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T Yagi

Publications and source records attributed to T Yagi.

At least 343 records · Page 19Linked to original sources
PubMed1995

Molecular cloning of a novel serine/threonine kinase, MRK, possibly involved in cardiac development.

We have isolated a novel member of putative serine/threonine kinase from a rat heart cDNA library using polymerase chain reaction methods. The novel kinase is transcribed as 2.6 kb mRNA encoding for a protein of 629 amino acids with the C-terminal non-catalytic portion. Amino acids analysis revealed that the N-terminal catalytic domain is 87% identical to the male-germ cell associated kinase (MAK), a cdc2-related serine/threonine kinase found to promote meiosis during spermatogenesis. Therefore, we designated this novel kinase as the MAK-related kinase (MRK). MRK protein, with a molecular weight of 66 kD, was shown to phosphorylate itself and the exogenous substrates, histone H1 and myelin basic protein. In addition, phosphoamino acid analysis confirmed the serine/threonine-specific protein kinase activity of MRK. Although MRK was ubiquitous in adult rat tissues, the expression of MRK protein in embryos was restricted primarily to embryonic myocardium during early organogenesis. This finding suggests that MRK may be a participant in cardiac development.

Amino Acid Sequence↗
PubMed1995

The effects of nutritional repletion on donors for liver transplantation in pigs.

It has been shown that the nutritional state of the donor may affect the outcome of liver transplantation. However, many donors staying in the intensive care unit for a long period are in a reduced nutritional state. In this study, we investigated the effects of various methods of nutritional repletion on the outcome of liver transplantation in pigs. Donor pigs were divided into three groups according to the nutritional pretreatment given for 7 days before harvesting: group I were fasted and received intravenous administration of saline; group II were fed orally; group III were fasted, but given 20% glucose intravenously. Donor livers were stored for 4 hr in cold Euro-Collins' solution and transplanted. The serum AST level 24 hr after reperfusion remained at a lower level in group III compared with those in groups I and II. Bile production of the liver after transplantation was also well recovered in group III. The glycogen content of the liver at harvesting, which was completely consumed in group 1, was well preserved in groups II and III. These storages in both groups were rapidly consumed 1 hr after reperfusion. On the other hand, ATP content of the liver in groups I, II, and III, which were at a similar level at harvesting, were markedly decreased 4 hr after cold preservation and, 1 hr after reperfusion, recovered to 26%, 48%, and 73% of that before preservation, respectively. The mean survival time in group III was 37.2 days, significantly longer than 5.8 +/- 0.7 and 9.8 +/- 2.0 days in groups I and II, respectively (P < 0.01). These results show that the favorable outcome of liver transplantation depends on the glycogen storage in the donor liver, and also on ATP generation after reperfusion. Furthermore, it was suggested that ATP generation was affected by some unknown factor related to the method of nutritional repletion.

Adenosine Triphosphate↗
PubMed1995

Stable production of mutant mice from double gene converted ES cells with puromycin and neomycin.

The antibiotic puromycin is an effective inhibitor of protein synthesis and puromycin N-acetyl transferase gene could be used as a dominant selection marker. We report the effective production of mutant mice from double gene-converted ES cells by selection with G418 and puromycin. We confirmed that (i) puromycin efficiently inhibited the growth of ES cells at a low-dose (0.1 microgram/ml) and for a short time (2 days), independent of G418 selection; (ii) when these selected ES cells were injected into eight-cell stage embryos, the cells produced chimeras with high levels of chimerism; (iii) these chimeric males were fertile and exclusively yielded ES cell-derived offspring; and (iv) each offspring contained both neomycin transferase and puromycin N-acetyl transferase genes.

Acetyltransferases↗
PubMed1995

Expression and characterization of the 66-kilodalton (NQO3) iron-sulfur subunit of the proton-translocating NADH-quinone oxidoreductase of Paracoccus denitrificans.

The proton-translocating NADH-quinone oxidoreductase (NDH-1) of Paracoccus denitrificans is composed of at least 14 dissimilar subunits which are designated NQO1-14 and contains one noncovalently bound FMN and at least five EPR-visible iron-sulfur clusters (N1a, N1b, N2, N3, and N4) as prosthetic groups. Comparison of the deduced primary structures of the subunits with consensus sequences for the cofactor binding sites has predicted that NQO1, NQO2, NQO3, NQO9, and probably NQO6 subunits are cofactor binding subunits. Previously, we have reported that the NQO2 (25 kDa) subunit was overexpressed as a water-soluble protein in Escherichia coli and was found to ligate a single [2Fe-2S] cluster with rhombic symmetry (gx,y,z = 1.92, 1.95, and 2.00) (Yano, T., Sled', V.D., Ohnishi, T., and Yagi, T. (1994) Biochemistry 33, 494-499). In the present study, the NQO3 (66 kDa) subunit, which is equivalent to the 75-kDa subunit of bovine heart Complex I, was overexpressed in E. coli. The expressed NQO3 subunit was found predominantly in the cytoplasmic phase and was purified by ammonium sulfate fractionation and anion-exchange chromatography. The chemical analyses and UV-visible and EPR spectroscopic studies showed that the expressed NQO3 subunit contains at least two distinct iron-sulfur clusters: a [2Fe-2S] cluster with axial EPR signals (g perpendicular, parallel = 1.934 and 2.026, and L perpendicular parallel = 1.8 and 3.0 millitesla) and a [4Fe-4S] cluster with rhombic symmetry (gx,y,z = 1.892, 1.928, and 2.063, and Lx,y,z = 2.40, 1.55, and 1.75 millitesla). The midpoint redox potentials of [2Fe-2S] and [4Fe-4S] clusters at pH 8.6 are -472 and -391 mV, respectively. The tetranuclear cluster in the isolated NQO3 subunit is sensitive toward oxidants and converts into [3Fe-4S] form. The assignment of these iron-sulfur clusters to those identified in the P. denitrificans NDH-1 enzyme complex and the possible functional role of the NQO3 subunit is discussed.

Base Sequence↗
PubMed1995

Superoxide generation by alveolar macrophages from aged rats: improvement by in vitro treatment with IFN-gamma.

Alveolar macrophages (AM) from aged rats show an impaired oxidative response, but it is unclear whether or not this is due to the inability of these cells to be activated. To elucidate this, we investigated the capacity of AM from young (16-week-old) and aged (100-week-old) rats to become primed with recombinant rat interferon-gamma (IFN-gamma) for increased phorbol myristate acetate (PMA)-elicited O2- production, utilizing an MCLA-dependent chemiluminescent assay. We also compared concanavalin A- or Bacillus Calmette Guerin (BCG)-induced IFN-gamma production by the spleen cells of young and aged animals. The data indicated that AM freshly harvested from non-sensitized aged rats produced less O2- than those from young animals. A similar result was obtained in BCG-sensitized rats. However, AM from aged rats were primed with in vitro treatment with IFN-gamma for increased rate of O2- production to an equivalent level of that by AM from young animals. In addition, the ability of spleen cells to produce IFN-gamma was well maintained in aged rats. These results suggest that AM function is suppressed in the lungs of aged animals. Our observation that the decreased AM function in aged rats can be reversed is important because it suggests that appropriate treatment may reduce the incidence and mortality of respiratory infections in the elderly.

Aging↗
PubMed1995

EPR and redox characterization of ferredoxins I and II from Desulfovibrio vulgaris Miyazaki.

Detailed redox titrations monitored by EPR and UV-visible spectroscopies have been carried out on the dimeric ferredoxins I and II from Desulfovibrio vulgaris Miyazaki. Ferredoxin II contains a unique [4Fe-4S] cluster per subunit characterized by a midpoint potential of -417 mV at 24 degrees C. The enthalpic and entropic contributions to the redox free energy variation of this cluster have been determined from the temperature dependence of the midpoint potential and compared to the data reported for other iron-sulfur proteins. The molecular arrangement of the two subunits is such that two [4Fe-4S]i+ clusters are magnetically coupled in the fully reduced state of the protein. Ferredoxin I contains one [3Fe-4S] and one [4Fe-4S] cluster per subunit, whose spectral and redox properties are very similar to those of the same clusters in ferredoxin III from Desulfovibrio africanus. The strong heterogeneity in the redox properties of the [3Fe-4S] center supports a bridging position between the N-terminal and C-terminal parts of the protein.

Desulfovibrio vulgaris↗
PubMed1995

Impairment of motor coordination, Purkinje cell synapse formation, and cerebellar long-term depression in GluR delta 2 mutant mice.

Of the six glutamate receptor (GluR) channel subunit families identified by molecular cloning, five have been shown to constitute either the AMPA, kainate, or NMDA receptor channel, whereas the function of the delta subunit family remains unknown. The selective localization of the delta 2 subunit of the GluR delta subfamily in cerebellar Purkinje cells prompted us to examine its possible physiological roles by the gene targeting technique. Analyses of the GluR delta 2 mutant mice reveal that the delta 2 subunit plays important roles in motor coordination, formation of parallel fiber-Purkinje cell synapses and climbing fiber-Purkinje cell synapses, and long-term depression of parallel fiber-Purkinje cell synaptic transmission. These results suggest a close relationship between synaptic plasticity and synapse formation in the cerebellum.

Animals↗
PubMed1995

Reduced hippocampal LTP and spatial learning in mice lacking NMDA receptor epsilon 1 subunit.

The NMDA (N-methyl-D-aspartate) receptor channel is important for synaptic plasticity, which is thought to underlie learning, memory and development. The NMDA receptor channel is formed by at least two members of the glutamate receptor (GluR) channel subunit families, the GluR epsilon (NR2) and GluR zeta (NR1) subunit families. The four epsilon subunits are distinct in distribution, properties and regulation. On the basis of the Mg2+ sensitivity and expression patterns, we have proposed that the epsilon 1 (NR2A) and epsilon 2 (NR2B) subunits play a role in synaptic plasticity. Here we show that targeted disruption of the mouse epsilon 1 subunit gene resulted in significant reduction of the NMDA receptor channel current and long-term potentiation at the hippocampal CA1 synapses. The mutant mice also showed a moderate deficiency in spatial learning. These results support the notion that the NMDA receptor channel-dependent synaptic plasticity is the cellular basis of certain forms of learning.

Animals↗
PubMed1995

Novel mode of hyper-oscillation in the paralyzed axoneme of a Chlamydomonas mutant lacking the central-pair microtubules.

The flagellar axoneme of the mutant pf18 lacking the central pair does not beat, but undergoes a nanometer-scale, high-frequency oscillation (hyper-oscillation) in the presence of ATP [Yagi et al., 1994: Cell Motil. Cytoskeleton 29:177-185]. The present study demonstrates that the amplitude of the hyper-oscillation increases significantly in the simultaneous presence of ATP and ADP. In addition, the hyper-oscillation under these conditions sometimes takes on an exceptionally simple asymmetric pattern, in which the maximal shearing velocity exceeds 50 microns/sec, much higher than the maximal velocity of ordinary dynein-microtubule sliding. The asymmetric oscillation thus appears to be at least partly driven by an internal elastic force. Its amplitude suggests that the axoneme has an elastic component that can be stretched by as long as 0.1 micron. Analyses of the asymmetric pattern further suggests that the axonemal dyneins have a tendency to attach to and detach from the doublets cooperatively and that the mechanochemical cycle of dynein has an inherent refractory period of about 2 msec, during which dynein cannot interact with microtubules.

Adenosine Diphosphate↗
PubMed1995

A new strategy of gene trapping in ES cells using 3'RACE.

"Gene trapping" in embryonic stem (ES) cells is a novel approach to identify a series of genes in mammals concomitant with the production of the corresponding mutant mice. However, this approach is currently unable to identify genes that are not expressed in ES cells. Here we describe a strategy to identify gene trapping clones which is not based on expression of a reporter gene. It uses the neor gene which lacks a polyadenylation signal and has a splice donor signal. Expression of the neor gene as fusion transcripts with the 3' end containing the polyadenylation signal of tagged genes allows the identification of these clones by 3' rapid amplification of the cDNA end in undifferentiated ES cells, even if the genes are not expressed in ES cells. Amplification was observed in about 25% of G418-resistant clones. Sequence analyses suggested the amplifications represent gene trapping events. The feasibility of this approach was further assessed by analysing one clone, PAT-12, in detail.

Amino Acid Sequence↗
PubMed1995

Computational studies on the interaction between red cone and H1 horizontal cell.

We propose an equivalent circuit model of a discrete formulation to describe the interaction between the red cone syncytium and the H1 horizontal cell syncytium in lower vertebrate retinas. Analytical solutions of the model provide intuitive understandings of spatio-temporal properties of light-induced responses in reference to membrane impedance, strength of chemical synapse and coupling resistance connecting neighbouring cells. Physiologically plausible values of these parameters are estimated using the solutions. Quantitative studies are made to elucidate the function of (1) the negative feedback from the H1 horizontal cell to the red cone, and (2) the resistance increase of H1 horizontal cell coupling by dopamine.

Animals↗
PubMed1995

Enhanced susceptibility of audiogenic seizures in Fyn-kinase deficient mice.

Mice with a mutation in fyn genes were examined for their susceptibility to acoustically primed audiogenic seizures. Homozygous mutant (fynz/fynz) mice were significantly more likely to have seizures and to show the stronger seizure syndrome (clonus). These results indicate that the susceptibility of acoustically primed audiogenic seizures is enhanced in the Fyn kinase deficient mice.

Animals↗