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Y Yoneda

Publications and source records attributed to Y Yoneda.

At least 19 recordsLinked to original sources
PubMed2002

Differences between D- and L-aspartate binding to the Na+-dependent binding sites on glutamate transporters in frozen sections of rat brain.

The Na+-dependent, "high-affinity" transport of L-glutamate (GluT) in brain tissue has become a significant focus of interest, particularly since it has been revealed that abnormalities of GluT may be associated with serious neurological disorders. Using quantitative autoradiography on 3H-sensitive films, we have studied, in thaw-mounted sections of rat brain, the distribution and pharmacology of radioligand binding to sites with characteristics of the substrate-recognition/binding locus on GluT. The technique makes it possible to determine not only the intensity of binding in brain regions but, with a high level of precision, pharmacological constants such as IC50 or nH. [3H]L-aspartate and [3H]D-aspartate are two classical radioligands used in studies of GluT. We have determined IC50 values for the inhibition of [3H]L- and [3H]D-aspartate binding by their non-radioactive counterparts in the cerebral neocortex. hippocampus, striatum, septal nuclei and the cerebellar cortex. The two radioligands did not appreciably differ from each other in their interactions with the binding sites in the forebrain, consistent with all Na+-dependent GluT binding sites in that region having no stereoselectivity for aspartate enantiomers. In the cerebellar cortex, however, the data indicated the presence of a GluT binding site that preferred L- over D-aspartate. These findings contrast with many previous observations and suggest that the pharmacological characteristics of the ligand binding sites on GluT in the mammalian cerebellar cortex may have to be re-assessed and/or a possibility of an existence of (a) hitherto unknown molecule(s) with properties of a glutamate transporter be considered.

Amino Acid Transport System X-AG↗
PubMed2001

Degradation of c-Fos protein expressed by N-methyl-D-aspartic acid in nuclear fractions of murine hippocampus.

In both nuclear and cytosolic fractions of murine hippocampus, constitutive expression was seen with Fra-2 protein, but not with other Fos family members tested including c-Fos, Fos-B and Fra-1 proteins. Fos-B protein was only detected in nuclear fractions. The systemic administration of N-methyl-D-aspartic acid (NMDA) induced marked and transient expression of c-Fos protein, but not other family members, in both hippocampal fractions 2 h later. In vitro incubation at 30 degrees C led to more rapid degradation of inducible c-Fos protein than constitutive Fra-2 protein in nuclear fractions obtained 2 h after the administration of NMDA, without significantly affecting that of both member proteins in cytosolic fractions. The addition of phosphatase inhibitors significantly delayed the initial degradation rate of inducible c-Fos protein, with concomitant facilitation of that of constitutive Fra-2 protein, in nuclear fractions. The addition of protease inhibitors also delayed the initial degradation of constitutive Fra-2 protein, without markedly altering that of inducible c-Fos protein, in nuclear fractions. Immunoprecipitation analysis revealed that NMDA induced phosphorylation of c-Fos protein on tyrosine residues in nuclear fractions to a lesser extent than that on serine residues 2 h after administration. These results suggest that NMDA signals may be propagated to the nucleus to induce both expression and degradation of c-Fos protein through a molecular mechanism associated with phosphorylation on serine and/or tyrosine residues in murine hippocampus.

Animals↗
PubMed2001

Androgen regulation of CYP4B1 responsible for mutagenic activation of bladder carcinogens in the rat bladder: detection of CYP4B1 mRNA by competitive reverse transcription-polymerase chain reaction.

Significant sex differences exist among cases of bladder cancer in humans as well as in experimental animals such as rats. Aromatic amines such as benzidine and 2-naphthylamine are known to induce bladder cancer. These carcinogenic amines are activated to genotoxic substances by cytochrome P 450 CYP4B1, which is present in bladder mucosa. In this study, regulation of CYP4B1 was investigated to elucidate sex difference in bladder carcinogenesis. Competitive reverse transcription-polymerase chain reaction was used to investigate the expression of rat CYP4B1 mRNA occurring in small amounts of tissue such as bladder tissue. Expression of CYP4B1 in the bladder of male rats increased with development but not in that of female rats. Moreover, mature male rats exhibited higher expression of CYP4B1 in the bladder than did mature female rats. Castration of male rats decreased CYP4B1 levels and treatment with testosterone led to a partial recovery of CYP4B1 levels. These results indicate that CYP4B1 levels in the rat bladder are partly regulated by androgens. Furthermore, the present findings suggest that the sex difference observed in bladder carcinogenesis was due to sex-different expression of CYP4B1 in bladder tissue.

Age Factors↗
PubMed2001

A novel nucleolar protein, NIFK, interacts with the forkhead associated domain of Ki-67 antigen in mitosis.

In a previous study, we demonstrated that the forkhead associated (FHA) domain of pKi-67 interacts with the novel kinesin-like protein, Hklp2 (Sueishi, M., Takagi, M., and Yoneda, Y. (2000) J. Biol. Chem. 275, 28888-28892). In this study, we report on the identification of a putative RNA-binding protein of 293 residues as another binding partner of the FHA domain of pKi-67 (referred to as NIFK for nucleolar protein interacting with the FHA domain of pKi-67). Human NIFK (hNIFK) interacted with the FHA domain of pKi-67 (Ki-FHA) efficiently in vitro when hNIFK was derived from mitotically arrested cells. In addition, a moiety of hNIFK was co-localized with pKi-67 at the peripheral region of mitotic chromosomes. The hNIFK domain that interacts with Ki-FHA was mapped in the yeast two-hybrid system to a portion encompassed by residues 226-269. In a binding assay utilizing Xenopus egg extracts, it was found that the mitosis-specific environment and two threonine residues within this portion of hNIFK (Thr-234 and Thr-238) were crucial for the efficient interaction of hNIFK and Ki-FHA, suggesting that hNIFK interacts with Ki-FHA in a mitosis-specific and phosphorylation-dependent manner. These findings provide a new clue to our understanding of the cellular function of pKi-67.

Amino Acid Sequence↗
PubMed2001

Differential in vitro degradation of particular Fos family members expressed by kainic acid in nuclear and cytosolic fractions of murine hippocampus.

Mice were injected with kainic acid (KA) at a convulsive dose, followed by homogenization of the hippocampus in the presence of different protease and phosphatase inhibitors, and subsequent preparation of nuclear and cytosolic fractions. An intraperitoneal injection of KA resulted in marked expression of particular Fos family members, including c-Fos, Fra-2, and Fos-B, but not Fra-1 proteins, in both fractions 2 to 18 h after administration. These fractions were individually incubated at 30 degrees C for 1 to 18 h for determination of in vitro degradation. Similarly rapid degradation was seen with c-Fos protein between nuclear fractions obtained 2 and 18 h after administration, while no significant degradation was found for c-Fos protein in cytosolic fractions obtained 2 h after administration during incubation. By contrast, in vitro incubation led to rapid degradation of c-Fos protein in cytosolic fractions obtained 18 h after administration. Degradation profiles were peculiar to each member protein in nuclear and cytosolic fractions obtained 2 and 18 h after administration. Dialysis prevented degradation of c-Fos protein in nuclear fractions without markedly affecting that in cytosolic fractions in a manner independent of the time after administration. The addition of inhibitors for phosphatases, but not for proteases, accelerated the degradation of c-Fos protein in nuclear fractions previously dialyzed. These results suggest that in vivo KA signals may modulate heterologous machineries responsible for breakdown of each Fos family member in a unique manner in nuclear fractions, rather than cytosolic fractions, of murine hippocampus.

Animals↗
PubMed2001

Group III metabotropic glutamate receptors in rat cultured calvarial osteoblasts.

Reverse transcription polymerase chain reaction revealed expression of mRNAs for particular receptors for the central neurotransmitter l-glutamic acid (Glu) in primary cultures of rat calvarial osteoblastic cells under premature to mature states according to the duration of days in vitro. These included metabotropic Glu receptors (mGluR) such as mGluR4 and mGluR8, in addition to several ionotropic Glu receptor subunits including NR1 and NR2D. Expression of mRNAs was not detected with other mGluR and NR2A-C subunits irrespective of the maturity of cultured cells. The agonist for group III mGluR L-(+)-2-amino-4-phosphonobutyric acid significantly inhibited the forskolin-induced accumulation of cAMP in premature osteoblasts, which occurred in a manner sensitive to prevention by the group III mGluR antagonist (RS)-alpha-cyclopropyl-4-phosphonophenylglycine. These results suggest that Glu may at least in part play a role in mechanisms associated with cellular proliferation and/or differentiation through group III mGluR functionally expressed in rat calvarial osteoblastic cells.

1-Methyl-3-isobutylxanthine↗
PubMed2001

Nuclear import of the U1A splicesome protein is mediated by importin alpha /beta and Ran in living mammalian cells.

U1A is a component of the uracil-rich small nuclear ribonucleoprotein. The molecular mechanism of nuclear import of U1A was investigated in vivo and in vitro. When recombinant deletion mutants of U1A are injected into the BHK21 cell cytoplasm, the nuclear localization signal (NLS) of U1A is found in the N-terminal half of the central domain (residues 100-144 in mouse U1A). In an in vitro assay, it was found that the U1A-NLS accumulated in only a portion of the nuclei in the absence of cytosolic extract. In contrast, the addition of importin alpha/beta and Ran induced the uniform nuclear accumulation of U1A-NLS in all cells. Furthermore, U1A was found to bind the C-terminal portion of importin alpha. In addition, the in vitro nuclear migration of full-length U1A was found to be exclusively dependent on importin alpha/beta and Ran. Moreover, in living cells, the full-length U1A accumulated in the nucleus in a Ran-dependent manner, and nuclear accumulation was inhibited by the importin beta binding domain of importin alpha. These results suggest that the nuclear import of U1A is mediated by at least two distinct pathways, an importin alpha/beta and Ran-dependent and an -independent pathway in permeabilized cells, and that the latter pathway may be suppressed in intact cells.

Amino Acid Sequence↗
PubMed2001

A role for Hsc70 in regulating nucleocytoplasmic transport of a temperature-sensitive p53 (p53Val-135).

Mouse temperature-sensitive p53(Val-135) accumulates in the nucleus and acts as a "wild-type" at 32 degrees C while it is sequestered in the cytoplasm at 37 degrees C. The cytoplasmic p53(Val-135) relocalized into the nucleus upon inhibition of the nuclear export at 37 degrees C, whereas a mutation in a major bipartite nuclear localization signal (NLS) caused constitutive cytoplasmic localization, indicating that it shuttled between the cytoplasm and the nucleus by its own nuclear export signal and NLS rather than tethered to cytoplasmic structures. Although the full-length p53(Val-135) did not bind the import receptor at 37 degrees C, a C-terminally truncated p53(Val-135) lacking residues 326-390 did bind it. Molecular chaperones such as Hsc70 were associated with p53(Val-135) at 37 degrees C but not at 32 degrees C. When the nuclear export was blocked by leptomycin B, only a fraction lacking Hsc70 was specifically accumulated in the nucleus. Immunodepletion of Hsc70 from the reticulocyte lysate caused p53(Val-135) to bind the import receptor. This binding was blocked by supplying the cell extract containing Hsc70 but not by the addition of recombinant Hsc70 alone. We suggest that the association with the Hsc70-containing complex prevents the NLS from the access of the import receptor through the C-terminal region of p53(Val-135) at 37 degrees C, whereas its dissociation at 32 degrees C allows rapid nuclear import.

Amino Acid Sequence↗
PubMed2001

Essential role of voltage-dependent anion channel in various forms of apoptosis in mammalian cells.

Through direct interaction with the voltage-dependent anion channel (VDAC), proapoptotic members of the Bcl-2 family such as Bax and Bak induce apoptogenic cytochrome c release in isolated mitochondria, whereas BH3-only proteins such as Bid and Bik do not directly target the VDAC to induce cytochrome c release. To investigate the biological significance of the VDAC for apoptosis in mammalian cells, we produced two kinds of anti-VDAC antibodies that inhibited VDAC activity. In isolated mitochondria, these antibodies prevented Bax-induced cytochrome c release and loss of the mitochondrial membrane potential (Deltapsi), but not Bid-induced cytochrome c release. When microinjected into cells, these anti-VDAC antibodies, but not control antibodies, also prevented Bax-induced cytochrome c release and apoptosis, whereas the antibodies did not prevent Bid-induced apoptosis, indicating that the VDAC is essential for Bax-induced, but not Bid-induced, apoptogenic mitochondrial changes and apoptotic cell death. In addition, microinjection of these anti-VDAC antibodies significantly inhibited etoposide-, paclitaxel-, and staurosporine-induced apoptosis. Furthermore, we used these antibodies to show that Bax- and Bak-induced lysis of red blood cells was also mediated by the VDAC on plasma membrane. Taken together, our data provide evidence that the VDAC plays an essential role in apoptogenic cytochrome c release and apoptosis in mammalian cells.

Amino Acid Sequence↗
PubMed2001

Fixed-height exit bender of synchrotron X-rays above 40 keV.

A crystal bender for sagittal focusing has been designed for standard monochromators at SPring-8. The bender does not move the position of the crystal center when the bending radius is changed. Sagittal focusing from 40 keV to 60 keV was achieved by using Si(311) double crystals. The flux density of the focused beam measured at 40 keV was 15 times higher than that of the unfocused beam. The height deviation of the focused beam throughout the measured energy range was within +/- 0.15 mm.

Crystallization↗
PubMed2001

Decrease in level of APG-2, a member of the heat shock protein 110 family, in murine brain following systemic administration of kainic acid.

APG-2 belongs to the heat shock protein 110 family. Although kainic acid (KA)-induced seizures are known to elicit expression of inducible heat shock protein 70 (HSP70) in the brain, no investigation has been carried out on the APG-2 level after excitatory amino acid-induced seizures. By means of an immunoblot assay, we determined the levels of HSP70 and APG-2 in discrete brain structures of mice after a single intraperitoneal injection of KA or N-methyl-D-aspartic acid (NMDA). APG-2 level was significantly decreased in frontal cortex, hippocampus, and striatum three days after the administration of KA, while HSP70 level was increased in these regions following the administration. In any of these regions, APG-2 levels were returned to the control levels 10 days after the administration. However, no significant changes were observed in levels of both HSP70 and APG-2 in hypothalamus, midbrain, medulla-pons, and cerebellum of the mice. By contrast, NMDA administration did not significantly affect both levels in any of the regions examined. These findings indicate that the transient decrease in APG-2 expression is one of the intracellular events elicited by signals peculiar to KA, but not by those peculiar to NMDA, in telencephalon of murine brain.

Animals↗
PubMed2001

Characterization with [3H]quisqualate of group I metabotropic glutamate receptor subtype in rat central and peripheral excitable tissues.

Radioligand binding studies were performed to label metabotropic glutamate receptor (mGluR) in rat brain synaptic membranes using [3H]quisqualic acid (QA) synthesized in our laboratory as a radioligand. In the presence of ionotropic glutamate receptor (iGluR) agonists, including N-methyl-D-aspartic (NMDA), DL-alpha-amino-3-hydroxy-5-methylisoxasole-4-propionic (AMPA) and kainic acids (KA), at concentrations maximally effective in displacing each receptor binding, the agonists for group I mGluR subtype (+/-)-1-aminocyclopentane-trans-1,3-dicarboxylic acid (trans-ACPD) and (S)-3,5-dihydroxyphenylglycine ((S)-3,5-DHPG) more potently displaced [3H]QA binding in a concentration-dependent manner than their absence. The addition of these three iGluR agonists did not significantly affect potencies of (2S,2'R,3'R)-2-(2',3'-dicarboxycyclopropyl)glycine (DCG-IV) and L-(+)-2-amino-4-phosphonobutyric acid (L-AP4) to displace [3H]QA binding. Scatchard analysis revealed that [3H]QA binding consisted of a single component with a maximal number of binding sites (B(max)) of 431.6 fmol/mg protein and a dissociation constant (K(d)) of 50.9 nM, in the presence of the three iGluR agonists. [3H]QA binding was markedly inhibited by GTP and its analogues; but not by GDP, GMP and ATP, under these conditions. Inhibition by GTP was seen in all central structures examined, but [3H]QA binding was not detectable in peripheral tissues, such as pituitary and adrenal glands. Neither reverses transcription polymerase chain reaction nor immunoblotting analysis demonstrated the expression of mGluR1 and mGluR5 subunits in the aforementioned two peripheral tissues. These results suggest that [3H]QA indeed labels group I mGluR subtype functionally coupled to GTP binding protein in rat brain synaptic membranes under the experimental conditions employed. Group I mGluR subtype seems to be selectively distributed in central structures but not in pituitary and adrenal glands.

Adrenal Glands↗
PubMed2001

Expression of GluR6/7 subunits of kainate receptors in rat adenohypophysis.

We have previously demonstrated the presence of unidentified [3H]glutamate (Glu) binding sites with stereo-selectivity, high affinity and saturability in rat peripheral excitable tissues such as the pituitary (Yoneda, Y., Ogita, K., 1986a. [3H]Glutamate binding sites in the rat pituitary. Neurosci. Res. 3, 430--435) and adrenal (Yoneda, Y., Ogita, K., 1986b. Localization of [3H]glutamate binding sites in rat adrenal medulla. Brain Res. 383, 387--391, 1986). In this study, peripheral binding sites were further evaluated for the ionotropic Glu receptor subtype insensitive to N-methyl-D-aspartate by using reverse transcription polymerase chain reaction (RT-PCR) and Western blotting, in addition to receptor binding using radiolabeled ligands other than [3H]Glu. Binding of [3H]kainate (KA) and [3H]DL-alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate was detected in membrane preparations obtained from the rat pituitary and adrenal irrespective of prior treatment with Triton X-100. An RT-PCR analysis revealed constitutive expression of mRNA for GluR1, GluR3, GluR5, KA1 and KA2 subunits in the rat adrenal and pituitary, as well as the brain and retina. The pituitary also expressed mRNA for GluR2, GluR4, GluR6 and GluR7 subunits in contrast to the adrenal. Under our experimental conditions employed, however, Western blotting assays failed to confirm the expression of receptor proteins for GluR1, GluR2/3 and GluR4 subunits in the adrenal cortex, adrenal medulla, adenohypophysis and neurohypophysis. Immunoreactive GluR6/7 subunits were only detectable in the adenohypophysis, but not in the adrenal cortex, adrenal medulla and neurohypophysis. An intraperitoneal injection of KA doubled DNA binding activity of the nuclear transcription factor activator protein-1 in the rat pituitary, with concomitant more potent potentiation of that in the hippocampus. These results suggest that GluR6/7 subunits of KA receptors may be constitutively expressed with responsiveness to the systemic administration of an agonist at least in the rat adenohypophysis.

Animals↗
PubMed2001

Consolidation of transient ionotropic glutamate signals through nuclear transcription factors in the brain.

Long-lasting alterations of neuronal functions could involve mechanisms associated with consolidation of transient extracellular signals through modulation of de novo synthesis of particular functional proteins in the brain. In eukaryotes, protein de novo synthesis is mainly under the control at the level of gene transcription by transcription factors in the cell nucleus. Transcription factors are nuclear proteins with an ability to recognize particular core nucleotides at the upstream and/or downstream of target genes, and thereby to modulate the activity of RNA polymerase II that is responsible for the formation of mRNA from double stranded DNA. Gel retardation electrophoresis is widely employed for conventional detection of DNA binding activities of a variety of transcription factors with different protein motifs. Extracellular ionotropic glutamate (Glu) signals lead to rapid and selective potentiation of DNA binding of the nuclear transcription factor activator protein-1 (AP1) that is a homo- and heterodimeric complex between Jun and Fos family members, in addition to inducing expression of the corresponding proteins, in a manner unique to each Glu signal in murine hippocampus. Therefore, extracellular Glu signals may be differentially transduced into the nucleus to express AP1 with different assemblies between Jun and Fos family members, and thereby to modulate de novo synthesis of the individual target proteins at the level of gene transcription in the hippocampus. Such mechanisms may be operative on synaptic plasticity as well as delayed neuronal death through consolidation of alterations of a variety of cellular functions induced by transient extracellular signals in the brain.

Animals↗
PubMed2001

Schistosoma japonicum egg granuloma formation in the interleukin-4 or interferon-gamma deficient host.

The roles of interleukin (IL)-4 and interferon (IFN)-gamma in Schistosoma japonicum egg granuloma formation were investigated in cercariae-infected (infection model) or after implantation of laid parasite eggs (egg implantation model) in cytokine deficient mice. Two weeks after hepatic egg-implantation, a markedly decreased mononuclear cell infiltration and lack of multinuclear cell formation were characteristic features in IL-4 deficient mice. By 4 weeks (late stage), the cellular reactions around the eggs were negligible in the deficient mice. Compared to the controls, there was a drastic reduction in the production of the Th2 cytokines, IL-4, IL-5 and IL-13. MCP-1 levels were also significantly lowered. In mice experimentally infected with cercariae, granuloma cellularity in both the wild-type and IL-4 deficient mice at 45 days and 10 weeks postinfection was analogous to the egg implantation model at 2 and 4 weeks. Overall, the effects of IFN-gamma deficiency on granuloma induction differed markedly from the IL-4 results. Two weeks after egg implantation, IFN-gamma deficient mice showed suppressed neutrophil response and hepatic necrosis with confluent mononuclear cell infiltration along the outer layer of granulomas. By 4 weeks, there was a decrease in cell infiltration, fibrosis and MCP-1 production while IL-10 production increased. While these early characteristic features for IFN-gamma deficiency were common to both the egg implantation (at 2 and 4 weeks) and cercariae infection model (at 45 days), there was a surprising difference, i.e. marked fibrosis was found in the late stages (at 10 weeks postinfection) of cercariae-infected mice, but not in parasite egg implanted mice. Furthermore, while IL-13 levels were unchanged, both MCP-1 and IL-4 production were significantly lower at 10 weeks in comparison with wild-type. The present study clearly demonstrates the importance of both Th1 and Th2 cytokine responses in S. japonicum egg-induced granuloma formation.

Animals↗
PubMed2001

Referral and care for acute ischemic stroke in a Japanese tertiary emergency hospital.

To examine the current emergency referral and care for acute stroke at a Japanese tertiary emergency hospital with a 24-h stroke team and care unit, we surveyed the presentations of patients with acute ischemic stroke or transient ischemic attack (TIA) seen within 7 days of onset. Delay from symptom onset to arrival at our hospital, from arrival to initial diagnostic brain computed tomography (CT), and the type of anti-thrombotic treatments were evaluated. During the 18-month period, there were 254 ischemic events in 244 patients; 239 (94%) had an ischemic stroke and 15 (6%) TIA. Eighty-two (32%) events presented within 3 h of onset, and 102 (40%) and 179 (70%) within the first 6 and 24 h, respectively. The median delay from hospital arrival to CT was 32 min, ranging 10 min to 22 h. Two hundred (79%) events underwent CT within 1 h of arrival (n=172) or at the referral hospitals before transfer (n=28). Direct ambulance transportation and more severe neurological deficits were independent predictors both for early arrival and short in-hospital delay to CT. Anti-thrombotic therapies including anticoagulant and/or antiplatelet medications were given in 237 (93%) episodes. Two (1%) patients received thrombolysis, although 18 (7%) patients fulfilled the National Institute of Neurological Disorders and Stroke guidelines for intravenous thrombolysis with tissue plasminogen activator. As in western communities, our pre-hospital emergency referral systems for acute stroke require substantial improvements including the wider use of ambulance calling. Although our in-hospital stroke management is functioning relatively well, further efforts are necessary in reducing the diagnostic delay.

Acute Disease↗