Authors: Nanjie Xu, Jay R Gibson, Mark HenkemeyerAbstract:Examining the role of Ephrin-B3 in dendritic and synaptic development in vivo, this study finds that Ephrin-B3 functions postsynaptically as a receptor to initiate reverse signaling events that organize dendritic branching complexity, spine maturation and formation of functional synapses.
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a dual shaping mechanism for postsynaptic Ephrin B3 as a receptor that sculpts dendrites and synapses
Nature Neuroscience, 2011Co-Authors: Suya Sun, Jay R Gibson, Mark HenkemeyerAbstract:As the neural network becomes wired, postsynaptic signaling molecules are thought to control the growth of dendrites and synapses. However, how these molecules are coordinated to sculpt postsynaptic structures is less well understood. We find that Ephrin-B3, a transmembrane ligand for Eph receptors, functions postsynaptically as a receptor to transduce reverse signals into developing dendrites of mouse hippocampal neurons. Both tyrosine phosphorylation-dependent GRB4 SH2/SH3 adaptor-mediated signals and PSD-95-discs large-zona occludens-1 (PDZ) domain-dependent signals are required for inhibition of dendrite branching, whereas only PDZ interactions are necessary for spine formation and excitatory synaptic function. PICK1 and syntenin, two PDZ domain proteins, participate with Ephrin-B3 in these postsynaptic activities. PICK1 has a specific role in spine and synapse formation, and syntenin promotes both dendrite pruning and synapse formation to build postsynaptic structures that are essential for neural circuits. The study thus dissects Ephrin-B reverse signaling into three distinct intracellular pathways and protein-protein interactions that mediate the maturation of postsynaptic neurons.
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a novel feedback mechanism by Ephrin b1 b2 in t cell activation concentration dependent switch from costimulation to inhibition
Blood, 2010Co-Authors: Hiroki Kawano, Mark Henkemeyer, Yoshio Katayama, Kentaro Minagawa, Manabu Shimoyama, Toshimitsu MatsuiAbstract:Abstract Abstract 277 Eph is the largest known family of receptor tyrosine kinases, and bind to a cell surface-associated ligand, Ephrin on neighboring cells upon direct cell-cell contact. The ensuing bidirectional signals have been recognized as a major form of contact-dependent cell communications, such as cell attraction and repulsion to control accurate spatial and temporal patterning in the development of the central nervous system. EphBs, EphB6 in particular, are expressed in T cells and its specific ligand, Ephrin-B2 has been shown to act as a costimulatory molecule for the T cell receptor (TCR)-mediated cell proliferation. Recently, another remarkable feature of Ephrins, a concentration-dependent transition from promotion to inhibition in axon growth has emerged in Ephrin-As. Thus, we postulated that this type of ligand concentration dependent functional transition would be suitable for the delicate tuning of immune responses to avoid reckless drive. To figure this out, we carefully evaluated the costimulatory effects of Ephrin-Bs by using murine primary T cells. Interestingly, low doses of solid phase Ephrin-B1 as well as Ephrin-B2 (at up to 5μ g/ml) costimulated, to the comparable level with anti-CD28, T cell proliferation induced by suboptimal concentration of immobilized anti-CD3 antibody, but high concentrations of Ephrin-B1/B2 inhibited the TCR-mediated proliferation significantly (by approximately 70% reduction from the baseline at 20μ g/ml). The similar concentration-dependent transition from coactivation to inhibition was also observed under the optimal CD3 stimulation. The concentration-dependent biphasic effects, positively at low concentration and negatively at high concentration, by Ephrin-B1/B2 in T cell activation were confirmed in the cytokine production such as TNF-α, IL-2, and IFN-γ. In contrast, Ephrin-B3 showed steadily increasing stimulatory effect even in higher concentrations in proliferation and cytokine production. We speculated that these unique modulations were partly mediated by EphB6 because EphB6 transfected in HEK293T cells has been shown to exert biphasic effects in cell adhesion and migration in response to different concentrations of Ephrin-B2. T cell derived from Ephb6 -/- mice showed decreased CD3-stimulated cell proliferation as reported previously. However, the unique comodulatory pattern by each Ephrin-B was virtually preserved in Ephb6 -/- T cells. Since the functions of Eph family could be redundant, we further investigated by generating multiple EphB knockout mice lacking four genes, Ephb1, Ephb2, EphB3 and Ephb6. Surprisingly, no further alteration was observed in T cells from the quadruple knockout mice compared to the Ephb6 single deficiency. We also confirmed that EphA4, an exception in EphA receptor family which binds Ephrin-Bs, was not expressed in T cells by RT-PCR. Taken together with the fact that EphB5 does not exist in mammals, the unique comodification by Ephrin-Bs might be regulated by EphB4. Next, we examined the cross-talk of EphB forward signaling with TCR pathway. The inhibitor of p38MAPK and p44/42MAPK significantly reduced the TCR-mediated proliferation, but did conserve the concentration-dependent effects of Ephrin-B1/B2, suggesting the interference with EphB signaling in TCR signal transduction at the upstream of MAPKs which are important for cell growth and survival. Immuno-blot analyses revealed that high concentrations of Ephrin-B1/B2, but not Ephrin-B3, clearly inhibited the anti-CD3 induced phosphorylation of Lck and its downstream signaling molecules such as ZAP70, c-Raf, MEK1/2, Erk, and Akt, although the phosphorylation of CD3ζ was not inhibited by high concentrations of any Ephrin-Bs. These data suggest that Eph signaling upon stimulation by high concentrations of Ephrin-B1/B2 may engage in negative feedback to TCR signals via Lck. The present studies demonstrate that TCR-mediated primary T cell activation may be highly governed by EphB/Ephrin-B axis with a complexity determined by the combination as well as the concentration of different Ephrin-Bs expressed in immunological microenvironments. EphB-involved in negative feedback of T cell activation could be a novel therapeutic target to inhibit the most proximal TCR signaling molecule, Lck. The generation of strong signaling molecule which mimics Ephrin-B1/B2 would be an effective strategy to control T cell mediated immune disorders. Disclosures: No relevant conflicts of interest to declare.
Matthew B Dalva - One of the best experts on this subject based on the ideXlab platform.
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Ephrin B3 regulates glutamate receptor signaling at hippocampal synapses
Molecular and Cellular Neuroscience, 2010Co-Authors: Marcia D Antion, Matthew B Dalva, Louisa A Christie, Allison M Bond, Anis ContractorAbstract:B-Ephrin-EphB receptor signaling modulates NMDA receptors by inducing tyrosine phosphorylation of NR2 subunits. Ephrins and EphB RTKs are localized to postsynaptic compartments in the CA1, and therefore potentially interact in a non-canonical cis- configuration. However, it is not known whether cis- configured receptor-ligand signaling is utilized by this class of RTKs, and whether this might influence excitatory synapses. We found that ablation of Ephrin-B3 results in an enhancement of the NMDA receptor component of synaptic transmission relative to the AMPA receptor component in CA1 synapses. Synaptic AMPA receptor expression is reduced in Ephrin-B3 knockout mice, and there is a marked enhancement of tyrosine phosphorylation of the NR2B receptor subunit. In a reduced system co-expression of Ephrin-B3 attenuated EphB2-mediated NR2B tyrosine phosphorylation. Moreover, phosphorylation of EphB2 was elevated in the hippocampus of Ephrin-B3 knockout mice, suggesting that regulation of EphB2 activity is lost in these mice. Direct activation of EphB RTKs resulted in phosphorylation of NR2B and a potential signaling partner, the non-receptor tyrosine kinase Pyk2. Our data suggests that Ephrin-B3 limits EphB RTK-mediated phosphorylation of the NR2B subunit through an inhibitory cis- interaction which is required for the correct function of glutamatergic CA1 synapses.
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trans synaptic ephb2 Ephrin B3 interaction regulates excitatory synapse density by inhibition of postsynaptic mapk signaling
Proceedings of the National Academy of Sciences of the United States of America, 2010Co-Authors: Andrew C Mcclelland, Martin Hruska, Andrew J Coenen, Mark Henkemeyer, Matthew B DalvaAbstract:Nervous system function requires tight control over the number of synapses individual neurons receive, but the underlying cellular and molecular mechanisms that regulate synapse number remain obscure. Here we present evidence that a trans-synaptic interaction between EphB2 in the presynaptic compartment and Ephrin-B3 in the postsynaptic compartment regulates synapse density and the formation of dendritic spines. Observations in cultured cortical neurons demonstrate that synapse density scales with Ephrin-B3 expression level and is controlled by Ephrin-B3–dependent competitive cell–cell interactions. RNA interference and biochemical experiments support the model that Ephrin-B3 regulates synapse density by directly binding to Erk1/2 to inhibit postsynaptic Ras/mitogen-activated protein kinase signaling. Together these findings define a mechanism that contributes to synapse maturation and controls the number of excitatory synaptic inputs received by individual neurons.
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Ephrin-B1 and Ephrin-B2 mediate EphB-dependent presynaptic development via syntenin-1
Proceedings of the National Academy of Sciences of the United States of America, 2009Co-Authors: Andrew C Mcclelland, Sean I. Sheffler-collins, Matthew S. Kayser, Matthew B DalvaAbstract:The development of central nervous system synapses requires precise coordination between presynaptic and postsynaptic components. The EphB family controls postsynaptic development by interacting with glutamate receptors and regulating dendritic filopodia motility, but how EphBs induce the formation of presynaptic specializations is less well understood. Here, we show that knockdown of presynaptic Ephrin-B1, Ephrin-B2, or syntenin-1, but not Ephrin-B3, prevents EphB-dependent presynaptic development. Ephrin-B1, Ephrin-B2, and syntenin-1 are clustered together with presynaptic markers, suggesting that these molecules function jointly in presynaptic development. Knockdown of Ephrin-B1 or Ephrin-B2 reduces the number of synaptic specializations and the colocalization of syntenin-1 with synaptic markers. Simultaneous knockdown of Ephrin-B1 and Ephrin-B2 suggests that they function independently in the formation of synaptic contacts, but act together to recruit syntenin-1 to presynaptic terminals. Taken together, these results demonstrate that Ephrin-B1 and Ephrin-B2 function with EphB to mediate presynaptic development via syntenin-1.
Toshimitsu Matsui - One of the best experts on this subject based on the ideXlab platform.
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a novel feedback mechanism by Ephrin b1 b2 in t cell activation involves a concentration dependent switch from costimulation to inhibition
European Journal of Immunology, 2012Co-Authors: Hiroki Kawano, Mark Henkemeyer, Yoshio Katayama, Kentaro Minagawa, Manabu Shimoyama, Toshimitsu MatsuiAbstract:Bidirectional signals via Eph receptors/Ephrins have been recognized as major forms of contact-dependent cell communications such as cell attraction and repulsion. T cells express EphBs, and their ligands, the Ephrin-Bs, have been known as costimulatory molecules for T-cell proliferation. Recently, another remarkable feature of Ephrin-As has emerged in the form of a concentration-dependent transition from promotion to inhibition in axon growth. Here we examined whether this modification plays a role in Ephrin-B costimulation in murine primary T cells. Low doses of Ephrin-B1 and Ephrin-B2 costimulated T-cell proliferation induced by anti-CD3, but high concentrations strongly inhibited it. In contrast, Ephrin-B3 showed a steadily increasing stimulatory effect. This modulation was virtually preserved in T cells from mice simultaneously lacking four genes, EphB1, EphB2, EphB3, and EphB6. High concentrations of Ephrin-B1/B2, but not Ephrin-B3, inhibited the anti-CD3-induced phosphorylation of Lck and its downstream signals such as Erk and Akt. Additionally, high doses of any Ephrin-Bs could phosphorylate EphB4. However, only Ephrin-B1/B2 but not Ephrin-B3 recruited SHP1, a phosphatase to suppress the phosphorylation of Lck. These data suggest that EphB4 signaling could engage in negative feedback to TCR signals. T-cell activation may be finely adjusted by the combination and concentration of Ephrin-Bs expressed in the immunological microenvironment.
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a novel feedback mechanism by Ephrin b1 b2 in t cell activation concentration dependent switch from costimulation to inhibition
Blood, 2010Co-Authors: Hiroki Kawano, Mark Henkemeyer, Yoshio Katayama, Kentaro Minagawa, Manabu Shimoyama, Toshimitsu MatsuiAbstract:Abstract Abstract 277 Eph is the largest known family of receptor tyrosine kinases, and bind to a cell surface-associated ligand, Ephrin on neighboring cells upon direct cell-cell contact. The ensuing bidirectional signals have been recognized as a major form of contact-dependent cell communications, such as cell attraction and repulsion to control accurate spatial and temporal patterning in the development of the central nervous system. EphBs, EphB6 in particular, are expressed in T cells and its specific ligand, Ephrin-B2 has been shown to act as a costimulatory molecule for the T cell receptor (TCR)-mediated cell proliferation. Recently, another remarkable feature of Ephrins, a concentration-dependent transition from promotion to inhibition in axon growth has emerged in Ephrin-As. Thus, we postulated that this type of ligand concentration dependent functional transition would be suitable for the delicate tuning of immune responses to avoid reckless drive. To figure this out, we carefully evaluated the costimulatory effects of Ephrin-Bs by using murine primary T cells. Interestingly, low doses of solid phase Ephrin-B1 as well as Ephrin-B2 (at up to 5μ g/ml) costimulated, to the comparable level with anti-CD28, T cell proliferation induced by suboptimal concentration of immobilized anti-CD3 antibody, but high concentrations of Ephrin-B1/B2 inhibited the TCR-mediated proliferation significantly (by approximately 70% reduction from the baseline at 20μ g/ml). The similar concentration-dependent transition from coactivation to inhibition was also observed under the optimal CD3 stimulation. The concentration-dependent biphasic effects, positively at low concentration and negatively at high concentration, by Ephrin-B1/B2 in T cell activation were confirmed in the cytokine production such as TNF-α, IL-2, and IFN-γ. In contrast, Ephrin-B3 showed steadily increasing stimulatory effect even in higher concentrations in proliferation and cytokine production. We speculated that these unique modulations were partly mediated by EphB6 because EphB6 transfected in HEK293T cells has been shown to exert biphasic effects in cell adhesion and migration in response to different concentrations of Ephrin-B2. T cell derived from Ephb6 -/- mice showed decreased CD3-stimulated cell proliferation as reported previously. However, the unique comodulatory pattern by each Ephrin-B was virtually preserved in Ephb6 -/- T cells. Since the functions of Eph family could be redundant, we further investigated by generating multiple EphB knockout mice lacking four genes, Ephb1, Ephb2, EphB3 and Ephb6. Surprisingly, no further alteration was observed in T cells from the quadruple knockout mice compared to the Ephb6 single deficiency. We also confirmed that EphA4, an exception in EphA receptor family which binds Ephrin-Bs, was not expressed in T cells by RT-PCR. Taken together with the fact that EphB5 does not exist in mammals, the unique comodification by Ephrin-Bs might be regulated by EphB4. Next, we examined the cross-talk of EphB forward signaling with TCR pathway. The inhibitor of p38MAPK and p44/42MAPK significantly reduced the TCR-mediated proliferation, but did conserve the concentration-dependent effects of Ephrin-B1/B2, suggesting the interference with EphB signaling in TCR signal transduction at the upstream of MAPKs which are important for cell growth and survival. Immuno-blot analyses revealed that high concentrations of Ephrin-B1/B2, but not Ephrin-B3, clearly inhibited the anti-CD3 induced phosphorylation of Lck and its downstream signaling molecules such as ZAP70, c-Raf, MEK1/2, Erk, and Akt, although the phosphorylation of CD3ζ was not inhibited by high concentrations of any Ephrin-Bs. These data suggest that Eph signaling upon stimulation by high concentrations of Ephrin-B1/B2 may engage in negative feedback to TCR signals via Lck. The present studies demonstrate that TCR-mediated primary T cell activation may be highly governed by EphB/Ephrin-B axis with a complexity determined by the combination as well as the concentration of different Ephrin-Bs expressed in immunological microenvironments. EphB-involved in negative feedback of T cell activation could be a novel therapeutic target to inhibit the most proximal TCR signaling molecule, Lck. The generation of strong signaling molecule which mimics Ephrin-B1/B2 would be an effective strategy to control T cell mediated immune disorders. Disclosures: No relevant conflicts of interest to declare.
Edward W C Bradley - One of the best experts on this subject based on the ideXlab platform.
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the role of grip1 and Ephrin B3 in blood pressure control and vascular smooth muscle cell contractility
Scientific Reports, 2016Co-Authors: Yujia Wang, Junzheng Peng, Hongyu Luo, John Verner Raelson, Georg Ehret, Patricia B Munroe, Ekatherina Stoyanova, Zhao Qin, Guy Cloutier, Edward W C BradleyAbstract:Several erythropoietin-producing hepatocellular receptor B family (EPHB) and their ligands, EphrinBs (EFNBs), are involved in blood pressure regulation in animal models. We selected 528 single nucleotide polymorphisms (SNPs) within the genes of EPHB6, EFNB2, EFNB3 and GRIP1 in the EPH/EFN signalling system to query the International Blood Pressure Consortium dataset. A SNP within the glutamate receptor interacting protein 1 (GRIP1) gene presented a p-value of 0.000389, approaching the critical p-value of 0.000302, for association with diastolic blood pressure of 60,396 individuals. According to echocardiography, we found that EfnB3 gene knockout mice showed enhanced constriction in the carotid arteries. In vitro studies revealed that in mouse vascular smooth muscle cells, siRNA knockdown of GRIP1, which is in the EFNB3 reverse signalling pathway, resulted in increased contractility of these cells. These data suggest that molecules in the EPHB/EFNB signalling pathways, specifically EFNB3 and GRIP1, are involved blood pressure regulation.
Di Stuart - One of the best experts on this subject based on the ideXlab platform.
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crystal structure and carbohydrate analysis of nipah virus attachment glycoprotein a template for antiviral and vaccine design
Journal of Virology, 2008Co-Authors: Max Crispin, David J. Harvey, Jonathan M. Grimes, E Y Jones, A R Aricescu, Di StuartAbstract:Two members of the paramyxovirus family, Nipah virus (NiV) and Hendra virus (HeV), are recent additions to a growing number of agents of emergent diseases which use bats as a natural host. Identification of Ephrin-B2 and Ephrin-B3 as cellular receptors for these viruses has enabled the development of immunotherapeutic reagents which prevent virus attachment and subsequent fusion. Here we present the structural analysis of the protein and carbohydrate components of the unbound viral attachment glycoprotein of NiV glycoprotein (NiV-G) at a 2.2-A resolution. Comparison with its Ephrin-B2-bound form reveals that conformational changes within the envelope glycoprotein are required to achieve viral attachment. Structural differences are particularly pronounced in the 579-590 loop, a major component of the Ephrin binding surface. In addition, the 236-245 loop is rather disordered in the unbound structure. We extend our structural characterization of NiV-G with mass spectrometric analysis of the carbohydrate moieties. We demonstrate that NiV-G is largely devoid of the oligomannose-type glycans that in viruses such as human immunodeficiency virus type 1 and Ebola virus influence viral tropism and the host immune response. Nevertheless, we find putative ligands for the endothelial cell lectin, LSECtin. Finally, by mapping structural conservation and glycosylation site positions from other members of the paramyxovirus family, we suggest the molecular surface involved in oligomerization. These results suggest possible pathways of virus-host interaction and strategies for the optimization of recombinant vaccines.
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structural basis of nipah and hendra virus attachment to their cell surface receptor Ephrin b2
Nature Structural & Molecular Biology, 2008Co-Authors: Radu A Aricescu, Jonathan M. Grimes, Robert J C Gilbert, Yvonne E Jones, Di StuartAbstract:Nipah and Hendra viruses are emergent paramyxoviruses, causing disease characterized by rapid onset and high mortality rates, resulting in their classification as Biosafety Level 4 pathogens. Their attachment glycoproteins are essential for the recognition of the cell-surface receptors Ephrin-B2 (EFNB2) and Ephrin-B3 (EFNB3). Here we report crystal structures of both Nipah and Hendra attachment glycoproteins in complex with human EFNB2. In contrast to previously solved paramyxovirus attachment complexes, which are mediated by sialic acid interactions, the Nipah and Hendra complexes are maintained by an extensive protein-protein interface, including a crucial phenylalanine side chain on EFNB2 that fits snugly into a hydrophobic pocket on the viral protein. By analogy with the development of antivirals against sialic acid binding viruses, these results provide a structural template to target antiviral inhibition of protein-protein interactions.