一般ポスター

一般ポスター神経細胞の分子基盤

7月7日（金）　13:50-14:50　ポスター会場① 2P①-1 神経細胞におけるBDNFのRNAスプライシング制御機構に関する解析 Analysis of regulatory mechanism for splicing of BDNF RNA in neurons 福地　守, 赤澤　祐斗, 柴崎　由実高崎健康福祉大薬　分子神経科学 Mamoru Fukuchi, Yuto Akazawa, Yumi Shibasaki Lab. of Mol. Neurosci., Fac. of Pharm., Takasaki Univ. of Hlth & Welfare, Gunma, Japan The expression of the BDNF gene is controlled by its unique mechanisms. BDNF gene consists of eight untranslated exons (exon I-VIII) and one exon containing coding sequence (exon IX). Because of alternative promoters located upstream of each exon and RNA splicing, multiple BDNF transcripts are produced. However, the mechanisms for the splicing of BDNF RNA remain unclear. In this study, we analyzed the regulatory mechanisms for the splicing of BDNF RNA. Here, we particularly focused on the splicing between exon I and exon IX of the BDNF gene (BDNF exon I-IX) using a mini-gene containing the regions around rat BDNF exon I and exon IX. We found that spliced BDNF exon I-IX RNA was mainly detected in cultured cortical neurons, whereas unspliced BDNF exon I-IX RNA was also detected in non-neuronal NIH3T3, suggesting that RNA splicing of BDNF exon I-IX could preferentially occur in neurons. To identify enhancers of neuronal splicing of BDNF exon I-IX, expression vectors of a series of RNA-binding proteins were co-transfected with the mini-gene into NIH3T3, and we found that the overexpression of Nova1 or Nova2 enhanced splicing of BDNF exon I-IX. In support, the expression levels of endogenous Nova1/2 in neurons were markedly higher than those in NIH3T3. Our current study strongly suggests that Nova1/2 would be candidates for splicing enhancers of BDNF exon I-IX in neurons.		7月7日（金）　13:50-14:50　ポスター会場① 2P①-2 Cdk5 organizes gatekeeper function of the axon initial segment. 吉村　武¹, ラズバンド　マシュー², 片山　泰一¹ 1. 大阪大・院・連合小児・分子生物遺伝学, 2. ベイラー医科大・神経科学・米国 Takeshi Yoshimura¹, Matthew Rasband², Taiichi Katayama¹ 1. Dept. of Child Dev. & Mol. Brain Sci., UGSCD, Osaka Univ., 2. Dept. of Neurosci., Baylor College of Med., USA The axon initial segment (AIS) is the gatekeeper of neurons. The AIS acts as a boundary between the somatodendritic and axonal compartments and maintains the distinct molecular identity of the axon. The AIS has a specific cytoskeletal structure consisting of αII-spectrin, βIV-spectrin, ankyrin-G and actin. Super-resolution microscopy has revealed that these proteins form a remarkable periodic lattice lining the cytoplasmic face of the AIS membrane. AIS proteins have been implicated in a variety of human diseases. αII-spectrin is associated with West syndrome. Human mutations in ankyrin-G were reported and shown to be associated with severe intellectual disability, attention deficit hyperactivity disorder, and autism spectrum disorder. While recent studies have identified the AIS proteome, the molecular mechanism by which AIS formation and gatekeeper function are regulated remains unclear. In this study, we report that Cdk5 phosphorylates AIS cytoskeletal proteins. Inhibition of Cdk5 impaired AIS formation and gatekeeper function. These results suggest that Cdk5 regulates AIS formation and gatekeeper function through the phosphorylation of AIS cytoskeletal proteins.

7月7日（金）　13:50-14:50　ポスター会場① 2P①-3 Propofol誘発性PKC Translocationの時空間的解析 Spatiotemporal analysis of propofol-induced PKC translocation 野口　颯真¹, 梶本　武利³, 卜部　智晶², 楢崎　壮志^1,2, 原田　佳奈¹, 田中　茂¹, 秀　和泉¹, 酒井　規雄¹ 1. 広島大学　大学院医系科学研究科　神経薬理学, 2. 広島大学　大学院医系科学研究科　麻酔蘇生学, 3. 神戸大学　大学院医学研究科　生化学・分子生物学講座　生化学分野 Soma Noguchi¹, Taketoshi Kajimoto³, Tomoaki Urabe², Soshi Narasaki^1,2, Kana Harada¹, Shigeru Tanaka¹, Izumi Hide¹, Norio Sakai¹ 1. Dept. of Pharmacol. Neurosci., Grad. Sch. of Biomedsci., Hiroshima Univ., Hiroshima, Japan, 2. Dept. of Anesthesiology and Critical Care, Grad. Sch. of Biomedsci., Hiroshima Univ., Hiroshima, Japan, 3. Div. of Biochem., Dept. of Biochem. and Mol. Biol., Kobe Univ. Grad. Sch. of Med., Kobe, Japan Protein kinase C (PKC) changes its localization upon various stimuli and exerts its function at the localized sites, which is called PKC translocation. We have previously shown that propofol, an intravenous anesthetic, induces PKC translocation. In this study, we attempted to further analyze propofol-induced PKC translocation.We selected PKCα of conventional PKC, PKCδ of novel PKC, and PKCζ of atypical PKC. We transiently expressed GFP-tagged PKCs in HeLa cells. Propofol-induced PKC translocation was observed by time-lapse imaging. Administration of propofol at 100 μM mainly translocated PKCα and PKCδ to the plasma membrane (PM), also to Golgi apparatus in case of PKCδ. PKCζ was translocated into nucleus. These results revealed that propofol-induced translocation is PKC subtypes specific. Analysis using C kinase activity reporter (CKAR) showed that PKC was activated at the PM and Golgi, suggesting that penetrated propofol locally activates PKC. Proteins other than PKCζ also translocated into the nucleus, indicating that the nuclear translocation mechanism is not PKC-specific. After the nuclear translocation, the protein concentration inside and outside the nucleus became uniform, suggesting that propofol alters nuclear membrane permeability independently of nuclear localization or export signals. These findings may contribute to the exertion of the various effects of propofol.		7月7日（金）　13:50-14:50　ポスター会場① 2P①-4 Aromatic-turmerone 類縁体がシャペロン介在性オートファジー及びミクロオートファジー活性に及ぼす影響 Effect of aromatic-turmerone analogues on the activities of chaperone-mediated autophagy and microautophagy 本村　健祐¹, アレックス　ボアテング², 杉浦　正晴², 倉内　祐樹¹, 香月　博志¹, 関　貴弘³ 1. 熊本大学　大学院薬学研究科　薬物活性学, 2. 崇城大学　大学院薬学研究科, 3. 姫路獨協大学　薬学部　薬理学 Kensuke Motomura¹, Boateng Alex², Masaharu Sugiura², Yuki Kurauchi¹, Hiroshi Katsuki¹, Takahiro Seki³ 1. Department of Chemico-Pharmacological Sciences, Graduate School of Pharmaceutical Sciences, Kumamoto Univercity, Kumamoto, Japan Autophagy-lysosome proteolysis regulates protein homeostasis in neurons and is classified into macroautophagy, microautophagy (mA), and chaperone-mediated autophagy (CMA). Among them, we focused on CMA and mA and established a novel method to monitor CMA/mA activity. Recently, we identified an analog (A2) from aromatic (ar)-turmerone that protects dopaminergic neurons via the activation of an antioxidant transcription factor, Nrf2, which is known to activate CMA/mA. In this study, we attempted to identify novel ar-turmerone analogs that can activate Nrf2 more efficiently and activate and CMA/mA. We synthesized four novel ar-turmerone analogues (A4-A7) and investigated the ability to activate Nrf2 and CMA/mA in SH-SY5Y cells. Immunoblot experiments revealed that all compounds significantly upregulated Nrf2 after 6 h treatment. In contrast, only A4 significantly activated CMA/mA after 24 h treatment. To investigate why these analogs differently affected CMA/mA activity, we focused on p38 that is reported to regulate CMA via the phosphorylation of LAMP2A. Although A5-A7 transiently increased the phosphorylation of p38 after 6 h treatment, A4 persistently increased the phosphorylation of p38 until 24 h after the treatment. We identified a novel ar-turmerone analog (A4) that activate CMA/mA via the upregulation of Nrf2 and sustained activation of p38.

7月7日（金）　13:50-14:50　ポスター会場① 2P①-5 細胞外マトリクスにより形成される微小環境は大脳皮質発生に重要である Microenvironment formed by the extracellular matrix is important for cortical development 武渕　明裕夢¹, 武智　美奈², 佐藤　ちひろ³, 北島　健³, 北川　裕之⁴, 宮田　真路¹ 1. 東京農工大学大学院連合農学研究科, 2. 名古屋大学大学院生命農学研究科, 3. 名古屋大学糖鎖生命コア研究所, 4. 神戸薬科大学生化学研究室 Ayumu Mubuchi¹, Mina Takechi², Chihiro Sato³, Ken Kitajima³, Hiroshi Kitagawa⁴, Shinji Miyata¹ 1. United Graduate School of Agricultural Science, Univ. of TUAT, Tokyo, Japan, 2. Grad Sch Bioagr Sci. Nagoya Univ, Nagoya, Japan,, 3. iGCORE, Nagoya Univ, Nagoya, Japan,, 4. Lab Biochem, Kobe Pharma Univ, Kobe, Japan The cerebral cortex, which controls higher brain functions such as cognition and memory, is formed during development as neurons differentiated from neural stem cells migrate toward the brain surface. Newly born neurons migrate in a complex three-dimensional environment generated by the extracellular matrix (ECM). The intermediate zone has a low cell density and abundant ECM. We previously reported that the intermediate zone is rich in hyaluronic acid (HA), a large linear polysaccharide. However, the physiological role of the ECM in cortical formation is not well understood. Neurocan (NCAN) is a major chondroitin sulfate proteoglycan highly expressed in developing cortex and a risk factor for neuropsychiatric disorders. We found that the N-terminal region of NCAN binds to HA and the C-terminal region to tenascin-C (TNC), forming a ternary complex of NCAN, TNC, and HA in the intermediate zone. To elucidate the function of this ternary complex, we generated double-knockout (DKO) mice deficient in both NCAN and TNC. Compared to wild-type mice, DKO mice showed delayed neuronal migration by suppressing the multipolar-to-bipolar transition. Furthermore, TNC in the ternary complex promotes neuronal morphological maturation. Our results suggest that the microenvironment formed by the ECM molecules is important for cortical development.		7月7日（金）　13:50-14:50　ポスター会場① 2P①-6 小頭症・セッケル症候群の原因遺伝子 Polo-like kinase 4 (Plk4)のマウス発達脳における発現解析 Expression analyses of Plk4, a responsible gene for microcephaly and Seckel syndrome, during mouse brain development 浜田　奈々子, 永田　浩一愛知県医療療育総合センター　発達障害研究所　分子病態研究部 Nanako Hamada, Koh-ichi Nagata Inst. for Developmental Res., Aichi Developmental Disability Center Plk4 (polo-like kinase 4) is a Ser/Thr-kinase which plays a central role in centriole duplication during the cell cycle. PLK4 gene abnormalities are responsible for autosomal recessive chorioretinopathy-microcephaly syndrome and Seckel syndrome. In this study, we performed expression analyses of Plk4 by focusing on mouse brain development. As to the central nervous sytem, Plk4 was expressed throughout the developmental process with drastic increase after P15, suggesting an essential role of Plk4 in differentiated neurons. In immunohistochemical analyses with mouse brain at embryonic day 14, Plk4 was detected dominantly at the cell-cell contact sites of neuronal progenitors in the ventricular zone. Plk4 was then diffusely distributed in the cell body of cortical neurons at P7 while it was enriched in the neuropil as well as soma of excitatory neurons in cerebral cortex and hippocampus at P30. Notably, biochemical fractionation analysis found an enrichment of Plk4 in the post-synaptic density fraction. Then, immunofluorescent analyses showed partial co-localization of Plk4 with excitatory synaptic markers, PSD-95 and synaptophysin, in differentiated primary cultured hippocampal neurons. These results suggest that Plk4 takes part in the regulation of synaptic function in differentiated neurons.