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RelA stability
Abstracts / Neuroscience Research 58S (2007) S1–S244
O1P-KØ1 Ras activation of Rac1 exchange factor Tiam1 mediates neurotrophin-3-induced Schwann cell migration Junji Yamauchi 1 , Yuki Miyamoto 1 , Atsushi Sanbe 1 , Chan R. Jonah 2 , Shooter M. Eric 2 , Akito Tanoue 1 1 Department of Pharmacology, NICHD, Tokyo, Japan; 2 Department of Neurobiology, Stanford University, Stanford, USA Endogenous neurotrophins positively and negatively regulate migration of premyelinating Schwann cells before the initiation of myelination. Neurotrophinp-3 (NT3) acting through the TrkC receptor tyrosine kinase stimulates Schwann cell migration through the Rho GTPases Rac1 and Cdc42. We previously demonstrated that the TrkC kinase phosphorylates and activates Dbs, the guanine-nucleotide exchange factor (GEF) for Cdc42, to mediate Schwann cell migration. Here we identify T-lymphoma invasion and metastasis (Tiam) 1 as the Rac1-specific GEF involved in NT3-induced Schwann cell migration. Furthermore, the interaction between a small GTPase Ras and Tiam1 plays an essential role in the activation of Rac1. Taken together, these results suggest that NT3 activation of TrkC stimulates Schwann cell migration through the two parallel signaling units, Ras/Tiam1/Rac1 and Dbs/Cdc42, and that the former is uniquely regulated by two different types of small GTPases.
O1P-KØ2 Tyrosine-phosphorylation of Tiam1 mediates BDNFinduced neurite outgrowth in cortical neurons Yuki Miyamoto 1 , Junji Yamauchi 1 , Atsushi Sanbe 1 , Mobley C. William 2 , Akito Tanoue 1 1 Department of Pharmacology, NICHD, Tokyo, Japan; 2 Department of Neurology, Stanford University, Stanford, USA Small GTPases of the Rho family play key roles in the formation of neuronal axons and dendrites by transducing signals from guidance cues, such as neurotrophins, to the actin cytoskeleton. However, there is little insight into the mechanism by which neurotrophins regulate Rho GTPases. Here we show the crucial role of the ubiquitous Rac1specific guanine-nucleotide exchange factor (GEF), Tiam1, in transducing neurotrophin-mediated neurite outgrowth. We demonstrated that the brain-derived neurotrophic factor (BDNF), acting through the TrkB receptor tyrosine kinase, directly binds and specifically activates Tiam1 by phosphorylating Tyr-829, leading to Rac1 activation and neurite outgrowth of cortical neurons. A point mutation in Tiam1, Tyr-829-to-Phe-829, blocked BDNF-induced changes in number of neurites of cortical neurons. The findings are evidence of a novel mechanism for the activation of Tiam1 and of a role for this effector in neurotrophin-mediated signal transduction leading to neurite outgrowth.
O1P-KØ4 Activation of STAT3 by pituitary adenylate cyclaseactivating polypeptide (PACAP) during PACAP-promoted neuritogenesis of PC12 cells Masami Ishido Environmental Risk Research Programme, National Institute for Environmental Studies PACAP is a 38-amino-acid neuropeptide that was first isolated from ovine hypothalamic extracts on the basis of its ability to stimulate cAMP formation in anterior pituitary cells. Addition of PACAP into the cultured PC12 cells promoted neurite outgrowth of the cells. Using DNA macroarray techniques, we have characterized the PC12 cell transcriptome, revealing that several genes were regulated by PACAP. Among many, we focused to investigate whether STAT3 molecule might be involved in PACAP signaling. PACAP increased gene expression of STAT3, at least up to 24 h, being maximum 9.5-fold with 3 h treatment of 1 nM PACAP. ELISA revealed that PACAP increased the amount of STAT3 proteins about 30%. PhosphoSTAT3 Tyr705 was increased in the nuclei by the neuropeptide. During the activation of STAT3, IL-6 was also secreted by PACAP in dose dependent manner. Treatment with IL-6-neutralizing antibody significantly diminished PACAP-activated STAT3 promoter activity. Together, our findings have demonstrated that STAT3 is involved in PACAP signaling and function.
S49
O1P-KØ5
Synaptic ubiquitin-specific protease (synUSP) is transported to nucleus and regulates p65/RelA stability Qing-Bao Tian, Tatsuo Suzuki Department of Neuroplasticity, Shinshu University Graduate School of Medicine, Matsumoto, Japan It is well known that the ubiquitin-mediated proteolysis of Ikappa B regulates nuclear factor-kappaB (NF-kappaB) signaling. Recently, presence of another, but a similar regulation mechanism has been suggested by the report in which the ubiquitination of p65/RelA, one of the NF-kappaB subunit, was mediated by E2 ubiquitin-conjugating enzyme, ubcH5alpha, and E3 ubiquitin ligase, SOSC-1 (Ryo et al., Mol. Cell 12, 1413–1426). However, the detailed mechanisms of the regulation through the p65/RelA ubiquitination remain unknown. Here we propose a novel mechanism of regulation of NF-kappaB signaling through the action of synaptic ubiquitin-specific protease (synUSP). The synUSP, also known as USP48 or USP31, is a de-ubiquitinating enzyme which we found in 2003 (Tian et al., J. Neurochem. 87, 665–675). We demonstrated that synUSP was transported to the nucleus and increased the stability of p65/RelA. Nuclear translocation of USP48 is blocked by deletion of its nuclear localization signal (NLS), which suggested a role of classic nuclear transport pathway.
O1P-KØ6 Involvement of ERK cascades in the anterior cingulate cortex in pain-related negative emotion in rat Hong Cao Institute of Neurobiology, Fudan University, Shanghai, China Activation of ERK in dorsal horn neurons of the spinal cord by peripheral noxious stimulation contributes to both short-term and long-term pain hypersensitivity. Mounting evidence indicates that the ACC is involved in pain emotion. Our study is to investigate the activation of ERK in the ACC and its contribution to pain-related negative emotion using formalininduced conditioned place avoidance animal model. Intraplantar injection of 5% formalin induced a rapid and sustained ERK activation in ACC neurons. Intra-ACC microinjection of MEK inhibitor PD098059 or U0126 could significantly block the formation of F-CPA. Incubation of ACC slices with NMDA, Forskolin or Sp-cAMP produced significant activation of ERK, which was suppressed by preincubation with their respective antagonists APV, SQ22536 and Rp-cAMP. Similarly, F-CPA was blocked by premicroinjection of SQ22536 or Rp-cAMP. Taken together, NMDARdependent AC-PKA-ERK signal transduction may play an important role in pain-related negative emotion. Research funds: NIH, FIRCA (USA, TW7180); NSFC (30330230 and 30370471); the NSFC for Distinguished Young Scholars (30425022)
O1P-KØ7 Fyn protein tyrosine kinase as a target for antipsychotic drugs Kotaro Hattori 1 , Shigeo Uchino 2 , Tomoko Isosaka 1,3 , Shinichi Kohsaka 2 , Takeshi Yagi 3 , Shigeki Yuasa 1 1 Department of Ultrastructural Research, National Institute of Neuroscience, NCNP, Japan; 2 Department of Neurochemistry, National Institute of Neuroscience, NCNP, Japan; 3 KOKORO Biology Group, FBS, Osaka University, Japan Antipsychotics exert their effect through the blockade of dopamine D2 receptor (D2-R), but the signal cascade triggered by D2-R blockade remains unclear. Recently, we have reported Fyn tyrosine kinase mediates that cascade. In the control mice, D2-R blockers induced (1) a behavioral response, catalepsy, (2) Fyn activation and enhanced tyrosine phosphorylation of a NMDA-R subunit in the striatum, and (3) increased NMDA-R channel activity of striatal neurons, but these responses were significantly reduced in Fyn-deficient mice. Thus, Fyn mediates intracellular signaling between D2-R and NMDA-R in the striatum which mediates a behavioral response. Further proteomics analysis on antipsychotics-induced tyrosine phosphorylation and spatiotemporal analysis of Fyn activation are now in progress. Our present findings strongly suggest that Fyn links the signaling downstream of D2-R, and can be a new target for antipsychotic drugs. Research funds: NIBIO Grant 05-32
O1P-KØ1 Ras activation of Rac1 exchange factor Tiam1 mediates neurotrophin-3-induced Schwann cell migration Junji Yamauchi 1 , Yuki Miyamoto 1 , Atsushi Sanbe 1 , Chan R. Jonah 2 , Shooter M. Eric 2 , Akito Tanoue 1 1 Department of Pharmacology, NICHD, Tokyo, Japan; 2 Department of Neurobiology, Stanford University, Stanford, USA Endogenous neurotrophins positively and negatively regulate migration of premyelinating Schwann cells before the initiation of myelination. Neurotrophinp-3 (NT3) acting through the TrkC receptor tyrosine kinase stimulates Schwann cell migration through the Rho GTPases Rac1 and Cdc42. We previously demonstrated that the TrkC kinase phosphorylates and activates Dbs, the guanine-nucleotide exchange factor (GEF) for Cdc42, to mediate Schwann cell migration. Here we identify T-lymphoma invasion and metastasis (Tiam) 1 as the Rac1-specific GEF involved in NT3-induced Schwann cell migration. Furthermore, the interaction between a small GTPase Ras and Tiam1 plays an essential role in the activation of Rac1. Taken together, these results suggest that NT3 activation of TrkC stimulates Schwann cell migration through the two parallel signaling units, Ras/Tiam1/Rac1 and Dbs/Cdc42, and that the former is uniquely regulated by two different types of small GTPases.
O1P-KØ2 Tyrosine-phosphorylation of Tiam1 mediates BDNFinduced neurite outgrowth in cortical neurons Yuki Miyamoto 1 , Junji Yamauchi 1 , Atsushi Sanbe 1 , Mobley C. William 2 , Akito Tanoue 1 1 Department of Pharmacology, NICHD, Tokyo, Japan; 2 Department of Neurology, Stanford University, Stanford, USA Small GTPases of the Rho family play key roles in the formation of neuronal axons and dendrites by transducing signals from guidance cues, such as neurotrophins, to the actin cytoskeleton. However, there is little insight into the mechanism by which neurotrophins regulate Rho GTPases. Here we show the crucial role of the ubiquitous Rac1specific guanine-nucleotide exchange factor (GEF), Tiam1, in transducing neurotrophin-mediated neurite outgrowth. We demonstrated that the brain-derived neurotrophic factor (BDNF), acting through the TrkB receptor tyrosine kinase, directly binds and specifically activates Tiam1 by phosphorylating Tyr-829, leading to Rac1 activation and neurite outgrowth of cortical neurons. A point mutation in Tiam1, Tyr-829-to-Phe-829, blocked BDNF-induced changes in number of neurites of cortical neurons. The findings are evidence of a novel mechanism for the activation of Tiam1 and of a role for this effector in neurotrophin-mediated signal transduction leading to neurite outgrowth.
O1P-KØ4 Activation of STAT3 by pituitary adenylate cyclaseactivating polypeptide (PACAP) during PACAP-promoted neuritogenesis of PC12 cells Masami Ishido Environmental Risk Research Programme, National Institute for Environmental Studies PACAP is a 38-amino-acid neuropeptide that was first isolated from ovine hypothalamic extracts on the basis of its ability to stimulate cAMP formation in anterior pituitary cells. Addition of PACAP into the cultured PC12 cells promoted neurite outgrowth of the cells. Using DNA macroarray techniques, we have characterized the PC12 cell transcriptome, revealing that several genes were regulated by PACAP. Among many, we focused to investigate whether STAT3 molecule might be involved in PACAP signaling. PACAP increased gene expression of STAT3, at least up to 24 h, being maximum 9.5-fold with 3 h treatment of 1 nM PACAP. ELISA revealed that PACAP increased the amount of STAT3 proteins about 30%. PhosphoSTAT3 Tyr705 was increased in the nuclei by the neuropeptide. During the activation of STAT3, IL-6 was also secreted by PACAP in dose dependent manner. Treatment with IL-6-neutralizing antibody significantly diminished PACAP-activated STAT3 promoter activity. Together, our findings have demonstrated that STAT3 is involved in PACAP signaling and function.
S49
O1P-KØ5
Synaptic ubiquitin-specific protease (synUSP) is transported to nucleus and regulates p65/RelA stability Qing-Bao Tian, Tatsuo Suzuki Department of Neuroplasticity, Shinshu University Graduate School of Medicine, Matsumoto, Japan It is well known that the ubiquitin-mediated proteolysis of Ikappa B regulates nuclear factor-kappaB (NF-kappaB) signaling. Recently, presence of another, but a similar regulation mechanism has been suggested by the report in which the ubiquitination of p65/RelA, one of the NF-kappaB subunit, was mediated by E2 ubiquitin-conjugating enzyme, ubcH5alpha, and E3 ubiquitin ligase, SOSC-1 (Ryo et al., Mol. Cell 12, 1413–1426). However, the detailed mechanisms of the regulation through the p65/RelA ubiquitination remain unknown. Here we propose a novel mechanism of regulation of NF-kappaB signaling through the action of synaptic ubiquitin-specific protease (synUSP). The synUSP, also known as USP48 or USP31, is a de-ubiquitinating enzyme which we found in 2003 (Tian et al., J. Neurochem. 87, 665–675). We demonstrated that synUSP was transported to the nucleus and increased the stability of p65/RelA. Nuclear translocation of USP48 is blocked by deletion of its nuclear localization signal (NLS), which suggested a role of classic nuclear transport pathway.
O1P-KØ6 Involvement of ERK cascades in the anterior cingulate cortex in pain-related negative emotion in rat Hong Cao Institute of Neurobiology, Fudan University, Shanghai, China Activation of ERK in dorsal horn neurons of the spinal cord by peripheral noxious stimulation contributes to both short-term and long-term pain hypersensitivity. Mounting evidence indicates that the ACC is involved in pain emotion. Our study is to investigate the activation of ERK in the ACC and its contribution to pain-related negative emotion using formalininduced conditioned place avoidance animal model. Intraplantar injection of 5% formalin induced a rapid and sustained ERK activation in ACC neurons. Intra-ACC microinjection of MEK inhibitor PD098059 or U0126 could significantly block the formation of F-CPA. Incubation of ACC slices with NMDA, Forskolin or Sp-cAMP produced significant activation of ERK, which was suppressed by preincubation with their respective antagonists APV, SQ22536 and Rp-cAMP. Similarly, F-CPA was blocked by premicroinjection of SQ22536 or Rp-cAMP. Taken together, NMDARdependent AC-PKA-ERK signal transduction may play an important role in pain-related negative emotion. Research funds: NIH, FIRCA (USA, TW7180); NSFC (30330230 and 30370471); the NSFC for Distinguished Young Scholars (30425022)
O1P-KØ7 Fyn protein tyrosine kinase as a target for antipsychotic drugs Kotaro Hattori 1 , Shigeo Uchino 2 , Tomoko Isosaka 1,3 , Shinichi Kohsaka 2 , Takeshi Yagi 3 , Shigeki Yuasa 1 1 Department of Ultrastructural Research, National Institute of Neuroscience, NCNP, Japan; 2 Department of Neurochemistry, National Institute of Neuroscience, NCNP, Japan; 3 KOKORO Biology Group, FBS, Osaka University, Japan Antipsychotics exert their effect through the blockade of dopamine D2 receptor (D2-R), but the signal cascade triggered by D2-R blockade remains unclear. Recently, we have reported Fyn tyrosine kinase mediates that cascade. In the control mice, D2-R blockers induced (1) a behavioral response, catalepsy, (2) Fyn activation and enhanced tyrosine phosphorylation of a NMDA-R subunit in the striatum, and (3) increased NMDA-R channel activity of striatal neurons, but these responses were significantly reduced in Fyn-deficient mice. Thus, Fyn mediates intracellular signaling between D2-R and NMDA-R in the striatum which mediates a behavioral response. Further proteomics analysis on antipsychotics-induced tyrosine phosphorylation and spatiotemporal analysis of Fyn activation are now in progress. Our present findings strongly suggest that Fyn links the signaling downstream of D2-R, and can be a new target for antipsychotic drugs. Research funds: NIBIO Grant 05-32