Oral Session

Oral Session 19 一般口演19

O19-1 n vivo phosphorylation of tau in brains of Marmoset, a non-human Primate マーモセット脳におけるタウのアイソフォーム発現とリン酸化：アルツハイマー病霊長類モデルとなりうるか Hisanaga Shin-Ichi（久永眞市）¹，塩澤　誠司²，小林玲央奈²，村山繁雄³，斎藤太郎¹，安藤香奈絵¹，岡野栄之²，木村妙子¹，Sharma Govinda¹ ¹Dept. of Biosci. Sci. Tokyo Metro. Univ. ²Keio University, Tokyo, Japan ³Tokyo Inst. Gerontology, Tokyo, Japan Tau is hyperphosphorylated in patient’s brains of Alzheimer’s disease. However, the cause of tau hyperphsophorylation is not known yet. There are unavoidable limitations in analysis of human brains’ tau, particularly dephosphorylation during postmortem interval is problematic. On the other hand, there are no proper rodent models reflecting tau pathology exactly. Marmoset, a new world monkey, is expected to be a primate model of neurodegenerative diseases. While tau hyperphosphorylation is reported histochemically, there is no biochemical report yet. In this study, we cloned tau cDNA from an adult marmoset brain and analyzed expression of tau isoforms and their phosphorylation. The longest isoform (2N4R) of marmoset tau showed 96.3 % identity to the corresponding human tau isoform. Interestingly, unlike tau from other primates, marmoset tau lacks 10 amino acids at the N-terminal region similar to rodent tau. Two isoforms, 0N4R and 2N4R, of tau were expressed in adult, whereas 0N3R was predominantly expressed with a small amount of 1N3R isoform in newborn marmoset. Marmoset tau displayed a slightly different banding pattern from those of human and mouse tau when they were expressed in Neuro2a cells and analyzed by Phos-tag (phospho-affinity) SDS-PAGE. Phos-tag SDS-PAGE analysis showed that tau is highly phosphorylated in marmoset brains, compared to those in autopsy samples of aged human brains. However, the phosphorylation at several pathological sites such as AT8, AT180 and PHF1 sites were insignificant in adult marmoset brains. These results provide valuable biochemical information of tau for using marmoset as the non-human primate model of tauopathies.		O19-2 Neuronal functions of ER stress sensor BBF2H7-derived small peptide fragments in neurotoxicity and fibrilization of amyloid β 小胞体ストレスセンサーBBF2H7由来小ペプチドの神経毒性とアミロイド線維形成促進作用 Matsuhisa Koji（松久幸司）¹，齋藤敦¹，蔡龍傑²，柳田寛太³，大河内正康³，工藤幸司⁴，松本雅記^5,6，中山敬一^5,6，今泉和則² ¹Department of Stress Protein Processing, Institute of Biomedical & Health Sciences, Hiroshima University, Hiroshima, Japan ²Department of Biochemistry, Institute of Biomedical & Health Sciences, Hiroshima University, Hiroshima, Japan ³Psychiatry, Department of Integrated Medicine, Division of Internal Medicine, Osaka University Graduate School of Medicine, Osaka, Japan ⁴Department of Gerontology and Geriatrics, Institute of Development, Aging and Cancer, Tohoku University, Sendai, Japan ⁵Division of Proteomics, Medical Institute of Bioregulation, Kyushu University, Fukuoka, Japan ⁶Department of Molecular and Cellular Biology, Medical Institute of Bioregulation, Kyushu University, Fukuoka, Japan The endoplasmic reticulum (ER) stress sensor BBF2H7 is a transmembrane transcription factor, and is received sequential cleavage in its transmembrane and luminal domains in response to ER stress. We recently found that approximately 40-50 amino acid polypeptides [BBF2H7-derived small peptide (BSP) fragments] located between the cleavage sites of BBF2H7 are generated in an ER stress-dependent manner. In this study, we examined biological characteristics of BSP fragments. The major type of BSP fragments was composed of 45 amino acid including partial transmembrane and luminal regions. The synthetic BSP fragments based on the identified sequence showed highly hydrophobic characteristics. The 100μM synthetic peptides were incubated in PBS at 37°C for 12, 24, 48 and 72 h to analyze its morphology and aggregable property by transmission electron microscope. The fibrilization was observed on 12 h and the fibrils with width of approximately 10 nm elongated until 72 h. These fibrils formed large bundle structures. Next, we treated SK-N-SH cells with the 5, 10 and 20 μM synthetic peptides for 48 h. The cells shrank and its number was decreased in a dose-dependent manner, indicating that BSP fragments exhibited neurotoxicity. Interestingly, the addition of 0.15 μM BSP fragments to 5 μM Amyloidβ_1-40(Aβ_1-40) promoted formation of fibrils, although 0.15 μM BSP fragments or 5 μM Aβ_1-40 alone did not form fibrils in these conditions. In conclusion, BSP fragments generated from BBF2H7 in response to ER stress have a potential to injure neuronal cells and accelerate fibrilization of Aβ_1-40. Our findings suggest that BSP fragments may link ER stress to the pathogenesis of ER stress-related neurodegenerative disorders.

O19-3 Genetic silencing of the mitochondrial protein p13 protects against experimental parkinsonism ミトコンドリア蛋白p13の遺伝子サイレンシングは実験的パーキンソン病モデルに保護的に働く Shintani Norihito（新谷紀人）¹，小椋紗恵¹，井上直紀¹，師田洋平¹，植野寛貴¹，橋本均^1,2,3 ¹Lab. of Mol. Neuropharmacol., Grad. Sch. of Pharmaceut. Sci., Osaka Univ. ²Mol. Res. Cent. for Children's Mental Dev., United Grad. Sch. Child Dev., Osaka Univ. ³Instit. for Datability Sci., Osaka Univ. We recently identified a novel 13-kDa protein (p13) that may be involved in mitochondrial function, however, its pathophysiological roles have been remained poorly understood. Here, we investigated the mitochondrial function of p13 and its involvement in the pathogenesis of Parkinson's disease (PD) using mitochondrial toxin-induced PD models. Our data show that p13 overexpression induces mitochondrial dysfunction and apoptosis and its knockdown attenuates toxin-induced mitochondrial dysfunction and apoptosis in dopaminergic SH-SY5Y cells. Importantly, heterozygous p13-deficient mice obtained by using the CRISPR/Cas9 system exhibit tolerances against toxin-induced motor deficits and the loss of dopaminergic neurons in the substantia nigra. Accordingly, these results firstly revealed the neuronal p13 function, and suggest that manipulating p13 expression might be a promising avenue for therapeutic intervention in PD.