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Manabu Negishi - One of the best experts on this subject based on the ideXlab platform.
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Plexin b1 is a gtpase activating protein for m ras remodelling dendrite morphology
EMBO Reports, 2009Co-Authors: Yasuhiro Saito, Izumi Oinuma, Satoshi Fujimoto, Manabu NegishiAbstract:Plexins are receptors for axonal guidance molecules known as semaphorins. We recently reported that the semaphorin 4D (Sema4D) receptor, Plexin-B1, induces axonal growth cone collapse by functioning as an R-Ras GTPase activating protein (GAP). Here, we report that Plexin-B1 shows GAP activity for M-Ras, another member of the Ras family of GTPases. In cortical neurons, the expression of M-Ras was upregulated during dendritic development. Knockdown of endogenous M-Ras—but not R-Ras—reduced dendritic outgrowth and branching, whereas overexpression of constitutively active M-Ras, M-Ras(Q71L), enhanced dendritic outgrowth and branching. Sema4D suppressed M-Ras activity and reduced dendritic outgrowth and branching, but this reduction was blocked by M-Ras(Q71L). M-Ras(Q71L) stimulated extracellular signal-regulated kinase (ERK) activation, inducing dendrite growth, whereas Sema4D suppressed ERK activity and down-regulation of ERK was required for a Sema4D-induced reduction of dendrite growth. Thus, we conclude that Plexin-B1 is a dual functional GAP for R-Ras and M-Ras, remodelling axon and dendrite morphology, respectively.
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Different Requirement for Rnd GTPases of R-Ras GAP Activity of Plexin-C1 and Plexin-D1
The Journal of biological chemistry, 2009Co-Authors: Kanami Uesugi, Hironori Katoh, Izumi Oinuma, Manabu NegishiAbstract:Plexins, comprising Plexin-A, -B, -C, and -D subfamilies, are receptors for semaphorins governing cell adhesion, migration, and axon guidance. Among Plexin subfamilies, Plexin-A1 and Plexin-B1 have been shown to function as an R-Ras GAP, inducing repulsive responses, and the expression of R-Ras GAP activity requires the binding of Rnd1, a member of Rnd subfamily of Rho GTPases. However, signaling pathways of Plexin-D1 and Plexin-C1 still remain obscure. Here, we found that Plexin-D1 displayed R-Ras GAP activity and inhibited migration of COS-7 cells, and these actions required Rnd2, another Rnd subfamily GTPase. Rnd2 bound to Plexin-D1 in cortical neurons, and Sema3E/Plexin-D1-induced inhibition of axon outgrowth of cortical neurons required Rnd2 and down-regulation of R-Ras activity. On the other hand, Plexin-C1 displayed R-Ras GAP activity and inhibited cell migration of COS-7 cells without Rnd proteins. Therefore, R-Ras GAP activity is a common function of Plexin subfamilies but the regulation of R-Ras GAP activity of Plexins by Rnd proteins is different among Plexin subfamilies.
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sema4d Plexin b1 activates gsk 3β through r ras gap activity inducing growth cone collapse
EMBO Reports, 2006Co-Authors: Yuri Ito, Hironori Katoh, Izumi Oinuma, Kozo Kaibuchi, Manabu NegishiAbstract:Plexins are receptors for the axonal guidance molecules known as semaphorins, and the semaphorin 4D (Sema4D) receptor Plexin-B1 induces repulsive responses by functioning as an R-Ras GTPase-activating protein (GAP). Here we characterized the downstream signalling of Plexin-B1-mediated R-Ras GAP activity, inducing growth cone collapse. Sema4D suppressed R-Ras activity in hippocampal neurons, in parallel with dephosphorylation of Akt and activation of glycogen synthase kinase (GSK)-3β. Ectopic expression of the constitutively active mutant of Akt or treatment with GSK-3 inhibitors suppressed the Sema4D-induced growth cone collapse. Constitutive activation of phosphatidylinositol-3-OH kinase (PI(3)K), an upstream kinase of Akt and GSK-3β, also blocked the growth cone collapse. The R-Ras GAP activity was necessary for Plexin-B1-induced dephosphorylation of Akt and activation of GSK-3β and was also required for phosphorylation of a downstream kinase of GSK-3β, collapsin response mediator protein-2. Plexin-A1 also induced dephosphorylation of Akt and GSK-3β through its R-Ras GAP activity. We conclude that Plexin-B1 inactivates PI(3)K and dephosphorylates Akt and GSK-3β through R-Ras GAP activity, inducing growth cone collapse.
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Molecular Dissection of the Semaphorin 4D Receptor Plexin-B1-Stimulated R-Ras GTPase-Activating Protein Activity and Neurite Remodeling in Hippocampal Neurons
The Journal of neuroscience : the official journal of the Society for Neuroscience, 2004Co-Authors: Izumi Oinuma, Hironori Katoh, Manabu NegishiAbstract:Plexins serve as receptors for repulsive axonal guidance molecules semaphorins. The cytoplasmic domain of the semaphorin 4D (Sema4D) receptor, Plexin-B1 has two separated Ras GTPase-activating protein (GAP)-homologous domains, C1 and C2. Recently, we reported that the Rho family small GTPase Rnd1 associates with Plexin-B1, and the Plexin-B1-Rnd1 complex stimulates GTPase activity of R-Ras, inducing growth cone collapse in hippocampal neurons in response to Sema4D. However, the molecular mechanisms by which Plexin-B1 exhibits the GAP activity remain unclear. In this report, critical roles of Rnd1 and Sema4D in Plexin-B1-stimulated R-Ras GAP activity and neurite remodeling were examined. The N-terminal region of the cytoplasmic domain of Plexin-B1 containing the C1 domain interacts with the C-terminal region containing the C2 domain, and Rnd1 disrupts this interaction. On the other hand, Sema4D induces clustering of Rnd1-bound Plexin-B1, in parallel with inactivation of R-Ras in cells. Antibody clustering of the recombinant cytoplasmic domain of Plexin-B1 in the presence of Rnd1 triggers the R-Ras GAP activity. Deletion of the extracellular domain of Plexin-B1 causes ligand-independent clustering of the receptor, rendering the receptor constitutively active in the presence of Rnd1, and induces contraction of COS-7 cells and inhibition of neurite outgrowth in hippocampal neurons. These results indicate that Rnd1 opens the two R-Ras GAP domains of Plexin-B1, and Sema4D-induced receptor clustering stimulates R-Ras GAP activity and neurite remodeling in hippocampal neurons.
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the semaphorin 4d receptor Plexin b1 is a gtpase activating protein for r ras
Science, 2004Co-Authors: Izumi Oinuma, Hironori Katoh, Yukio Ishikawa, Manabu NegishiAbstract:Plexins are cell surface receptors for semaphorin molecules, and their interaction governs cell adhesion and migration in a variety of tissues. We report that the Semaphorin 4D (Sema4D) receptor Plexin-B1 directly stimulates the intrinsic guanosine triphosphatase (GTPase) activity of R-Ras, a member of the Ras superfamily of small GTP-binding proteins that has been implicated in promoting cell adhesion and neurite outgrowth. This activity required the interaction of Plexin-B1 with Rnd1, a small GTP-binding protein of the Rho family. Down-regulation of R-Ras activity by the Plexin-B1-Rnd1 complex was essential for the Sema4D-induced growth cone collapse in hippocampal neurons. Thus, Plexin-B1 mediates Sema4D-induced repulsive axon guidance signaling by acting as a GTPase activating protein for R-Ras.
Luca Tamagnone - One of the best experts on this subject based on the ideXlab platform.
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Plexin-Bs enhance their GAP activity with a novel activation switch loop generating a cooperative enzyme
Cellular and Molecular Life Sciences, 2020Co-Authors: Zhen-lu Li, Luca Tamagnone, Jeannine Müller-greven, Matthias BuckAbstract:Plexins receive guidance cues from semaphorin ligands and transmit their signal through the plasma membrane. This family of proteins is unique amongst single-pass transmembrane receptors as their intracellular regions interact directly with several small GTPases, which regulate cytoskeletal dynamics and cell adhesion. Here, we characterize the GTPase Activating Protein (GAP) function of Plexin-B1 and find that a cooperative GAP activity towards the substrate GTPase, Rap1b, is associated with the N-terminal Juxtamembrane region of Plexin-B1. Importantly, we unveil an activation mechanism of Plexin-B1 by identifying a novel functional loop which partially blocks Rap1b entry into the Plexin GAP domain. Consistent with the concept of allokairy developed for other systems, Plexin-B activity is increased by an apparent substrate-mediated cooperative effect. Simulations and mutagenesis suggest the repositioned JM conformation is stabilized by the new activation switch loop when the active site is occupied, giving rise to faster enzymatic turnover and cooperative behavior. The biological implications, essentially those of a threshold behavior for cell migration, are discussed.
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Plexin bs enhance their gap activity with a novel activation switch loop
bioRxiv, 2020Co-Authors: Jeannine Mullergreven, Soonjeung Kim, Luca Tamagnone, Matthias BuckAbstract:Plexins receive guidance cues from semaphorin ligands and transmit their signal through the plasma membrane. This family of proteins is unique amongst single-pass transmembrane receptors as their intracellular regions interact directly with several small GTPases, which regulate cytoskeletal dynamics and cell adhesion. Here, we characterize the GTPase Activating Protein (GAP) function of Plexin-B1 and find that a cooperative GAP activity towards the substrate GTPase, Rap1b, is associated with the N-terminal Juxtamembrane region of Plexin-B1. Importantly, we unveil an activation mechanism of Plexin-B1 by identifying a novel functional loop which partially blocks Rap1b entry to the Plexin GAP domain. Consistent with the concept of allokairy developed for other systems, Plexin-B activity is increased by an apparent substrate mediated cooperative effect. Simulations and mutagenesis suggest the repositioned JM conformation is stabilized by the new activation switch loop when the active site is occupied, giving rise to faster enzymatic turnover and cooperative behavior.
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Structure and function of the intracellular region of the Plexin-b1 transmembrane receptor.
The Journal of biological chemistry, 2009Co-Authors: Yufeng Tong, Prasanta K. Hota, Limin Shen, Soonjeung Kim, Wolfram Tempel, Junia Y Penachioni, Mehdi Bagheri Hamaneh, Rebecca S. Alviani, Luca TamagnoneAbstract:Members of the Plexin family are unique transmembrane receptors in that they interact directly with Rho family small GTPases; moreover, they contain a GTPase-activating protein (GAP) domain for R-Ras, which is crucial for Plexin-mediated regulation of cell motility. However, the functional role and structural basis of the interactions between the different intracellular domains of Plexins remained unclear. Here we present the 2.4 A crystal structure of the complete intracellular region of human Plexin-B1. The structure is monomeric and reveals that the GAP domain is folded into one structure from two segments, separated by the Rho GTPase binding domain (RBD). The RBD is not dimerized, as observed previously. Instead, binding of a conserved loop region appears to compete with dimerization and anchors the RBD to the GAP domain. Cell-based assays on mutant proteins confirm the functional importance of this coupling loop. Molecular modeling based on structural homology to p120(GAP).H-Ras suggests that Ras GTPases can bind to the Plexin GAP region. Experimentally, we show that the monomeric intracellular Plexin-B1 binds R-Ras but not H-Ras. These findings suggest that the monomeric form of the intracellular region is primed for GAP activity and extend a model for Plexin activation.
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Plexin b2 but not Plexin b1 critically modulates neuronal migration and patterning of the developing nervous system in vivo
The Journal of Neuroscience, 2007Co-Authors: Suhua Deng, Esther T. Stoeckli, Olivier Mauti, Alexandra Hirschberg, Thomas Worzfeld, Junia Y Penachioni, Alexander Korostylev, Jakub M Swiercz, Peter Vodrazka, Luca TamagnoneAbstract:Semaphorins and their receptors, Plexins, have emerged as important cellular cues regulating key developmental processes. B-type Plexins directly regulate the actin cytoskeleton in a variety of cell types. Recently, B-type Plexins have been shown to be expressed in striking patterns in the nervous system over critical developmental windows. However, in contrast to the well characterized Plexin-A family, the functional role of Plexin-B proteins in neural development and organogenesis in vertebrates in vivo is not known. Here, we have elucidated the functional contribution of the two neuronally expressed Plexin-B proteins, Plexin-B1 or Plexin-B2, toward the development of the peripheral nervous system and the CNS by generating and analyzing constitutive knock-out mice. The development of the nervous system was found to be normal in mice lacking Plexin-B1, whereas mice lacking Plexin-B2 demonstrated defects in closure of the neural tube and a conspicuous disorganization of the embryonic brain. After analyzing mutant mice, which bypassed neural tube defects, we observed a key requirement for Plexin-B2 in proliferation and migration of granule cell precursors in the developing dentate gyrus, olfactory bulb, and cerebellum. Furthermore, we identified semaphorin 4C as a high-affinity ligand for Plexin-B2 in binding and functional assays. Semaphorin 4C stimulated activation of ErbB-2 and RhoA via Plexin-B2 and enhanced proliferation and migration of granule cell precursors. Semaphorin 4C-induced proliferation of ventricular zone neuroblasts was abrogated in mice lacking Plexin-B2. These genetic and functional analyses reveal a key requirement for Plexin-B2, but not Plexin-B1, in patterning of the vertebrate nervous system in vivo.
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p190 rho gtpase activating protein associates with Plexins and it is required for semaphorin signalling
Journal of Cell Science, 2005Co-Authors: Davide Barberis, Simona Corso, Paolo M. Comoglio, Stefania Artigiani, Andrea Casazza, Raffaella Sordella, Jeff Settleman, Luca TamagnoneAbstract:Plexins are transmembrane receptors for semaphorins, guiding cell migration and axon extension. Plexin activation leads to the disassembly of integrin-based focal adhesive structures and to actin cytoskeleton remodelling and inhibition of cell migration; however, the underlying molecular mechanisms are unclear. We consistently observe a transient decrease of cellular RhoA-GTP levels upon Plexin activation in adherent cells. One of the main effectors of RhoA downregulation is p190, a ubiquitously expressed GTPase activating protein (GAP). We show that, in p190-deficient fibroblasts, the typical functional activities mediated by Plexins (such as cell collapse and inhibition of integrin-based adhesion) are blocked or greatly impaired. Notably, the functional response can be rescued in these cells by re-expressing exogenous p190, but not a mutant form specifically lacking RhoGAP activity. We furthermore demonstrate that semaphorin function is blocked in epithelial cells, primary endothelial cells and neuroblasts upon treatment with small interfering RNAs that knockdown p190 expression. Finally, we show that p190 transiently associates with Plexins, and its RhoGAP activity is increased in response to semaphorin stimulation. We conclude that p190-RhoGAP is crucially involved in semaphorin signalling to the actin cytoskeleton, via interaction with Plexins.
Stefan Offermanns - One of the best experts on this subject based on the ideXlab platform.
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Plexin b1 mutation drives prostate cancer metastasis
bioRxiv, 2020Co-Authors: Boris Shorning, Thomas Worzfeld, Stefan Offermanns, Neil Trent, David Fr Griffiths, John R Masters, Alan Richard Clarke, Matthew J Smalley, Magali WilliamsonAbstract:Abstract Prostate cancer mortality is associated with the metastatic spread of tumour cells. A better understanding of the mechanisms which allow a locally advanced tumour to disseminate around the body will identify new therapeutic targets to block this process. One of set of genes implicated in metastasis are Plexins, which can promote or suppress tumour progression depending on cancer type and cellular context. We have taken a mouse genetics approach to gain insight into the role of Plexin-B1 in prostate cancer progression in vivo. We show here that genetic deletion of Plexin-B1 in PbCre+Ptenfl/flKrasG12V and PbCre+Ptenfl/flp53fl/fl mouse prostate cancer models significantly decreased metastasis. High levels of prostate epithelial cell-specific expression of wild-type Plexin-B1 in knock-in mice with a PbCre+Ptenfl/flKrasG12V background also significantly decreased metastasis. In contrast, expression of a Plexin-B1 mutant (P1597L; identified from metastatic deposits in prostate cancer patients) in prostate epithelial cells in PbCre+Ptenfl/flKrasG12V and PbCre+Ptenfl/flp53fl/fl mice significantly increased metastasis, in particular metastasis to distant sites. In line with these findings, both deletion and overexpression of wild-type Plexin-B1 reduced invasion of tumour cells into the prostate stroma, while overexpression of mutant Plexin-B1 significantly increased invasion, suggesting that Plexin-B1 has a role in the initial stages of metastasis. Invasion and metastasis also correlated with phosphorylation of myosin light chain, suggesting that Plexin-B1 signals via the Rho/ROCK pathway to promote metastasis. Our results demonstrate that mutant Plexin-B1 promotes metastasis in prostate cancer and represents a new therapeutic target to suppress tumour spread.
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semaphorin 4c Plexin b2 signaling in peripheral sensory neurons is pronociceptive in a model of inflammatory pain
Nature Communications, 2017Co-Authors: Eszter Paldy, Thomas Worzfeld, Manuela Simonetti, Kiran Kumar Bali, Lucas Vicuna, Stefan Offermanns, Rohini KunerAbstract:Semaphorins and their transmembrane receptors, Plexins, are key regulators of axon guidance and development of neuronal connectivity. B-type Plexins respond to Class IV semaphorins and mediate a variety of developmental functions. Here we report that the expression of Plexin-B2 and its high-affinity ligand, Sema4C, persists in peripheral sensory neurons in adult life and is markedly increased in states of persistent pain in mice. Genetic deletion of Sema4C as well as adult-onset loss of Plexin-B2 leads to impairment of the development and duration of inflammatory hypersensitivity. Remarkably, unlike the neurodevelopmental functions of Plexin-B2 that solely rely on Ras signaling, we obtained genetic and pharmacological evidence for a requirement of RhoA-ROCK-dependent mechanisms as well as TRPA1 sensitization in pronociceptive functions of Sema4C-Plexin-B2 signaling in adult life. These results suggest important roles for Plexin-B2 signaling in sensory function that may be of therapeutic relevance in pathological pain. Semaphorins and their receptors are involved in neurodevelopment, but their functions in the adult nervous system are not fully understood. This study finds that semaphorin 4C and its receptor Plexin B are expressed in sensory neurons and are pronociceptive in a mouse model of inflammatory pain.
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a systematic expression analysis implicates Plexin b2 and its ligand sema4c in the regulation of the vascular and endocrine system
Experimental Cell Research, 2010Co-Authors: Matthias Zielonka, Roland H. Friedel, Thomas Worzfeld, Jingjing Xia, Stefan OffermannsAbstract:Plexins serve as receptors for semaphorins and play important roles in the developing nervous system. Plexin-B2 controls decisive developmental programs in the neural tube and cerebellum. However, whether Plexin-B2 also regulates biological functions in adult nonneuronal tissues is unknown. Here we show by two methodologically independent approaches that Plexin-B2 is expressed in discrete cell types of several nonneuronal tissues in the adult mouse. In the vasculature, Plexin-B2 is selectively expressed in functionally specialized endothelial cells. In endocrine organs, Plexin-B2 localizes to the pancreatic islets of Langerhans and to both cortex and medulla of the adrenal gland. Plexin-B2 expression is also detected in certain types of immune and epithelial cells. In addition, we report on a systematic comparison of the expression patterns of Plexin-B2 and its ligand Sema4C, which show complementarity or overlap in some but not all tissues. Furthermore, we demonstrate that Plexin-B2 and its family member Plexin-B1 display largely nonredundant expression patterns. This work establishes Plexin-B2 and Sema4C as potential regulators of the vascular and endocrine system and provides an anatomical basis to understand the biological functions of this ligand-receptor pair.
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semaphorin 4d signaling requires the recruitment of phospholipase cγ into the Plexin b1 receptor complex
Molecular and Cellular Biology, 2009Co-Authors: Jakub M Swiercz, Thomas Worzfeld, Stefan OffermannsAbstract:The semaphorin 4D (Sema4D) receptor Plexin-B1 constitutively interacts with particular Rho guanine nucleotide exchange factors (RhoGEFs) and thereby mediates Sema4D-induced RhoA activation, a process which involves the tyrosine phosphorylation of Plexin-B1 by ErbB-2. It is, however, unknown how Plexin-B1 phosphorylation regulates RhoGEF activity. We show here that activation of Plexin-B1 by Sema4D and its subsequent tyrosine phosphorylation creates docking sites for the SH2 domains of phospholipase Cγ (PLCγ). PLCγ is thereby recruited into the Plexin-B1 receptor complex and via its SH3 domain activates the Rho guanine nucleotide exchange factor PDZ-RhoGEF. PLCγ-dependent RhoGEF activation is independent of its lipase activity. The recruitment of PLCγ has no effect on the R-Ras GTPase-activating protein activity of Plexin-B1 but is required for Sema4D-induced axonal growth cone collapse as well as for the promigratory effects of Sema4D on cancer cells. These data demonstrate a novel nonenzymatic function of PLCγ as an important mechanism of Plexin-mediated signaling which links tyrosine phosphorylation of Plexin-B1 to the regulation of a RhoGEF protein and downstream cellular processes.
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Plexin b1 rhogef mediated rhoa activation involves the receptor tyrosine kinase erbb 2
Journal of Cell Biology, 2004Co-Authors: Jakub M Swiercz, Rohini Kuner, Stefan OffermannsAbstract:Plexins are widely expressed transmembrane proteins that mediate the effects of semaphorins. The molecular mechanisms of Plexin-mediated signal transduction are still rather unclear. Plexin-B1 has recently been shown to mediate activation of RhoA through a stable interaction with the Rho guanine nucleotide exchange factors PDZ-RhoGEF and LARG. However, it is unclear how the activity of Plexin-B1 and its downstream effectors is regulated by its ligand Sema4D. Here, we show that Plexin-B family members stably associate with the receptor tyrosine kinase ErbB-2. Binding of Sema4D to Plexin-B1 stimulates the intrinsic tyrosine kinase activity of ErbB-2, resulting in the phosphorylation of both Plexin-B1 and ErbB-2. A dominant-negative form of ErbB-2 blocks Sema4D-induced RhoA activation as well as axonal growth cone collapse in primary hippocampal neurons. Our data indicate that ErbB-2 is an important component of the Plexin-B receptor system and that ErbB-2–mediated phosphorylation of Plexin-B1 is critically involved in Sema4D-induced RhoA activation, which underlies cellular phenomena downstream of Plexin-B1, including axonal growth cone collapse.
Thomas Worzfeld - One of the best experts on this subject based on the ideXlab platform.
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Plexin b1 mutation drives prostate cancer metastasis
bioRxiv, 2020Co-Authors: Boris Shorning, Thomas Worzfeld, Stefan Offermanns, Neil Trent, David Fr Griffiths, John R Masters, Alan Richard Clarke, Matthew J Smalley, Magali WilliamsonAbstract:Abstract Prostate cancer mortality is associated with the metastatic spread of tumour cells. A better understanding of the mechanisms which allow a locally advanced tumour to disseminate around the body will identify new therapeutic targets to block this process. One of set of genes implicated in metastasis are Plexins, which can promote or suppress tumour progression depending on cancer type and cellular context. We have taken a mouse genetics approach to gain insight into the role of Plexin-B1 in prostate cancer progression in vivo. We show here that genetic deletion of Plexin-B1 in PbCre+Ptenfl/flKrasG12V and PbCre+Ptenfl/flp53fl/fl mouse prostate cancer models significantly decreased metastasis. High levels of prostate epithelial cell-specific expression of wild-type Plexin-B1 in knock-in mice with a PbCre+Ptenfl/flKrasG12V background also significantly decreased metastasis. In contrast, expression of a Plexin-B1 mutant (P1597L; identified from metastatic deposits in prostate cancer patients) in prostate epithelial cells in PbCre+Ptenfl/flKrasG12V and PbCre+Ptenfl/flp53fl/fl mice significantly increased metastasis, in particular metastasis to distant sites. In line with these findings, both deletion and overexpression of wild-type Plexin-B1 reduced invasion of tumour cells into the prostate stroma, while overexpression of mutant Plexin-B1 significantly increased invasion, suggesting that Plexin-B1 has a role in the initial stages of metastasis. Invasion and metastasis also correlated with phosphorylation of myosin light chain, suggesting that Plexin-B1 signals via the Rho/ROCK pathway to promote metastasis. Our results demonstrate that mutant Plexin-B1 promotes metastasis in prostate cancer and represents a new therapeutic target to suppress tumour spread.
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semaphorin 4c Plexin b2 signaling in peripheral sensory neurons is pronociceptive in a model of inflammatory pain
Nature Communications, 2017Co-Authors: Eszter Paldy, Thomas Worzfeld, Manuela Simonetti, Kiran Kumar Bali, Lucas Vicuna, Stefan Offermanns, Rohini KunerAbstract:Semaphorins and their transmembrane receptors, Plexins, are key regulators of axon guidance and development of neuronal connectivity. B-type Plexins respond to Class IV semaphorins and mediate a variety of developmental functions. Here we report that the expression of Plexin-B2 and its high-affinity ligand, Sema4C, persists in peripheral sensory neurons in adult life and is markedly increased in states of persistent pain in mice. Genetic deletion of Sema4C as well as adult-onset loss of Plexin-B2 leads to impairment of the development and duration of inflammatory hypersensitivity. Remarkably, unlike the neurodevelopmental functions of Plexin-B2 that solely rely on Ras signaling, we obtained genetic and pharmacological evidence for a requirement of RhoA-ROCK-dependent mechanisms as well as TRPA1 sensitization in pronociceptive functions of Sema4C-Plexin-B2 signaling in adult life. These results suggest important roles for Plexin-B2 signaling in sensory function that may be of therapeutic relevance in pathological pain. Semaphorins and their receptors are involved in neurodevelopment, but their functions in the adult nervous system are not fully understood. This study finds that semaphorin 4C and its receptor Plexin B are expressed in sensory neurons and are pronociceptive in a mouse model of inflammatory pain.
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genetic dissection of Plexin signaling in vivo
Proceedings of the National Academy of Sciences of the United States of America, 2014Co-Authors: Suhua Deng, Thomas Worzfeld, Alexander Korostylev, Jakub M Swiercz, Aycan Senturk, Berit Genz, Jingjing Xia, Mikio HoshinoAbstract:Mammalian Plexins constitute a family of transmembrane receptors for semaphorins and represent critical regulators of various processes during development of the nervous, cardiovascular, skeletal, and renal system. In vitro studies have shown that Plexins exert their effects via an intracellular R-Ras/M-Ras GTPase-activating protein (GAP) domain or by activation of RhoA through interaction with Rho guanine nucleotide exchange factor proteins. However, which of these signaling pathways are relevant for Plexin functions in vivo is largely unknown. Using an allelic series of transgenic mice, we show that the GAP domain of Plexins constitutes their key signaling module during development. Mice in which endogenous Plexin-B2 or Plexin-D1 is replaced by transgenic versions harboring mutations in the GAP domain recapitulate the phenotypes of the respective null mutants in the developing nervous, vascular, and skeletal system. We further provide genetic evidence that, unexpectedly, the GAP domain-mediated developmental functions of Plexins are not brought about via R-Ras and M-Ras inactivation. In contrast to the GAP domain mutants, Plexin-B2 transgenic mice defective in Rho guanine nucleotide exchange factor binding are viable and fertile but exhibit abnormal development of the liver vasculature. Our genetic analyses uncover the in vivo context-dependence and functional specificity of individual Plexin-mediated signaling pathways during development.
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a systematic expression analysis implicates Plexin b2 and its ligand sema4c in the regulation of the vascular and endocrine system
Experimental Cell Research, 2010Co-Authors: Matthias Zielonka, Roland H. Friedel, Thomas Worzfeld, Jingjing Xia, Stefan OffermannsAbstract:Plexins serve as receptors for semaphorins and play important roles in the developing nervous system. Plexin-B2 controls decisive developmental programs in the neural tube and cerebellum. However, whether Plexin-B2 also regulates biological functions in adult nonneuronal tissues is unknown. Here we show by two methodologically independent approaches that Plexin-B2 is expressed in discrete cell types of several nonneuronal tissues in the adult mouse. In the vasculature, Plexin-B2 is selectively expressed in functionally specialized endothelial cells. In endocrine organs, Plexin-B2 localizes to the pancreatic islets of Langerhans and to both cortex and medulla of the adrenal gland. Plexin-B2 expression is also detected in certain types of immune and epithelial cells. In addition, we report on a systematic comparison of the expression patterns of Plexin-B2 and its ligand Sema4C, which show complementarity or overlap in some but not all tissues. Furthermore, we demonstrate that Plexin-B2 and its family member Plexin-B1 display largely nonredundant expression patterns. This work establishes Plexin-B2 and Sema4C as potential regulators of the vascular and endocrine system and provides an anatomical basis to understand the biological functions of this ligand-receptor pair.
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semaphorin 4d signaling requires the recruitment of phospholipase cγ into the Plexin b1 receptor complex
Molecular and Cellular Biology, 2009Co-Authors: Jakub M Swiercz, Thomas Worzfeld, Stefan OffermannsAbstract:The semaphorin 4D (Sema4D) receptor Plexin-B1 constitutively interacts with particular Rho guanine nucleotide exchange factors (RhoGEFs) and thereby mediates Sema4D-induced RhoA activation, a process which involves the tyrosine phosphorylation of Plexin-B1 by ErbB-2. It is, however, unknown how Plexin-B1 phosphorylation regulates RhoGEF activity. We show here that activation of Plexin-B1 by Sema4D and its subsequent tyrosine phosphorylation creates docking sites for the SH2 domains of phospholipase Cγ (PLCγ). PLCγ is thereby recruited into the Plexin-B1 receptor complex and via its SH3 domain activates the Rho guanine nucleotide exchange factor PDZ-RhoGEF. PLCγ-dependent RhoGEF activation is independent of its lipase activity. The recruitment of PLCγ has no effect on the R-Ras GTPase-activating protein activity of Plexin-B1 but is required for Sema4D-induced axonal growth cone collapse as well as for the promigratory effects of Sema4D on cancer cells. These data demonstrate a novel nonenzymatic function of PLCγ as an important mechanism of Plexin-mediated signaling which links tyrosine phosphorylation of Plexin-B1 to the regulation of a RhoGEF protein and downstream cellular processes.
Izumi Oinuma - One of the best experts on this subject based on the ideXlab platform.
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Plexin b1 is a gtpase activating protein for m ras remodelling dendrite morphology
EMBO Reports, 2009Co-Authors: Yasuhiro Saito, Izumi Oinuma, Satoshi Fujimoto, Manabu NegishiAbstract:Plexins are receptors for axonal guidance molecules known as semaphorins. We recently reported that the semaphorin 4D (Sema4D) receptor, Plexin-B1, induces axonal growth cone collapse by functioning as an R-Ras GTPase activating protein (GAP). Here, we report that Plexin-B1 shows GAP activity for M-Ras, another member of the Ras family of GTPases. In cortical neurons, the expression of M-Ras was upregulated during dendritic development. Knockdown of endogenous M-Ras—but not R-Ras—reduced dendritic outgrowth and branching, whereas overexpression of constitutively active M-Ras, M-Ras(Q71L), enhanced dendritic outgrowth and branching. Sema4D suppressed M-Ras activity and reduced dendritic outgrowth and branching, but this reduction was blocked by M-Ras(Q71L). M-Ras(Q71L) stimulated extracellular signal-regulated kinase (ERK) activation, inducing dendrite growth, whereas Sema4D suppressed ERK activity and down-regulation of ERK was required for a Sema4D-induced reduction of dendrite growth. Thus, we conclude that Plexin-B1 is a dual functional GAP for R-Ras and M-Ras, remodelling axon and dendrite morphology, respectively.
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Different Requirement for Rnd GTPases of R-Ras GAP Activity of Plexin-C1 and Plexin-D1
The Journal of biological chemistry, 2009Co-Authors: Kanami Uesugi, Hironori Katoh, Izumi Oinuma, Manabu NegishiAbstract:Plexins, comprising Plexin-A, -B, -C, and -D subfamilies, are receptors for semaphorins governing cell adhesion, migration, and axon guidance. Among Plexin subfamilies, Plexin-A1 and Plexin-B1 have been shown to function as an R-Ras GAP, inducing repulsive responses, and the expression of R-Ras GAP activity requires the binding of Rnd1, a member of Rnd subfamily of Rho GTPases. However, signaling pathways of Plexin-D1 and Plexin-C1 still remain obscure. Here, we found that Plexin-D1 displayed R-Ras GAP activity and inhibited migration of COS-7 cells, and these actions required Rnd2, another Rnd subfamily GTPase. Rnd2 bound to Plexin-D1 in cortical neurons, and Sema3E/Plexin-D1-induced inhibition of axon outgrowth of cortical neurons required Rnd2 and down-regulation of R-Ras activity. On the other hand, Plexin-C1 displayed R-Ras GAP activity and inhibited cell migration of COS-7 cells without Rnd proteins. Therefore, R-Ras GAP activity is a common function of Plexin subfamilies but the regulation of R-Ras GAP activity of Plexins by Rnd proteins is different among Plexin subfamilies.
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sema4d Plexin b1 activates gsk 3β through r ras gap activity inducing growth cone collapse
EMBO Reports, 2006Co-Authors: Yuri Ito, Hironori Katoh, Izumi Oinuma, Kozo Kaibuchi, Manabu NegishiAbstract:Plexins are receptors for the axonal guidance molecules known as semaphorins, and the semaphorin 4D (Sema4D) receptor Plexin-B1 induces repulsive responses by functioning as an R-Ras GTPase-activating protein (GAP). Here we characterized the downstream signalling of Plexin-B1-mediated R-Ras GAP activity, inducing growth cone collapse. Sema4D suppressed R-Ras activity in hippocampal neurons, in parallel with dephosphorylation of Akt and activation of glycogen synthase kinase (GSK)-3β. Ectopic expression of the constitutively active mutant of Akt or treatment with GSK-3 inhibitors suppressed the Sema4D-induced growth cone collapse. Constitutive activation of phosphatidylinositol-3-OH kinase (PI(3)K), an upstream kinase of Akt and GSK-3β, also blocked the growth cone collapse. The R-Ras GAP activity was necessary for Plexin-B1-induced dephosphorylation of Akt and activation of GSK-3β and was also required for phosphorylation of a downstream kinase of GSK-3β, collapsin response mediator protein-2. Plexin-A1 also induced dephosphorylation of Akt and GSK-3β through its R-Ras GAP activity. We conclude that Plexin-B1 inactivates PI(3)K and dephosphorylates Akt and GSK-3β through R-Ras GAP activity, inducing growth cone collapse.
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Molecular Dissection of the Semaphorin 4D Receptor Plexin-B1-Stimulated R-Ras GTPase-Activating Protein Activity and Neurite Remodeling in Hippocampal Neurons
The Journal of neuroscience : the official journal of the Society for Neuroscience, 2004Co-Authors: Izumi Oinuma, Hironori Katoh, Manabu NegishiAbstract:Plexins serve as receptors for repulsive axonal guidance molecules semaphorins. The cytoplasmic domain of the semaphorin 4D (Sema4D) receptor, Plexin-B1 has two separated Ras GTPase-activating protein (GAP)-homologous domains, C1 and C2. Recently, we reported that the Rho family small GTPase Rnd1 associates with Plexin-B1, and the Plexin-B1-Rnd1 complex stimulates GTPase activity of R-Ras, inducing growth cone collapse in hippocampal neurons in response to Sema4D. However, the molecular mechanisms by which Plexin-B1 exhibits the GAP activity remain unclear. In this report, critical roles of Rnd1 and Sema4D in Plexin-B1-stimulated R-Ras GAP activity and neurite remodeling were examined. The N-terminal region of the cytoplasmic domain of Plexin-B1 containing the C1 domain interacts with the C-terminal region containing the C2 domain, and Rnd1 disrupts this interaction. On the other hand, Sema4D induces clustering of Rnd1-bound Plexin-B1, in parallel with inactivation of R-Ras in cells. Antibody clustering of the recombinant cytoplasmic domain of Plexin-B1 in the presence of Rnd1 triggers the R-Ras GAP activity. Deletion of the extracellular domain of Plexin-B1 causes ligand-independent clustering of the receptor, rendering the receptor constitutively active in the presence of Rnd1, and induces contraction of COS-7 cells and inhibition of neurite outgrowth in hippocampal neurons. These results indicate that Rnd1 opens the two R-Ras GAP domains of Plexin-B1, and Sema4D-induced receptor clustering stimulates R-Ras GAP activity and neurite remodeling in hippocampal neurons.
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the semaphorin 4d receptor Plexin b1 is a gtpase activating protein for r ras
Science, 2004Co-Authors: Izumi Oinuma, Hironori Katoh, Yukio Ishikawa, Manabu NegishiAbstract:Plexins are cell surface receptors for semaphorin molecules, and their interaction governs cell adhesion and migration in a variety of tissues. We report that the Semaphorin 4D (Sema4D) receptor Plexin-B1 directly stimulates the intrinsic guanosine triphosphatase (GTPase) activity of R-Ras, a member of the Ras superfamily of small GTP-binding proteins that has been implicated in promoting cell adhesion and neurite outgrowth. This activity required the interaction of Plexin-B1 with Rnd1, a small GTP-binding protein of the Rho family. Down-regulation of R-Ras activity by the Plexin-B1-Rnd1 complex was essential for the Sema4D-induced growth cone collapse in hippocampal neurons. Thus, Plexin-B1 mediates Sema4D-induced repulsive axon guidance signaling by acting as a GTPase activating protein for R-Ras.