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Y Ohshima

Publications and source records attributed to Y Ohshima.

At least 73 records · Page 4Linked to original sources
PubMed1997

The UNC-14 protein required for axonal elongation and guidance in Caenorhabditis elegans interacts with the serine/threonine kinase UNC-51.

Certain unc mutants in the nematode Caenorhabditis elegans, such as unc-14 and unc-51, show abnormal axonal elongation and axonal structures. We cloned the unc-51 gene previously and predicted that it encodes a novel serine/threonine protein kinase. In this study, we precisely localized the activity to rescue an unc-14 mutation. Also, we identified four cDNA clones encoded by the unc-14 rescuing region, in screens for proteins that bind to UNC-51 using a yeast two-hybrid system. A mutation site in the cDNA was identified for each of the six unc-14 mutants, establishing that the unc-14 gene was cloned. The unc-14 gene encodes a novel protein of 665 amino acids, and is coexpressed with the unc-51 gene in the cell bodies and axons of almost all neurons including DD/VD and hermaphrodite-specific neurons. Another clone recovered in the two-hybrid screen encodes a carboxy-terminal region of UNC-51. Analysis using the yeast two-hybrid system suggested that a central region of UNC-14 bound to a carboxy-terminal region of UNC-51, and that the UNC-51 carboxy-terminal region oligomerized. In in vitro binding studies using recombinant fusion proteins, UNC-14 interacted with UNC-51 directly. We propose that UNC-51 protein kinase acts as an oligomer, and that UNC-14 is a regulator of UNC-51, in axonal elongation and guidance.

Amino Acid Sequence↗
PubMed1997

An activating mutation in a Caenorhabditis elegans Gs protein induces neural degeneration.

Heterotrimeric guanine nucleotide-binding proteins (G proteins) act as signal-transducing molecules that connect serpentine-transmembrane receptors to a variety of intracellular effectors. We characterized a Caenorhabditis elegans G(s) gene, gsa-1, which encodes a G(s) alpha-subunit (G alpha(s)) that is expressed throughout the nervous system and in muscle cells. gsa-1 is an essential gene; a loss-of-function mutation in gsa-1 results in lethality at the first stage of larval development. Partial (mosaic) loss of G alpha(s) expression or overexpression of the protein results in reciprocal defects in movement and egg-laying, suggesting a role for G alpha(s) in the regulation of these behaviors. Expression of a constitutively active form of G alpha(s) from an inducible promotor results in hypercontraction of body-wall muscle cells and vacuolization and degeneration of neurons within hours of induction. Neurons that are susceptible to the degeneration induced by activated G alpha(s) are predominantly motoneurons located within the ventral nerve cord. Phenotypic analysis shows that the induced neural degeneration is not the result of programmed cell death but is probably caused by the activation of ion channels. A genetic suppressor of activated G alpha(s) was isolated that identifies a putative downstream target of G(s) signaling.

Animals↗
PubMed1997

T cell-derived IL-4 and dendritic cell-derived IL-12 regulate the lymphokine-producing phenotype of alloantigen-primed naive human CD4 T cells.

Previous studies on human Th subset development were restricted to the analysis of naive T cells activated with anti-CD3 mAb in the absence of physiologic APC. In this study, we have analyzed the role of cytokines and physiologic APC on T cell maturation in an Ag-specific system, in which naive neonatal CD4 T cells were primed with allogeneic dendritic cells (DC). We found that the cytokine profile of primed cells was dependent upon 1) the ratio between T cells and allogeneic DC and 2) the endogenous production of IL-4 and IL-12. Neutralization of IL-4 during primary MLR increased IFN-gamma production at priming and shifted the phenotype of primed cells from Th0 to Th1. These effects were IL-12 dependent, in that they were suppressed by anti-IL-12 Abs. The production of IL-12 in primary MLR was further evidenced by the presence of IL-12 p40 in the culture supernatant fluids. IL-12 production was suppressed by exogenous IL-4 and increased by anti-IL-4 blocking mAbs, indicating that endogenous IL-4 down-regulated IL-12 production by DC. Finally, IL-12 was produced as a result of T cell/DC interaction involving the CD40/CD40 ligand and CD28/B7 costimulation pathways, as revealed by the inhibitory effect of anti-CD40 ligand mAb and CTLA-4Ig. These observations suggest that in neutral conditions, Ag presentation by DC results in the coordinate production of naive T cell-derived IL-4 and DC-derived IL-12 that in concert shape the cytokine profile of Th cells.

CD4-Positive T-Lymphocytes↗
PubMed1997

Molecular neurogenetics of chemotaxis and thermotaxis in the nematode Caenorhabditis elegans.

Chemotaxis and thermotaxis in Caenorhabditis elegans are based on the chemical senses (smell and taste) and the thermal sense, respectively, which are important for the life of the animal. Laser ablation experiments have allowed identification of sensory neurons and some interneurons required for these senses. Many mutants that exhibit various abnormalities have been isolated and analyzed. These studies have predicted novel signaling pathways whose components include a putative odorant specific transmembrane receptor (ODR-10) and a cyclic nucleotide-gated channel (TAX-4/TAX-2) functioning in taste and thermosensation as well as in smell. The emerging picture of the mechanisms of sensory transduction in C. elegans seems to be basically similar to what is known of visual and olfactory sensory transduction in vertebrates. Thus, molecular and cellular analyses of chemotaxis and thermotaxis in C. elegans have proved useful and will continue to provide significant implications for the molecular basis of sensory systems in higher animals.

Animals↗
PubMed1997

Fourier-Transform Microwave Spectroscopy of the Argon-Diacetylene van der Waals Complex

The rotational spectrum of the argon-diacetylene van der Waals complex, produced in a supersonic molecular beam at 1 K, has been observed with a Fourier-transform microwave spectrometer. We observed 22 a-type rotational transitions with Ka up to 3. Three rotational constants, five centrifugal distortion constants, and one higher-order centrifugal distortion constant were determined precisely by least-squares analysis. The complex is shown to have a planer T-shaped structure with C2v symmetry. The structural analysis provides that the Ar atom is located 3.68 A from the center of mass of diacetylene. Force constants for the van der Waals vibrations were determined from the centrifugal distortion constants. It has been found that this complex has a much steeper and more harmonic intermolecular potential than the argon-acetylene complex. Copyright 1997 Academic Press. Copyright 1997Academic Press

Journal Article↗
PubMed1997

5-Hydroxytryptamine-induced Ca2+ -independent cGMP formation is mediated by nitric oxide in a nitric oxide synthase-independent manner in NG108-15 cells.

A novel pathway of 5-hydroxytryptamine (5-HT)-induced cGMP formation, which does not require Ca2+ and is distinct from the 5-HT, receptor-mediated pathway, is reported to exist in NG108-15 cells. Although the possible involvement of undefined 5-HT receptors and membrane-bound guanylyl cyclase is suggested. the mechanism is not clarified in detail in this Ca2+ -independent cGMP formation. In the present study, we investigated the activation mechanism of guanylyl cyclase activity. 5-HT-induced Ca2+ -independent cGMP formation was not observed in the cell membrane preparation. In intact cells, the 5-HT-induced Ca2+ -independent cGMP formation was inhibited by 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazolin-1-oxyl 3-oxide (carboxyPTIO), a nitric oxide (NO)-specific trapper, and by 3,7-bis(dimethylamino)-phenothiazinium chloride (methylene blue), a cytosolic guanylyl cyclase-specific inhibitor, suggesting the involvement of NO and cytosolic guanylyl cyclase. Ca2+ -independent cGMP formation was not inhibited by 1,2-bis(o-aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid tetra(acetoxymethyl)ester (BAPTA-AM), N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide (W-7), and various arginine-derivative nitric oxide synthase (NOS) inhibitors. Our findings suggest that 5-HT stimulation results in the generation of NO followed by cGMP formation in an NOS-independent manner in NG108-15 cells.

Animals↗
PubMed1997

Regulation of interneuron function in the C. elegans thermoregulatory pathway by the ttx-3 LIM homeobox gene.

Neural pathways, which couple temperature-sensing neurons to motor and autonomic outputs, allow animals to navigate away from and adjust metabolism rates in response to the temperature extremes often encountered. ttx-3 is required for the specification of the AIY interneuron in the C. elegans neural pathway that mediates thermoregulation. ttx-3 null mutant animals exhibit the same thermotactic behavioral defect as that seen with laser ablation of AIY in wild type, suggesting that AIY does not signal in this mutant. ttx-3 encodes a LIM homeodomain protein. A ttx-3-GFP fusion gene is expressed specifically in the adult AIY interneuron pair, which connects to thermosensory neurons. In ttx-3 mutant animals, the AIY interneuron is generated but exhibits patterns of abnormal axonal outgrowth. Thus, the TTX-3 LIM homeodomain protein is likely to regulate the expression of target genes required late in AIY differentiation for the function of this interneuron in the thermoregulatory pathway. The ttx-3-dependent thermosensory pathway also couples to the temperature-modulated dauer neuroendocrine signaling pathway, showing that ttx-3 specifies AIY thermosensory information processing of both motor and autonomic outputs.

Amino Acid Sequence↗
PubMed1997

Some biochemical properties of the components of Staphylococcus aureus binding to human platelets.

The binding properties of Staphylococcus aureus in relation to human platelets were investigated. Protease digestion (pronase E, proteinase K, trypsin), heat treatment (80 degrees C, 30 min), and sonication for 5 min significantly reduced the binding abilities of the staphylococcal cells to 0% (p < .01), 50 +/- 5% (p < .05), and 38 +/- 9% (p < .05), respectively, while mixed glycosidases did not. Inhibition experiments indicated that protein A and various sugars were ineffective. A binding study using biotinylated cell surface fractions extracted from the whole cells of S. aureus indicated that the proteins having apparent molecular weights of 14400 and 16500 estimated by sodium dodecylsulfate-polyacrylamide gel electrophoresis were involved in the binding between S. aureus and human platelets.

Bacterial Adhesion↗
PubMed1997

Isolation and molecular characterization of mRNA transport mutants in Schizosaccharomyces pombe.

Nucleocytoplasmic transport of mRNA is essential for eukaryotic gene expression. However, how mRNA is exported from the nucleus is mostly unknown. To elucidate the mechanisms of mRNA transport, we took a genetic approach to identify genes, the products of which play a role in that process. From about 1000 temperature -sensitive (ts- or cs-) mutants, we identified five ts- mutants that are defective in poly(A)+ RNA transport by using a situ hybridization with an oligo(dT)50 as a probe. These mutants accumulate poly(A)+ RNA in the nuclei when shifted to a nonpermissive temperature. All five mutations are tightly linked to the ts- growth defects, are recessive, and fall into four different groups designated as ptr 1-4 (poly(A)+ RNA transport). Interestingly, each group of mutants has a differential localization pattern of poly(A)+ RNA in the nuclei at the nonpermissive temperature, suggesting that they have defects at different steps of the mRNA transport pathway. Localization of a nucleoplasmin-green fluorescent protein fusion suggests that ptr2 and ptr3 have defects also in nuclear protein import. Among the isolated mutants, only ptr2 showed a defect in pre-mRNA splicing. We cloned the ptr2+ and ptr3+ genes and found that they encode Schizosaccharomyces pombe homologues of the mammalian RCC1, a guanine nucleotide exchange factor for RAN/TC4, and the ubiquitin-activating enzyme E1 involved in ubiquitin conjugation, respectively. The ptr3+ gene is essential for cell viability, and Ptr3p tagged with green fluorescent protein was localized in both the nucleus and the cytoplasm. This is the first report suggesting that the ubiquitin system plays a role in mRNA export.

Amino Acid Sequence↗
PubMed1997

The prp1+ gene required for pre-mRNA splicing in Schizosaccharomyces pombe encodes a protein that contains TPR motifs and is similar to Prp6p of budding yeast.

The prp (pre-mRNA processing) mutants of the fission yeast Schizosaccharomyces pombe have a defect in pre-mRNA splicing and accumulate mRNA precursors at a restrictive temperature. One of the prp mutants, prp1-4, also has a defect in poly(A)+ RNA transport. The prp1+ gene encodes a protein of 906 amino acid residues that contains 19 repeats of 34 amino acids termed tetratrico peptide repeat (TPR) motifs, which were proposed to mediate protein-protein interactions. The amino acid sequence of Prp1p shares 29.6% identity and 50.6% similarity with that of the PRP6 protein of Saccharomyces cerevisiae, which is a component of the U4/U6 snRNP required for spliceosome assembly. No functional complementation was observed between S. pombe prp1+ and S. cerevisiae PRP6. We examined synthetic lethality of prp1-4 with the other known prp mutations in S. pombe. The results suggest that Prp1p interacts either physically or functionally with Prp4p, Prp6p and Prp13p. Interestingly, the prp1+ gene was found to be identical with the zer1+ gene that functions in cell cycle control. These results suggest that Prp1p/Zer1p is either directly or indirectly involved in cell cycle progression and/or poly(A)+ RNA nuclear export, in addition to pre-mRNA splicing.

Alleles↗
PubMed1997

[Mitral valve replacement and closure of ventricular septal defect in the first two months of life].

A 2-month-old female infant weighing 4.2 kg was admitted with severe congestive heart failure and respiratory distress. The patient was operated upon under a diagnosis of severe congenital mitral regurgitation and ventricular septal defect. There was thickening of the anterior leaflet and short thickened chordae. The valve was judged not amenable to repair and it was replaced with a 17 mm St. Jude Medical valve. She was extubated on the 5th postoperative day. Post operative anti-coagulant therapy was initiated with warfarin potassium, dipyridamole and ticlopidine hydrochloride. The postoperative course was uneventful and the patient is doing well 9 months after the operation.

Female↗
PubMed1997

[Hemodynamic effects of amrinone in children during cardiopulmonary bypass and postoperative 12 hours].

Hemodynamic effects of amrinone in children during cardiopulmonary bypass (CPB) and postoperative 12 hours were studied. In 10 patients undergoing open heart surgery, 1 mg/kg of amrinone was infused as the initial CPB dose and 5 - 10 micrograms/kg/min of amrinone was continuously administered as the maintenance dose during CPB and postoperative 12 hours. Amrinone levels ranged from 0.9 to 1.4 micrograms/ml during CPB and postoperative 12 hours. After infusion of amrinone, mean arterial blood pressure decreased significantly, but other parameters did not show remarkable change. The administration of amrinone during CPB showed enough vasodilating effect and decreased the need of conventionally used other vasodilators (nitroglycerin or prostaglandin E1). The postoperative course of 10 patients was clinically uneventful. The administration of amrinone in 10 patients did not produce thrombocytopenia compared with the control group (5 patients) in the postoperative period. In conclusion, the administration of amrinone during CPB and postoperative 12 hours in children was useful in producing enough vasodilating effect without major side effect.

Amrinone↗
PubMed1996

Isolation of novel pre-mRNA splicing mutants of Schizosaccharomyces pombe.

New prp (pre-mRNA processing) mutants of the fission yeast Schizosaccharomyces pombe were isolated from a bank of 700 mutants that were either temperature sensitive (ts-) or cold sensitive (cs-) for growth. The bank was screened by Northern blot analysis with probes complementary to S. pombe U6 small nuclear RNA (sn RNA), the gene for which has a splicesomal (mRNA-type) intron. We identified 12 prp mutants that accumulated the U6 snRNA precursor at the nonpermissive temperature. All such mutants were also found to have defects in an early step of TFIID pre-mRNA splicing at the nonpermissive temperature. Complementation analyses showed that seven of the mutants belong to six new complementation groups designated as prp8 and prp10-prp14, whereas the five other mutants were classified into the known complementation groups prp1, prp2 and prp3. Interestingly, some of the isolated prp mutants produced elongated cells at the nonpermissive temperature, which is a phenotype typical of cell division cycle (cdc) mutants. Based on these findings, we propose that some of the wild-type products from these prp+ genes play important roles in the cellular processes of pre-mRNA splicing and cell cycle progression.

Cell Cycle↗
PubMed1996

Cyclic GMP elevation by 5-hydroxytryptamine is due to nitric oxide derived from endogenous nitrosothiol in NG108-15 cells.

To clarify the involvement of nitric oxide (NO) derived from nitrosothiols (RSNO) in 5-hydroxytryptamine (5-HT)-induced Ca(2+)-independent cGMP formation (CIGF) in NG108-15 cells, we investigated the effects of 5-HT on intracellular contents of RSNO as well as of NO metabolites. 5-HT stimulation resulted in an increase in the intracellular contents of nitrate and cGMP. RSNO was detected in NG108-15 cells and was decreased by 5-HT stimulation. Furthermore, the time course of nitrate increase was coincident with that of RSNO decrease. CarboxyPTIO inhibited 5-HT-induced CIGF, whereas oxyhemoglobin failed to inhibit it. The data suggest that NO is stored in a stable form as RSNO and that 5-HT stimulates NO generation from endogenous RSNO, which is followed by elevation of cGMP via activation of cytosolic guanylyl cyclase by NO in NG108-15 cells. We suggest the existence of a novel 5-HT signal transduction pathway involved in NO generation in NG108-15 cells.

Animals↗
PubMed1996

Genomic structure and 5' regulatory regions of the let-23 gene in the nematode C. elegans.

The let-23 gene in the nematode Caenorhabditis elegans encodes a receptor tyrosine kinase and is necessary for the induction of a vulva, survival past the L1 stage, hermaphrodite fertility and for male spicule development. We sequenced the entire let-23 genomic region of over 12 kb. The 5' end of the let-23 mRNA was mapped by sequencing polymerase chain reaction products, and two mRNAs were found which had alternative exons and were probably initiated at different sites. One transcript was trans-spliced to SL1. Expression of either cDNA rescued a let-23 vulvaless mutation in germline transformation. These results suggest that the let-23 gene encodes two proteins of 1323 or 1335 amino acid residues. We prepared various 5' deletion constructs of the let-23 gene, and examined their rescue activities for a let-23 lethal or vulvaless mutation. The results revealed that two regions were required for let-23 expression, one for larval survival and the other for vulva formation. We also cloned and analyzed a let-23 homologue from Caenorhabditis vulgaris. It can encode two proteins of 77% amino acid residue identity with the Let-23 proteins. The 12 kb fragment carrying the C. vulgaris gene rescued the let-23 vulvaless mutation in C. elegans. Seventeen sequences highly conserved between the two species were identified in the 5' upstream region or within an intron. Three of these sequences are contained in the two regions required for let-23 expression, suggesting that they are cis-acting elements for let-23 expression.

Amino Acid Sequence↗
PubMed1996

Mutations in a cyclic nucleotide-gated channel lead to abnormal thermosensation and chemosensation in C. elegans.

The C. elegans tax-4 mutants are abnormal in multiple sensory behaviors: they fail to respond to temperature or to water-soluble or volatile chemical attractants. We show that the predicted tax-4 gene product is highly homologous to vertebrate cyclic nucleotide-gated channels. Tax-4 protein expressed in cultured cells functions as a cyclic nucleotide-gated channel. The green fluorescent protein (GFP)-tagged functional Tax-4 protein is expressed in thermosensory, gustatory, and olfactory neurons mediating all the sensory behaviors affected by the tax-4 mutations. The Tax-4::GFP fusion is partly localized at the sensory endings of these neurons. The results suggest that a cyclic nucleotide-gated channel is required for thermosensation and chemosensation and that cGMP is an important intracellular messenger in C. elegans sensory transduction.

Amino Acid Sequence↗
PubMed1996

A connection between pre-mRNA splicing and the cell cycle in fission yeast: cdc28+ is allelic with prp8+ and encodes an RNA-dependent ATPase/helicase.

The fission-yeast gene cdc28+ was originally identified in a screen for temperature-sensitive mutants that exhibit a cell-division cycle arrest and was found to be required for mitosis. We undertook a study of this gene to understand more fully the general requirements for entry into mitosis. Cells carrying the conditional lethal cdc28-P8 mutation divide once and arrest in G2 after being shifted to the restrictive temperature. We cloned the cdc28+ gene by complementation of the temperature-sensitive growth arrest in cdc28-P8. DNA sequence analysis indicated that cdc28+ encodes a member of the DEAH-box family of putative RNA-dependent ATPases or helicases. The Cdc28 protein is most similar to the Prp2, Prp16, and Prp22 proteins from budding yeast, which are required for the splicing of mRNA precursors. Consistent with this similarity, the cdc28-P8 mutant accumulates unspliced precursors at the restrictive temperature. Independently, we isolated a temperature-sensitive pre-mRNA splicing mutant prp8-1 that exhibits a cell-cycle phenotype identical to that of cdc28-P8. We have shown that cdc28 and prp8 are allelic. These results suggest a connection between pre-mRNA splicing and progression through the cell cycle.

Adenosine Triphosphatases↗