||Watanabe T, Sakiyama R, Iimi Y, Sekine S, Abe E, Nomura H, Ishibashi Y, Okino N, Ito M, Regulation of TG accumulation and lipid droplet morphology by the novel TLDP1 in Aurantiochytrium limacinum F26-b, Journal Lipid Research, 2017.10.
||Nozomu Okino, Makoto Ito, Molecular mechanism for sphingosine-induced Pseudomonas ceramidase expression through the transcriptional regulator SphR, 10.1038/screp38797, 2016.11.
||Takashi Watanabe, Motohiro Tani, Yohei Ishibashi, Ikumi Endo, NOZOMU OKINO, Makoto Ito, Ergosteryl-β-glucosidase (Egh1) involved in sterylglucoside catabolism and vacuole formation in Saccharomyces cerevisiae.
, Glycobiology, 25, 10, 1079-1089, 2015.04.
||Takashi Watanabe, Tomohiro Ito, Hatsumi Goda, Yohei Ishibashi, NOZOMU OKINO, Makoto Ito, Sterylglucoside catabolism in Cryptococcus neoformans with endoglycocoramidase-related protein2(EGCrP2), the first steryl-β-glucosidase identifed in fungi.
, Journal of Biological Chemistry, 290, 1005-1019, 2015.01, Cryptococcosis is an infectious disease caused by pathogenic fungi such as Cryptococcus neoformans and C. gattii. The ceramide structure (methyl-d18:2/h18:0) of C. neoformans glucosylceramide (GlcCer) is characteristic and strongly related to their pathogenicity. Recently, we reported that endoglycoceramidase-related protein 1 (EGCrP1) is a glucocerebrosidase in C. neoformans and involved in the quality control of GlcCer by eliminating immature GlcCer during the synthesis of GlcCer (Ishibashi et al, J. Biol. Chem., 2012). We report here the identification and characterization of EGCrP2, a homologue of EGCrP1, as an enzyme responsible for sterylglucoside catabolism in C. neoformans. In contrast to EGCrP1 specific to GlcCer, EGCrP2 was found to hydrolyze a variety of β-glucosides including cholesteryl-β-glucoside, ergosteryl-β-glucoside, sitosteryl-β-glucoside, GlcCer, and para-nitrophenyl-β-glucoside, but not α-glucosides or β-galactosides, under the acidic condition. Disruption of the EGCrP2 gene (egcrp2) resulted in the accumulation of glycolipid, and the structure was determined following purification to ergosteryl-3-β-glucoside by mass spectrometric and two-dimensional nuclear magnetic resonance analyses. This glycolipid was found to accumulate in vacuole where EGCrP2 is localized. These results indicated that EGCrP2 is involved in the catabolism of ergosteryl-β-glucoside in the vacuole of C. neoformans. The egcrp2-disrupted mutants showed distinct growth arrest, dysfunction of cell budding, and an abnormal vacuole morphology, suggesting that EGCrP2 is a promising target for anti-cryptococcal drugs. EGCrP2, classified into glycohydrolase family 5, is the first identified steryl-β-glucosidase and a missing link in steryl glucoside metabolism in fungi. .
||Yohei Ishibashi, NOZOMU OKINO, Makoto Ito, A novel ether-linked phytol-containing digalactosylglycerolipid in the marine green alga, Ulva pertusa., Biochem Biophys Res Commun, 10.1016/j.bbrc.2014.08.056., 452, 873-80, 2014.10.
||Eriko Abe, Makoto Ito, Novel Lysophospholipid Acyltransferase PLAT1 of Aurantiochytrium limacinum F26-b Responsible for Generation of Palmitate-Docosahexaenoate-Phosphatidylcholine and Phosphatidylethanolamine, PLOS ONE, 10.1371/journal.pone.0102377, 9, 8, 2014.08.
||Makoto Ito, NOZOMU OKINO, Motohiro Tani, New insights into the structure, reaction mechanism, and biological functions of neutral ceramidase, Biochimica ET BIOPHYSICA ACTA-MOLECULAR AND CELL BIOLOGY OF LIPIDS, 1841, 5, 682-691, 2014.05.
||Junichiro Ohara, Keishi Sakaguchi, NOZOMU OKINO, Makoto Ito, Two fatty acid elongates possessing C18-Δ6/C18-Δ9/C20-Δ5 or C16-Δ9 Elongase activity in Thraustochytrium sp. ATCC 26185, Marine Biotechnology, 10.1007/s10126-013-9496-1, 2013.04.
||Chisada, Shin-ichi, Shimizu, Kohei, Kamada, Haruna, Matsunaga, Naoyuk, NOZOMU OKINO, Makoto Ito, Vibrios adhere to epithelial cells in the intestinal tract of red sea bream, Pagrus major, utilizing GM4 as an attachment site, FEMS MICROBIOLOGY LETTERS, 10.1111/1574-6968.12082, 341, 1, 18-26, 2013.04.
||Ishibashi, Yohei, Ikeda, Kazutaka, Sakaguchi, Keishi, NOZOMU OKINO, Taguchi, Ryo, Makoto Ito, Quality Control of Fungus-specific Glucosylceramide in Cryptococcus neoformans by Endoglycoceramidase-related Protein 1 (EGCrP1), JOURNAL OF BIOLOGICAL CHEMISTRY, 10.1074/jbc.M111.311340, 287, 1, 368-381, 2012.01.
||Sakaguchi, Keishi, Matsuda, Takanori;, Kobayashi, Takumi, Ohara, Junichiro, Hamaguchi, Rie, Abe, Eriko, Hayashi, Masahiro, Honda, Daiske, NOZOMU OKINO, Makoto Ito, Versatile Transformation System That Is Applicable to both Multiple Transgene Expression and Gene Targeting for Thraustochytrids, APPLIED AND ENVIRONMENTAL MICROBIOLOGY, 10.1128/AEM.07129-11, 78, 9, 3193-3202, 2012.05.
||Takanori Matsuda, Hamaguchi, Rie, Kobayashi, Takumi, Abe, Eriko, Hama, Yoichiro, Hayashi, Masahiro, Honda, Daiske, NOZOMU OKINO, Makoto Ito, Analysis of Delta 12-fatty acid desaturase function revealed that two distinct pathways are active for the synthesis of PUFAs in T. aureum ATCC 34304, JOURNAL OF LIPID RESEARCH, 10.1194/jlr.M024935, 53, 6, 1210-1222, 2012.06.
||Ishibashi, Yohei;, Kobayashi, Utaro, Hijikata, Atsushi, Sakaguchi, Keish, 合田 初美, NOZOMU OKINO, Makoto Ito, Preparation and characterization of EGCase I, applicable to the comprehensive analysis of GSLs, using a rhodococcal expression system, JOURNAL OF LIPID RESEARCH, 10.1194/jlr.D028951, 53, 10, 2242-2251, 2012.10.
||Kobayashi T, Sakaguchi K, Matsuda T, Abe E, Hama Y, Hayashi M, Honda D, Okita Y, Sugimoto S, Okino N, Ito M.
, Increase of EPA in Thraustochytrids through Expression of a Fatty Acid Δ5 Desaturase Gene Driven by the Thraustochytrid Ubiquitin Promoter.
, Appl. Environ. Microbiol. , 77, 11, 3870-3876, 2011.11.
||Sumida T, Fujimoto K, Ito M, Molecular Cloning and Catalytic Mechanism of a Novel Glycosphingolipid-degrading β-N-Acetylgalactosaminidase from Paenibacillus sp. TS12.
J. Biol. Chem. , 286, 16, 14065-14072, 2011.04.
||Takara T, Nakagawa T, Isobe M, Okino N, Ichinose S, Omori A, Ito M., Purification, molecular cloning, and application of a novel sphingomyelin-binding protein (clamlysin) from the brackishwater clam, Corbicula japonica.
, Biochim. Biophys. Acta. , 1811, 5, 323-332, 2011.05.
||Togayachi A, Kozono Y, Ikehara Y, Ito H, Suzuki N, Tsunoda Y, Abe S, Sato T, Nakamura K, Suzuki M, Goda HM, Ito M, Kudo T, Takahashi S, Narimatsu H.
, Lack of lacto/neolacto-glycolipids enhances the formation of glycolipid-enriched microdomains, facilitating B cell activation.
, Proc. Natl. Acad. Sci. USA, 107, 26, 11900-11905 , 2010.04, In a previous study, we demonstrated that β1,3-N-acetylglucosaminyltransferase
5 (B3gnt5) is a lactotriaosylceramide (Lc3Cer) synthase
that synthesizes a precursor structure for lacto/neolacto-series glycosphingolipids
(GSLs) in in vitro experiments. Here, we generated
B3gnt5-deficient (B3gnt5−/−) mice to investigate the in vivo biological
functions of lacto/neolacto-series GSLs. In biochemical analyses,
lacto/neolacto-series GSLs were confirmed to be absent and no
Lc3Cer synthase activity was detected in the tissues of these mice.
These results demonstrate that β3GnT5 is the sole enzyme synthesizing
Lc3Cer in vivo. Ganglioside GM1, known as a glycosphingolipid-
enriched microdomain (GEM) marker, was found to be upregulated
in B3gnt5−/− B cells by flow cytometry and fluorescence
microscopy. However, no difference in the amount of GM1 was observed
by TLC-immunoblotting analysis. The GEM-stained puncta on
the surface of B3gnt5−/− resting B cells were brighter and larger
than those of WT cells. These results suggest that structural alteration
of GEM occurs in B3gnt5−/− B cells.We next examinedwhether
BCR signaling-related proteins, such as BCR, CD19, and the signaling
molecule Lyn, had moved into or out of the GEM fraction. In
B3gnt5−/− B cells, thesemolecules were enriched in the GEMfraction
or adjacent fraction. Moreover, B3gnt5−/− B cells were more sensitive
to the induction of intracellular phosphorylation signals on BCR
stimulation and proliferated more vigorously than WT B cells. Together,
these results suggest that lacto/neolacto-series GSLs play
an important role in clustering of GEMs and tether-specific proteins,
such as BCR, CD19, and related signaling molecules to the GEMs..
||Nishie T, Hikimochi Y, Zama K., Fukusumi Y, Ito M, Yokoyama H, Baruse C, Ito
M, Asano M.
, β-Galactosyltransferase-5 is a lactosylceramide synthase essential for
mouse extra-embryonic development.
, Glycobiology, 20, 10, 1311-1322, 2010.05.
||Inoue T, Okino N, Kakuta Y, Hijikata A, Okano H, Goda HM, Tani M, Sueyoshi N, Kambayashi K, Matsumura H, Kai Y, Ito M., Mechanistic insights into the hydrolysis and synthesis of ceramide by neutral ceramidase., J Biol Chem. , 284, 14, 9566-9577, 2009.04.
||Ishibashi Y, Nagamatsu Y, Meyer S, Imamura A, Ishida H, Kiso M, Okino N, Geyer R, Ito M., Transglycosylation-based Fluorescent Labeling of 6-Gala Series Glycolipids by EGALC, Glycobiology , 19, 7, 797-807, 2009.04.
||Zama K, Hayashi Y, Ito Y, Hirabayashi Y, Inoue T, Ohno K, Okino N, Ito M., Simultaneous quantification of glucosylceramide and galactosylceramide by normal-phase HPLC using O-phtalaldehyde derivatives prepared with sphingolipid ceramide N-deacylase, Glycobiology , 19, 7, 767-775, 2009.05.
||Kiyohara M, Sakaguchi K, Yamaguchi K, Araki T, and Ito M., Characterization and application of carbohydrate-binding modules of b-1,3-xylanase XUL4., J Biochem, 146, 5, 633-641, 2009.07.
||Chisada S, Yoshimura Y, Sakaguchi K, Uchima H, Matsunaga N, Okino N, Uemura S, Go S, Ogura K, Tai T, Ikeda K, Taguchi R, Inokuchi J, and Ito M., Zebrafish and mouse a2,3-sialyltransferases responsible for synthesizing GM4 ganglioside., J Biol Chem., 284, 44, 30534-30546, doi:10.1074/jbc.M109.016188, 2009.10.
||Sumida T., Ishii R, Yanagisawa T., Yokoyama S., and Ito M., Molecular cloning and crystal structural analysis of a novel b-N-acetylhexosaminidase from Paenibacillus sp. TS12 capable of degrading glycosphingoliids., J Mol Biol , 392, 87-99, 2009.04.
||Xu X, Horibata Y, Inagaki M, Hama Y, Sakaguchi K, Okino N, and Ito M., A novel fucosyl glycosphingolipid of brine shrimp that is highly sensitive to endoglycoceramidase., Glycobiology , 19, 1446-1451, 2009.10.
||Y. Hayashi, K. Zama, E. Abe, N. Okino, T. Inoue, K. Ohono, and M. Ito, A sensitive and reproducible fluorescent-based HPLC assay to measure the activity of acid as well as neutral b-glucocerebraosidases., Analytical Biochemistry, 383, 122-129, 2008.05.
||H. M. Goda, K. Ushigusa, N. Okino, H. Narimatsu, and M. Ito, Molecular cloning, expression and characterization of a novel endo-a-N-acetylgalactosaminidase from Enterococcus faecalis., Biochemistry Biophysics Research Communication, 375, 541-546, 2008.05.
||Y. Ishibashi, T. Nakasone, M. Kiyohara, K. Sakaguchi, A. Hijikata, N. Okino, M. Ito, A novel endogalactosylceramidase hydrolyzes oligogalactosylceramides to produce galactooligosaccharides and ceramides., Journal of Biological Chemistry, 282, 11386-11389, 2007.05.
||Y. Hayashi, N. Okino, Y. Kakuta, T. Shikanai, H. Narimatsu, M. Ito, Klotho-related protein is a novel cytosolic neutral b-glycosylceramidase., Journal of Biological Chemistry, 282, 30889-30900, 2007.10.
||Eriko Abe, Yasuhiro Hayashi, Yoichiro Hama, Masahiro Hayashi, Masanori Inagaki, and Makoto Ito, A novel phosphatidylcholine which contains pentadecanoic acid at sn-1 and docosahexaenoic acid at sn-2 in Shizochytrium sp. F26-b., Journal of Biochemistry, 140, 247-253, 2006.01.
||Motohiro Tani, Yasuyuki Igarashi and Makoto Ito, Involvement of neutral ceramidase in ceramide metabolism at the plasma membrane and in extracellular milieu, Journal of Biological Chemistry, 10.1074/jbc.M506827200, 280, 44, 36592-36600, 280(44),36592-36600, 2005.11.
||Shin-ichi Chisada, Yasuhiro Horibata, Yoichiro Hama, Masanori Inagaki, Naruto Furuya, Nozomu Okino and Makoto Ito, The glycosphingolipid receptor for Vibrio trachuri in the red sea bream intestine is a GM4 ganglioside which contains 2-hydroxy fatty acids., Biochemical and Biophysical Research Communications, 10.1016/j.bbrc.2005.05.110, 333, 2, 367-373, 333, 367-373, 2005.01.
||Tetsuto Nakagawa, Akio Morotomi, Motohiro Tani, Noriyuki Sueyoshi, Hironobu Komori, and Makoto Ito, C18:3-GM1a induces appoptosis in neuro2a cells: enzymatic remodeling of fatty acyl chains of glycosphingolipids, Journal of Lipid Reserch, 10.1194/jlr.M400516-JLR200, 46, 6, 1103-1112, 46, 1103-1112, 2005.01.
||Motohiro Tani, Nozomu Okino, Noriyuki Sueyoshi, and Makoto Ito, Conserved amino acid residues in the COOH-terminal tail are indispenzable for the correct folding and localization and enzyme activity of neutral ceramidase., Journal of Biological Chemistry, 10.1074/jbc.M404012200, 279, 28, 29351-29358, 2004.07.
||Yukihiro Yoshimura, Motohiro Tani, Nozomu Okino, Hiroshi Iida, and Makoto Ito, Molecular cloning and functional analysis of zebrafish neutral ceramidase., Journal of Biological Chemistry, 10.1074/jbc.M405598200, 279, 42, 44012-44022, 279(42), 44012-44022, 2004.10.
||Yasuhiro Horibata, Keishi Sakaguchi, Nozomu Okino, Hiroshi Iida, Masanori Inagaki, Yoichiro Hama and Makoto Ito, Unique catabolic pathway of glycosphingolipids in a Hydrozoan, Hydra magnipapillata, involving endoglycoceramidase, Journal of Biological Chemistry, 10.1074/jbc.M401460200, 279, 32, 33379-33389, 279(32), 33379-33389, 2004.08.
||Motohiro Tani, Hiroshi Iida, Makoto Ito, O-Glycosylation of mucin-like domain retains the neutral ceamidase on the plasma membranes as a type II integral membrane protein, Journal of Biological Chemistry, 10.1074/jbc.M207932200, 278, 12, 10523-10530, 278 (12), 10523-10530, 2003.03.
||Masako Furusato, Noriyuki Sueyoshi, Susumu Mitsutake, Makoto Ito et. al., Molecular cloning and characterization of sphingolipid ceramide N-deacylase from a marine bacterium, Shewanella alga G8, Journal of Biological Chemistry, 10.1074/jbc.m110688200, 277, 19, 17300-17307, 277(19), 17300-17307, 2002.05.
||Susumu Mitsutake, Motohiro Tani, Nozomu Okino, Makoto Ito et al., Purification, characterization, molecular cloning , and subcellular distribution of neutral ceramidase of rat kidney., Journal of Biological Chemistry, 10.1074/jbc.M102233200, 276, 28, 26249-26259, 276(28), 26249-26259, 2001.07.
||Motohiro Tani, Nozomu Okino, Susumu Mitsutake, Makoto Ito et al., Purification and characterization of a neutral ceamidase from mouse liver: A single protein catalyzes the reversile reaction in which ceramide is both hydrolyzed and synthesized, Journal of Biological Chemistry, 10.1074/jbc.275.5.3462, 275, 5, 3462-3468, 275(5), 3462-3468, 2000.02.
||Motohiro Tani, Nozomu Okino, Kaoru Mori, Makoto Ito et al., Molecular cloning ot the full-length cDNA encoding mouse neutral ceamidase: A novel but highly conserved gene family of neutral ceramidase, Journal of Biological Chemistry, 10.1074/jbc.275.15.11229, 275, 15, 11229-11234, 275(15), 11229-11234, 2000.04.
||Yasuhiro Horibata, Nozomu Okino, Sachiyo Ichinose, Akira Omori, Makoto Ito, Purification, characteriation, and cDNA cloning of a novel acidic endoglycoceramidase from the jellyfish, Cyanea nozakii, Journal of Biological Chemistry, 10.1074/jbc.M003575200, 275, 40, 31297-31304, 275(40), 31297-31304, 2000.10.
||Hironobu Komori, Shinichi Ichikawa, Yoshio Hirabayashi, and Makoto Ito, Regulation of intracelular ceramide content in B16 melanoma cells: Biological implications of ceramide glycosylation, Journal of Biological Chemistry, 10.1074/jbc.274.13.8981, 274, 13, 8981-8987, 274(13), 8981-8987, 1999.03.
||Nozomu Okino, Motohiro Tani, Shuhei Imayama, and Makoto Ito, Purification and characterization of a novel ceramidase from Pseudomonas aeruginosa, Journal of Biological Chemistry, 10.1074/jbc.273.23.14368, 273, 23, 14368-14373, 273(23), 14368-14373, 1999.06.