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�Research projects and annual reports We have been investigating roles of O-linked sugar chains with the linkage structures, GalNAcα1→Ser(Thr), Manα1→Ser(Thr), or GlcNAcβ1→Ser(Thr). O-GalNAc-type sugar chains are called mucin-type carbohydrates and their biosynthesis is initiated by a group of enzymes, UDP-GalNAc: polypeptide N-acetyl- galactosaminyltransferases (GalNAc-Ts). GalNAc-Ts consist of a large gene family with 20 isozymes in humans. Interestingly, GalNAc-T8, -T9, -T17, and

-T18 are catalytically inactive, while GalNAc-T9 and -T17, which were identified by us, are brain-specific and biologically important for the neural differentiation. Based on these backgrounds, we have focused on the functions of glycosyltransferases to make O-glycan carbohydrate-linkage structures, and obtained the following findings. 1) Analysis of mice defective of a GalNAc-T17 gene We found that GalNAc-T17 KO mice showed decreased body weight compared with wild-type mice (Fig. 1). Analysis of GalNAc-T17 expression demonstrated that it was expressed not only in hippocampus and amygdala, but in ventromedial nucleus of hypothalamus, which is known as satiety center. GalNAc-T17 may therefore positively regulate feeding behavior. 2) Production of zebrafish GalNAc-T mutants To produce zebrafish mutants defective of GalNAc-T genes, we employed recently developed genome-editing technology, TALEN and CRISPR/Cas systems. We found that mutations were successfully introduced at the target sites in the zebrafish embryos using TALEN and CRISPR/Cas vectors for GalNAc-T9 and -T18. 3) A rapid and efficient method for neuronal induction of P19 embryonic carcinoma cell line We revised a novel method to efficiently differentiate P19 cells into neurons by an adherent culture in laminin-coated dish in the presence of several reagents to induce neural differentiation and suppress growth of non-neural cells. The new method induced neurons within 4 days without gliogenesis. The mature neurons obtained were responsive to several neurotransmitters with functional neuronal networks. 4) Analysis of extracellular O-glycosylation We investigated the function of a glycosyltransferase in zebrafish, which catalyzes extracellular novel O-glycosylation. We found that it was strongly expressed in the brain of zebrafish embryos, and its suppression caused alterations in the brain development. 96��5��32!

Y. Nakayama, A. Wada, R. Inoue, K. Terasawa, I. Kimura, N.

Nakamura, A. Kurosaka: A Rapid and Efficient Method for

Neuronal Induction of the P19 Embryonic Carcinoma Cell Line.

Journal of Neuroscience Methods, in press.

M. Ogawa, N. Nakamura, Y. Nakayama, A. Kurosaka, H. Manya,

M. Kanagawa, T. Endo, K. Furukawa, T. Okajima: GTDC2

modifies O-mannosylated ŷ-dystroglycan in the endoplasmic

reticulum to generate N-acetyl-glucosamine epitopes reactive

with CTD110.6 antibody. (2013) Biochem Biophys Res

Commun 440(1), 88-93

I. Kimura, Y. Nakayama, Y. Zhao, M. Konishi, N. Itoh:

Neurotrophic effects of neudesine in the central nervous system.

(2013) Front Neurosci 7 (article111), 1-5

Y. Nakayama, N. Nakamura, D. Tsuji, K. Itoh, A. Kurosaka:

Genetic diseases associated with protein glycosylation

disorders in mammals. pp. 244-269 in Genetic Disorders,

Maria Puiu (Ed.), ISBN: 978-953-51-08866-3, InTech.

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Õ36?�� T¸²V�d�ǪÎybǪ2013.12.3-6

Y. Nakayama, K. Kato, N. Nakamura, A. Kurosaka: The roles

of a putative polypeptide GalNAc-transferase/Wbscr17.

International Symposium on Glyco-Neuroscience, Awaji city

(Japan) 2014.1.9-11

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