研究者詳細 - 佐藤　ちひろ

Ganglioside contained in the neuronal tissue-enriched acidic protein of 22 kDa (NAP-22) fraction prepared from the detergent-resistant membrane microdomain of rat brain inhibits the phosphatase activity of calcineurin.

Kobayashi Y, da Silva R, Kumanogoh H, Miyata S, Sato C, Kitajima K, Nakamura S, Morita M, Hayashi F, Maekawa S

Journal of neuroscience research 93 巻 ( 9 ) 頁： 1462 - 70 2015年9月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：Journal of Neuroscience Research

Neurons have well-developed membrane microdomains called "rafts" that are recovered as a detergent-resistant membrane microdomain fraction (DRM). Neuronal tissue-enriched acidic protein of 22 kDa (NAP-22) is one of the major protein components of neuronal DRM. To determine the cellular function of NAP-22, interacting proteins were screened with an immunoprecipitation assay, and calcineurin (CaN) was detected. Further studies with NAP-22 prepared from DRM and CaN expressed in bacteria showed the binding of these proteins and a dose-dependent inhibitory effect of the NAP-22 fraction on the phosphatase activity of CaN. On the other hand, NAP-22 expressed in bacteria showed low binding to CaN and a weak inhibitory effect on phosphatase activity. To solve this discrepancy, identification of a nonprotein component that modulates CaN activity in the DRM-derived NAP-22 fraction was attempted. After lyophilization, a lipid fraction was extracted with chloroform/methanol. The lipid fraction showed an inhibitory effect on CaN without NAP-22, and further fractionation of the extract with thin-layer chromatography showed the presence of several lipid bands having an inhibitory effect on CaN. The mobility of these bands coincided with that of authentic ganglioside (GM1a, GD1a, GD1b, and GT1b), and authentic ganglioside showed an inhibitory effect on CaN. Treatment of lipid with endoglycoceramidase, which degrades ganglioside to glycochain and ceramide, caused a diminution of the inhibitory effect. These results show that DRM-derived NAP-22 binds several lipids, including ganglioside, and that ganglioside inhibits the phosphatase activity of CaN.

DOI： 10.1002/jnr.23599

Rapid Trimming of Cell Surface Polysialic Acid (PolySia) by Exovesicular Sialidase Triggers Release of Preexisting Surface Neurotrophin. Open Access

Sumida M, Hane M, Yabe U, Shimoda Y, Pearce OM, Kiso M, Miyagi T, Sawada M, Varki A, Kitajima K, Sato C

The Journal of biological chemistry 290 巻 ( 21 ) 頁： 13202 - 14 2015年5月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：Journal of Biological Chemistry

As acidic glycocalyx on primary mouse microglial cells and a mouse microglial cell line Ra2, expression of polysialic acid (polySia/PSA), a polymer of the sialic acid Neu5Ac (N-acetylneuraminic acid), was demonstrated. PolySia is known to modulate cell adhesion, migration, and localization of neurotrophins mainly on neural cells. PolySia on Ra2 cells disappeared very rapidly after an inflammatory stimulus. Results of knockdown and inhibitor studies indicated that rapid surface clearance of polySia was achieved by secretion of endogenous sialidase Neu1 as an exovesicular component. Neu1-mediated polySia turnover was accompanied by the release of brain-derived neurotrophic factor normally retained by polySia molecules. Introduction of a single oxygen atom change into polySia by exogenous feeding of the non-neural sialic acid Neu5Gc (N-glycolylneuraminic acid) caused resistance to Neu1-induced polySia turnover and also inhibited the associated release of brain-derived neurotrophic factor. These results indicate the importance of rapid turnover of the poly-Sia glycocalyx by exovesicular sialidases in neurotrophin regulation.

DOI： 10.1074/jbc.M115.638759

Discovery, primary, and crystal structures and capacitation-related properties of a prostate-derived heparin-binding protein WGA16 from boar sperm.

Sulfatide-Hsp70 interaction promotes Hsp70 clustering and stabilizes binding to unfolded protein. Open Access

Harada Y, Sato C, Kitajima K

Biomolecules 5 巻 ( 2 ) 頁： 958 - 73 2015年5月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：Biomolecules

The 70-kDa heat shock protein (Hsp70), one of the major stress-inducible molecular chaperones, is localized not only in the cytosol, but also in extracellular milieu in mammals. Hsp70 interacts with various cell surface glycolipids including sulfatide (3'-sulfogalactosphingolipid). However, the molecular mechanism, as well as the biological relevance, underlying the glycolipid-Hsp70 interaction is unknown. Here we report that sulfatide promotes Hsp70 oligomerization through the N-terminal ATPase domain, which stabilizes the binding of Hsp70 to unfolded protein in vitro. We find that the Hsp70 oligomer has apparent molecular masses ranging from 440 kDa to greater than 669 kDa. The C-terminal peptide-binding domain is dispensable for the sulfatide-induced oligomer formation. The oligomer formation is impaired in the presence of ATP, while the Hsp70 oligomer, once formed, is unable to bind to ATP. These results suggest that sulfatide locks Hsp70 in a high-affinity state to unfolded proteins by clustering the peptide-binding domain and blocking the binding to ATP that induces the dissociation of Hsp70 from protein substrates.

DOI： 10.3390/biom5020958

Garénaux E, Kanagawa M, Tsuchiyama T, Hori K, Kanazawa T, Goshima A, Chiba M, Yasue H, Ikeda A, Yamaguchi Y, Sato C, Kitajima K

The Journal of biological chemistry 290 巻 ( 9 ) 頁： 5484 - 501 2015年2月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：Journal of Biological Chemistry

Mammalian sperm acquire fertility through a functional maturation process called capacitation, where sperm membrane molecules are drastically remodeled. In this study, we found that a wheat germ agglutinin (WGA)-reactive protein on lipid rafts, named WGA16, is removed from the sperm surface on capacitation. WGA16 is a prostate-derived seminal plasma protein that has never been reported and is deposited on the sperm surface in themalereproductive tract. BasedonproteinandcDNAsequences for purified WGA16, it is a homologue of human zymogen granule protein16(ZG16) belonging to the Jacalin-related lectin (JRL) family in crystal and primary structures. A glycan array shows that WGA16 binds heparin through a basic patch containing Lys-53/ Lys-73 residues but not the conventional lectin domain of the JRL family. WGA16 is glycosylated, contrary to other ZG16 members, and comparative mass spectrometry clearly shows its unique N-glycosylation profile among seminal plasma proteins. It has exposed GlcNAc and GalNAc residues without additional Gal residues. The GlcNAc/GalNAc residues can work as binding ligands foraspermsurface galactosyltransferase,whichactually galactosylatesWGA16in situ in the presence of UDP-Gal. Interestingly, surface removal of WGA16 is experimentally induced by either UDPGal or heparin. In the crystal structure, N-glycosylated sites and a potential heparin-binding site face opposite sides. This geography of two functional sites suggest that WGA16 is deposited on the sperm surface through interaction between its N-glycans and the surface galactosyltransferase, whereas its heparin-binding domain may be involvedinbindingto sulfatedglycosaminoglycansinthefemaletract, enabling removal ofWGA16from the sperm surface.

DOI： 10.1074/jbc.M114.635268

Polysialic acid: Biosynthesis, novel functions and applications

Adaptive evolution of the promoter region of the Sialyltransferase 8B (STX) gene

Fujito Naoko T., Hayakawa Toshiyuki, Hane Masaya, Kitajima Ken, Sato Chihiro, Satta Yoko

GENES & GENETIC SYSTEMS 89 巻 ( 6 ) 頁： 327 - 327 2014年12月

An anti-schizophrenic drug affects the surface expression of polySia-NCAM in IMR-32 human neuroblastoma cells

Nishimura Saki, Hane Masaya, Niimi Yuki, Kitajima Ken, Sato Chihiro

GLYCOBIOLOGY 24 巻 ( 11 ) 頁： 1193 - 1193 2014年11月

Insight into single nucleotide polymorphisms (SNPs) of the polysialyltransferase ST8SIA2/STX in psychiatric disorders

Hane Masaya, Nishimura Saki, Hayakawa Toshiyuki, Kitajima Ken, Sato Chihiro

GLYCOBIOLOGY 24 巻 ( 11 ) 頁： 1206 - 1206 2014年11月

Colley, KJ; Kitajima, K; Sato, C

CRITICAL REVIEWS IN BIOCHEMISTRY AND MOLECULAR BIOLOGY 49 巻 ( 6 ) 頁： 498 - 532 2014年11月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：Critical Reviews in Biochemistry and Molecular Biology

As an anti-adhesive, a reservoir for key biological molecules, and a modulator of signaling, polysialic acid (polySia) is critical for nervous system development and maintenance, promotes cancer metastasis, tissue regeneration and repair, and is implicated in psychiatric diseases. In this review, we focus on the biosynthesis and functions of mammalian polySia, and the use of polySia in therapeutic applications. PolySia modifies a small subset of mammalian glycoproteins, with the neural cell adhesion molecule, NCAM, serving as its major carrier. Studies show that mammalian polysialyltransferases employ a unique recognition mechanism to limit the addition of polySia to a select group of proteins. PolySia has long been considered an anti-adhesive molecule, and its impact on cell adhesion and signaling attributed directly to this property. However, recent studies have shown that polySia specifically binds neurotrophins, growth factors, and neurotransmitters and that this binding depends on chain length. This work highlights the importance of considering polySia quality and quantity, and not simply its presence or absence, as its various roles are explored. The capsular polySia of neuroinvasive bacteria allows these organisms to evade the host immune response. While this "stealth" characteristic has made meningitis vaccine development difficult, it has also made polySia a worthy replacement for polyetheylene glycol in the generation of therapeutic proteins with low immunogenicity and improved circulating half-lives. Bacterial polysialyltransferases are more promiscuous than the protein-specific mammalian enzymes, and new studies suggest that these enzymes have tremendous therapeutic potential, especially for strategies aimed at neural regeneration and tissue repair.

DOI： 10.3109/10409238.2014.976606