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M Fukuda

Publications and source records attributed to M Fukuda.

At least 37 records · Page 2Linked to original sources
PubMed2001

Brain lateralization for mismatch response to across- and within-category change of vowels.

Differences in hemispheric predominance between across- and within-category change perception of vowels were assessed using a whole-head magnetoencephalography. The magnetic mismatch responses (MMNm) to pure-tone and vowel within-category changes were significantly predominant in the right hemisphere; on the other hand, vowel across-category MMNm did not differ in power between hemispheres. The results suggest that both hemispheres are symmetrically activated in the preattentive across-category change perception of vowels, while the within-category change of a vowel is analyzed as the change in physical features of the stimuli, thus predominantly activating the right hemisphere. Thus, the relative contribution of the left auditory cortex in the preattentive speech processing may occur only at the level of perception of the vowel across-category change.

Acoustic Stimulation↗
PubMed2001

Synaptotagmin VI participates in the acrosome reaction of human spermatozoa.

Acrosomal exocytosis is a calcium-dependent secretion event causing the release of the acrosomal contents and the loss of the membranes surrounding the acrosome. The synaptotagmins are a family of calcium-binding proteins that participate in the exocytosis of synaptic vesicles. The ubiquitous synaptotagmin VI isoform was found in human sperm cells by Western blot analysis. Immunocytochemistry at the optical and electron microscopy levels localized the protein to the outer acrosomal membrane. Calcium-triggered acrosomal exocytosis in permeabilized sperm cells was abrogated by a specific anti-synaptotagmin VI antibody, indicating that the protein is required for the process. Moreover, a recombinant fusion protein between glutathione S-transferase and the two calcium and phospholipid binding domains of synaptotagmin VI completely inhibited calcium-triggered exocytosis. Interestingly, phorbol ester-dependent in vitro phosphorylation of this recombinant protein abolished its inhibitory effect. We previously showed that, in permeabilized spermatozoa, addition of active Rab3A triggers acrosomal exocytosis at very low calcium concentration. Rab3A-promoted exocytosis was inhibited by the cytosolic domain of synaptotagmin VI and by the anti-synaptotagmin VI antibody, indicating that synaptotagmin is also necessary for Rab-mediated acrosomal content release. In conclusion, the results strongly indicate that synaptotagmin VI is a key component of the secretory machinery involved in acrosomal exocytosis.

Acrosome Reaction↗
PubMed2001

Novel sulfated lymphocyte homing receptors and their control by a Core1 extension beta 1,3-N-acetylglucosaminyltransferase.

L-selectin mediates lymphocyte homing by facilitating lymphocyte adhesion to addressins expressed in the high endothelial venules (HEV) of secondary lymphoid organs. Peripheral node addressin recognized by the MECA-79 antibody is apparently part of the L-selectin ligand, but its chemical nature has been undefined. We now identify a sulfated extended core1 mucin-type O-glycan, Gal beta 1-->4(sulfo-->6)GlcNAc beta 1-->3Gal beta 1-->3GalNAc, as the MECA-79 epitope. Molecular cloning of a HEV-expressed core1-beta 1,3-N-acetylglucosaminyltransferase (Core1-beta 3GlcNAcT) enabled the construction of the 6-sulfo sialyl Lewis x on extended core1 O-glycans, recapitulating the potent L-selectin-mediated, shear-dependent adhesion observed with novel L-selectin ligands derived from core2 beta1,6-N-acetylglucosaminyltransferase-I null mice. These results identify Core1-beta 3GlcNAcT and its cognate extended core1 O-glycans as essential participants in the expression of the MECA-79-positive, HEV-specific L-selectin ligands required for lymphocyte homing.

Animals↗
PubMed2001

Crystal structure of 2-hydroxyl-6-oxo-6-phenylhexa-2,4-dienoic acid (HPDA) hydrolase (BphD enzyme) from the Rhodococcus sp. strain RHA1 of the PCB degradation pathway.

2-Hydroxyl-6-oxo-6-phenylhexa-2,4-dienoic acid (HPDA) hydrolase (the BphD enzyme) hydrolyzes a ring-cleavage product of an aromatic compound generated in a biphenyl/polychlorinated biphenyl (PCB) degradation pathway of bacteria. The crystal structure of the BphD enzyme has been determined at 2.4 A resolution by the multiple isomorphous replacement method. The final refined model of the BphD enzyme yields an R-factor of 17.5 % at 2.4 A resolution with reasonable geometry. The BphD enzyme is an octameric enzyme with a 422 point-group symmetry. The subunit can be divided into core and lid domains. The active site of the enzyme is situated in the substrate-binding pocket, which is located between the two domains. The substrate-binding pocket can be divided into hydrophobic and hydrophilic regions. This feature of the pocket seems to be necessary for substrate binding, as the substrate is composed of hydrophilic and hydrophobic parts. The proposed orientation of the substrate seems to be consistent with the general catalytic mechanism of alpha/beta-hydrolases.

Amino Acid Sequence↗
PubMed2001

Mechanism of the calcium-dependent multimerization of synaptotagmin VII mediated by its first and second C2 domains.

The Ca(2+)-dependent oligomerization activity of the second C2 (C2B) domain of synaptotagmin I (Syt I) has been hypothesized to regulate neurotransmitter release. We previously showed that the cytoplasmic domains of several other Syt isoforms also show Ca(2+)-dependent oligomerization activity (Fukuda, M., and Mikoshiba, K. (2000) J. Biol. Chem. 275, 28180-28185), but little is known about the involvement of their C2 domains in Ca(2+)-dependent oligomerization. In this study, we analyzed the Ca(2+)-dependent oligomerization properties of the first (C2A) and the second C2 (C2B) domains of Syt VII. Unlike Syt I, both C2 domains of Syt VII contribute to Ca(2+)-dependent homo- and hetero-oligomerization with other isoforms. For instance, the Syt VII C2A domain Ca(2+)-dependently binds itself and the C2A domain of Syt VI but not its C2B domain, whereas the Syt VII C2B domain Ca(2+)-dependently binds itself and the C2B domain of Syt II but not its C2A domain. In addition, we showed by gel filtration that a single Syt VII C2 domain is sufficient to form a Ca(2+)-dependent multimer of very high molecular weight. Because of this "two handed" structure, the Syt VII cytoplasmic domain has been found to show the strongest Ca(2+)-dependent multimerization activity in the Syt family. We also identified Asn-328 in the C2B domain as a crucial residue for the efficient Ca(2+)-dependent switch for multimerization by site-directed mutagenesis. Our results suggest that Syt VII is a specific isoform that can cluster different Syt isoforms with two hands in response to Ca(2+).

Amino Acid Sequence↗
PubMed2001

The C2A domain of double C2 protein gamma contains a functional nuclear localization signal.

The C2 domain was originally defined as a homologous domain to the C2 regulatory region of Ca2+ -dependent protein kinase C and has been identified in more than 50 different signaling molecules. The original C2 domain of protein kinase Calpha functions as a Ca2+ binding module, and the Ca2+ binding to the C2 domain allows translocation of proteins to phospholipid membranes. By contrast, however, some C2 domains do not exhibit Ca2+ binding activity because of amino acid substitutions at Ca2+ -binding sites, and their physiological meanings remain largely unknown. In this study, we discovered an unexpected function of the Ca2+ -independent C2A domain of double C2 protein gamma (Doc2gamma) in nuclear localization. Deletion and mutation analyses revealed that the putative Ca2+ binding loop 3 of Doc2gamma contains six Arg residues ((177)RLRRRRR(183)) and that this basic cluster is both necessary and sufficient for nuclear localization of Doc2gamma. Because of the presence of the basic cluster, the C2A domain of Doc2gamma did not show Ca2+ -dependent phospholipid binding activity. Our findings indicate that by changing the nature of the putative Ca2+ binding loops the C2 domain has more diversified function in cellular signaling than a simple Ca2+ binding motif.

Amino Acid Sequence↗
PubMed2001

Herniation of cervical intervertebral disc: immunohistochemical examination and measurement of nitric oxide production.

STUDY DESIGN: Surgically obtained cervical herniated intervertebral discs were examined histologically and immunohistochemically. The production of nitric oxide (NO) in the local tissue was examined using the electron spin resonance (ESR) method. OBJECTIVES: To investigate the local histologic and immunohistochemical changes in cervical disc herniation, including NO production, and to compare such changes with those in autopsy cases. SUMMARY OF BACKGROUND DATA: Very little is known about the histopathologic processes of cervical disc herniation. In addition, no information is available on the level of in vivo NO production in cervical disc herniation. METHODS: Thirty-six herniated cervical discs obtained from 31 patients were immunohistochemically examined for localization of blood vessels, matrix metalloproteinase (MMP)-3, and inducible NO synthetase (iNOS). We also compared the production of NO, measured by the ESR method, in eight specimens with that of five control discs obtained from fresh cadavers. RESULTS: The presence of herniated discs correlated with the degeneration of cartilaginous endplate and torn anulus fibrosus. Formation of new blood vessels around the herniated discs was detected, using von Willebrand factor antibody, in seven uncontained hernias and 20 contained hernias. Immunohistochemical studies showed the presence of cells positive for MMP-3 (chondrocytes), iNOS (chondrocytes and granulation tissue) in cervical disc hernias. ESR analysis showed a significantly higher NO production in herniated cervical discs than in disc samples of fresh cadavers. CONCLUSIONS: Herniated cervical intervertebral disc is characterized by the presence of an inflammatory process associated with neovascularization and increased expression of MMP-3. Production of NO was markedly high in both contained- and uncontained-type hernias.

Adult↗
PubMed2001

Novel splicing isoforms of synaptotagmin-like proteins 2 and 3: identification of the Slp homology domain.

Slp1-3 (synaptotagmin-like protein 1-3) is a new family of carboxyl-terminal-type (C-type) tandem C2 proteins that show higher sequence similarity to the C2 domains of granuphilin-a/Slp-4 than those of other C-type tandem C2 proteins (e.g., synaptotagmin and the Doc2 family). However, the amino (N)-terminal domains of the original Slp1-3 do not contain any known protein motifs and do not show any sequence similarities to each other. We report four alternative splicing isoforms of Slp2 (designated Slp2-a-d, with the original Slp2 renamed Slp2-c) and two alternative splicing isoforms of Slp3 (Slp3-a and Slp3-b, the original Slp3). These isoforms share the same C-terminal tandem C2 structures, but their N-terminal nucleotide sequences are completely different due to the alternate use of different exons. Sequence alignment of the Slp1, Slp2-a, Slp3-a, and Slp4 amino terminal domains reveals the presence of two conserved regions among the Slp family, designated SHD1 (Slp homology domain 1) and SHD2, which may function as protein interaction sites. The SHD1 and SHD2 of Slp3-a and Slp4 are separated by a putative Zn(2+)-binding sequence, whereas Slp1 and Slp2 lack such Zn(2+)-binding sequences and their SHD1 and SHD2 are linked together. In addition, we show that the Slp2-a/c/d mRNAs are differentially distributed in different mouse tissues and at different stages of development, suggesting that these transcripts may be regulated by different promoters.

Alternative Splicing↗
PubMed2001

Molecular cloning and expression of a novel human beta-Gal-3-O-sulfotransferase that acts preferentially on N-acetyllactosamine in N- and O-glycans.

A novel cDNA-encoding galactose 3-O-sulfotransferase was cloned by screening the expressed sequence tag data base using the previously cloned cDNA encoding a galactosyl ceramide 3-O-sulfotransferase, which we term Gal3ST-1. The newly isolated cDNA encodes a novel 3-O-sulfotransferase, termed Gal3ST-3, that acts exclusively on N-acetyllactosamine present in N-glycans and core2-branched O-glycans. These conclusions were confirmed by analyzing CD43 chimeric proteins in Chinese hamster ovary cells expressing core2 beta1,6-N-acetylglucosaminyltransferase. The acceptor specificity of Gal3ST-3 contrasts with that of the recently cloned galactose 3-O-sulfotransferase (Honke, K., Tsuda, M., Koyota, S., Wada, Y., Iida-Tanaka, N., Ishizuka, I., Nakayama, J., and Taniguchi, N. (2001) J. Biol. Chem. 276, 267-274), which we term Gal3ST-2 in the present study because the latter enzyme can also act on core1 O-glycan and type 1 oligosaccharides, Galbeta1-->3GlcNAc. Moreover, Gal3ST-3 but not Gal3ST-2 can act on Galbeta1-->4(sulfo-->6)GlcNAc, indicating that disulfated sulfo-->3Galbeta1-->4(sulfo-->6) GlcNAc-->R may be formed by Gal3ST-3 in combination with GlcNAc 6-O-sulfotransferase. Although both Gal3ST-2 and Gal3ST-3 do not act on galactosyl ceramide, Gal3ST-3 is only moderately more homologous to Gal3ST-2 (40.1%) than to Gal3ST-1 (38.0%) at the amino acid level. Northern blot analysis demonstrated that transcripts for Gal3ST-3 are predominantly expressed in the brain, kidney, and thyroid where the presence of 3'-sulfation of N-acetyllactosamine has been reported. These results indicate that the newly cloned Gal3ST-3 plays a critical role in 3'-sulfation of N-acetyllactosamine in both O- and N-glycans.

Amino Acid Sequence↗
PubMed2001

Electromagnetic moments of the beta-emitting nucleus 16N.

The nuclear magnetic dipole moment mu and electric quadrupole moment Q of the beta-emitting 16N(Ipi = 2(-), T(1/2) = 7.13 s) nucleus have been determined for the first time by detecting its beta-NMR in a MgO crystal and beta-NQR (nuclear quadrupole resonance) in a TiO (2) crystal to be /mu/ = (1.9859+/-0.0011) mu(N) and /Q/ = (17.9+/-1.7) mb, respectively. Although the prediction of mu given by the Hartree-Fock calculation agrees well with the experiment, an abnormally small effective charge for neutrons is required to account for the experimental Q.

Journal Article↗
PubMed2001

Impaired suppression of processing in schizophrenic patients suggested by ERPs obtained in a selective attention task.

In the present study, we focused on the ability of suppression of processing in schizophrenic patients, using event-related potentials (ERPs) recorded during a selective attention task. During the task, subjects were required to focus on one ear, counting deviant stimuli, those deviating in duration from a sequence of standard stimuli. We compared amplitude data of two positive components differing in latency elicited by standard stimuli, which reflect suppression of stimulus processing, between schizophrenic patients and normal controls. Significant between-group differences were obtained specifically in the right ear attended condition, suggesting impaired suppression of processing in schizophrenics mediated in the left hemisphere.

Adult↗
PubMed2001

In vitro ubiquitination of cyclin D1 by ROC1-CUL1 and ROC1-CUL3.

Overexpression of cyclin D1 has been implicated in a variety of tumors, such as breast cancers, gastrointestinal cancers and lymphomas. Both gene amplification and protein degradation mediated by ubiquitin (Ub)-dependent proteolysis regulate the abundance of cyclin D1. Here we report that ROC1 interacted with all three D type cyclins in vivo but did not bind to other cyclins tested. The ROC1-CUL1 and ROC1-CUL3, but not ROC1-CUL2, -CUL3 and -CUL4, immunocomplexes promoted polyubiquitination of bacterially purified cyclin D1 in vitro. RING finger mutations of ROC1 eliminated the Ub ligase activity toward cyclin D1. In all cases the ubiquitination of cyclin D1 was accompanied by autoubiquitination of the cullins. The results suggest the involvement of ROC1-cullin ligases in cyclin D1 ubiquitination and a potential mechanism whereby the cullin subunit is ubiquitinated itself while ubiquitinating a substrate.

Anaphase-Promoting Complex-Cyclosome↗
PubMed2001

Cutting edge: CD43 functions as a T cell counterreceptor for the macrophage adhesion receptor sialoadhesin (Siglec-1).

Sialoadhesin (Siglec-1) is a macrophage-restricted sialic acid-binding receptor that mediates interactions with hemopoietic cells, including lymphocytes. In this study, we identify sialoadhesin counterreceptors on T lymphocytes. Several major glycoproteins (85, 130, 240 kDa) were precipitated by sialoadhesin-Fc fusion proteins from a murine T cell line (TK-1). Binding of sialoadhesin to these glycoproteins was sialic acid dependent and was abolished by mutation of a critical residue (R97A) of the sialic acid binding site in the membrane distal Ig-like domain of sialoadhesin. The 130- and 240-kDa sialoadhesin-binding glycoproteins were identified as the sialomucins CD43 and P-selectin glycoprotein ligand 1 (CD162), respectively. CD43 expressed in COS cells supported increased binding to immobilized sialoadhesin. Finally, sialoadhesin bound different glycoforms of CD43 expressed in Chinese hamster ovary cells, including unbranched (core 1) and branched (core 2) O:-linked glycans, that are normally found on CD43 in resting and activated T cells, respectively. These results identify CD43 as a T cell counterreceptor for sialoadhesin and suggest that in addition to its anti-adhesive role CD43 may promote cell-cell interactions.

Animals↗
PubMed2001

C2 domains from different Ca2+ signaling pathways display functional and mechanistic diversity.

The ubiquitous C2 domain is a conserved Ca2+ triggered membrane-docking module that targets numerous signaling proteins to membrane surfaces where they regulate diverse processes critical for cell signaling. In this study, we quantitatively compared the equilibrium and kinetic parameters of C2 domains isolated from three functionally distinct signaling proteins: cytosolic phospholipase A2-alpha (cPLA2-alpha), protein kinase C-beta (PKC-beta), and synaptotagmin-IA (Syt-IA). The results show that equilibrium C2 domain docking to mixed phosphatidylcholine and phosphatidylserine membranes occurs at micromolar Ca2+ concentrations for the cPLA2-alpha C2 domain, but requires 3- and 10-fold higher Ca2+ concentrations for the PKC-beta and Syt-IA C2 domains ([Ca2+](1/2) = 4.7, 16, 48 microM, respectively). The Ca2+ triggered membrane docking reaction proceeds in at least two steps: rapid Ca2+ binding followed by slow membrane association. The greater Ca2+ sensitivity of the cPLA2-alpha domain results from its higher intrinsic Ca2+ affinity in the first step compared to the other domains. Assembly and disassembly of the ternary complex in response to rapid Ca2+ addition and removal, respectively, require greater than 400 ms for the cPLA2-alpha domain, compared to 13 ms for the PKC-beta domain and only 6 ms for the Syt-IA domain. Docking of the cPLA2-alpha domain to zwitterionic lipids is triggered by the binding of two Ca2+ ions and is stabilized via hydrophobic interactions, whereas docking of either the PKC-beta or the Syt-IA domain to anionic lipids is triggered by at least three Ca2+ ions and is maintained by electrostatic interactions. Thus, despite their sequence and architectural similarity, C2 domains are functionally specialized modules exhibiting equilibrium and kinetic parameters optimized for distinct Ca2+ signaling applications. This specialization is provided by the carefully tuned structural and electrostatic parameters of their Ca2+ and membrane-binding loops, which yield distinct patterns of Ca2+ coordination and contrasting mechanisms of membrane docking.

Binding Sites↗
PubMed2001

The RING heterodimer BRCA1-BARD1 is a ubiquitin ligase inactivated by a breast cancer-derived mutation.

BRCA1-BARD1 constitutes a heterodimeric RING finger complex associated through its N-terminal regions. Here we demonstrate that the BRCA1-BARD1 heterodimeric RING finger complex contains significant ubiquitin ligase activity that can be disrupted by a breast cancer-derived RING finger mutation in BRCA1. Whereas individually BRCA1 and BARD1 have very low ubiquitin ligase activities in vitro, BRCA1 combined with BARD1 exhibits dramatically higher activity. Bacterially purified RING finger domains comprising residues 1-304 of BRCA1 and residues 25-189 of BARD1 are capable of polymerizing ubiquitin. The steady-state level of transfected BRCA1 in vivo was increased by co-transfection of BARD1, and reciprocally that of transfected BARD1 was increased by BRCA1 in a dose-dependent manner. The breast cancer-derived BARD1-interaction-deficient mutant, BRCA1(C61G), does not exhibit ubiquitin ligase activity in vitro. These results suggest that the BRCA1-BARD1 complex contains a ubiquitin ligase activity that is important in prevention of breast and ovarian cancer development.

BRCA1 Protein↗
PubMed2001

Characterization of KIAA1427 protein as an atypical synaptotagmin (Syt XIII).

Synaptotagmin (Syt) belongs to a family of type-I membrane proteins and is a protein that consists of a short extracellular N-terminus, a single transmembrane domain, two C2 domains and a short C-terminus. Here, we cloned and characterized a mouse orthologue of human KIAA1427 protein as an atypical Syt (named Syt XIII). Subcellular fractionation and antibody-uptake experiments indicate that Syt XIII is indeed a type-I membrane protein, but, unlike other Syt isoforms, lacks an N-terminal extracellular domain. Syt XIII C2 domains show relatively little similarity to Syt I (less than 35% identity at the amino acid level), and lack key amino acids responsible for Ca2+ binding. Because of these substitutions, the Syt XIII C2 domains did not show Ca2+-dependent phospholipid-binding activity, and Syt XIII is thus classified as a Ca2+ -independent isoform. By contrast, the Syt XIII C-terminal domain is highly homologous with other Syt isoforms and can function as a common receptor for neurexin Ialpha in vitro. Since Syt XIII is expressed in various tissues outside the brain, Syt XIII may be involved in constitutive vesicle transport.

Amino Acid Sequence↗
PubMed2001

Clinicopathological significance of core 2 beta1,6-N-acetylglucosaminyltransferase messenger RNA expressed in the pulmonary adenocarcinoma determined by in situ hybridization.

Cell surface carbohydrates of epithelial cells play important roles in tumor progression. Previously, we have shown that expression of core 2 branched O-glycans in colorectal cancer is closely correlated with the vessel invasion and depth of invasion (K. Shimodaira et al., Cancer Res., 57: 5201-5206, 1997). To test whether this is also the case in human lung cancer, we have examined the expression pattern of core 2 beta1,6-N-acetylglucosaminyltransferase (C2GnT) mRNA responsible for the biosynthesis of core 2 branched O-glycans in 41 cases of lung cancer. Using in situ hybridization, C2GnT mRNA was detected in 73.2% of the lung cancer cells, irrespective of the histopathological type; whereas in normal lung tissues, its expression was restricted to the basal cells of bronchial mucosa. These results indicate that the expression level of C2GnT mRNA was significantly enhanced in association with malignant transformation. Statistical analysis between the C2GnT mRNA expressed in pulmonary adenocarcinoma and clinicopathological variables revealed that the expression of C2GnT was correlated with vessel invasion and lymph node metastasis with significant difference (P < 0.05), but expression of sialyl Le(x), which is frequently expressed in the adenocarcinoma, was not significantly correlated with lymph node metastasis. These results indicate that C2GnT mRNA detected by in situ hybridization reflects the malignant potentials of pulmonary adenocarcinoma, because lymph node metastasis is the most affecting factor to the patients' prognosis.

Adenocarcinoma↗