Chromosome engineering in yeast with a site-specific recombination system from a heterologous yeast plasmid.
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Chromosome Aberrations↗
Biomedical subjects
Y Oshima
Publications and source records attributed to Y Oshima.
Two new genes, PHO86 and PHO87, involved in inorganic phosphate uptake in Saccharomyces cerevisiae.
The PHO84 gene in Saccharomyces cerevisiae encodes a Pi transporter, mutation of which confers constitutive synthesis of repressible acid phosphatase (rAPase), in medium containing repressible amounts of Pi, and an arsenate-resistant phenotype. We selected an arsenate-resistant mutant showing the constitutive synthesis of rAPase on nutrient plates containing 4.5 mM arsenate. This mutant has double mutations designated as pho86 and pho87. The mutant transcribes PHO84 even in the repressible condition but has a severe defect in Pi uptake. The constitutive rAPase+ phenotype of the pho86 pho87 mutant was partially suppressed by an increased dosage of the PHO84 gene. The PHO87 gene was found to be identical with YCR524, according to the published nucleotide sequence of chromosome III, which encodes a protein of 923 amino-acid residues with a highly charged N-terminal half followed by a C-terminal half consisting of 12 membrane-spanning segments as in Pho84p. These and the other findings suggest that the Pho86p and Pho87p proteins collaborate with Pho84p in Pi uptake.
Acid Phosphatase↗
[Inhibitory effect of vitamin E succinate on the proliferation of cultured bovine choroidal endothelial cells].
We report the effect of vitamin E succinate (VE succinate) on the proliferation of cultured bovine choroidal endothelial cells (BCECs). BCECs were incubated with a medium containing vitamin E (VE) or one of the VE derivatives gamma-tocopherol, VE phosphate, VE succinate, VE nicotinate, VE acetate, or trolox, at a concentration of 10 microM. The proliferation of BCECs was assessed by 3H-thymidine uptake and cell counting. Especially in VE and VE succinate, the proliferation assay was performed on BCECs at two different stages, that is, the proliferating stage and the quiescent stage. The effect of protein kinase C (PKC) stimulator phorbol ester (PMA) on the VE succinate-induced inhibition of BCEC proliferation was also examined. VE succinate was found to significantly inhibit BCEC proliferation at a concentration of 10 microM or greater both by 3H-thymidine uptake assay and by cell counting. This inhibitory effect was not noted in other VE derivatives. The inhibitory effect was the most prominent in the proliferating BCECs and co culture of PMA. VE succinate inhibits the proliferation of cultured BCECs and PKC is involved in this action at least in part.
Animals↗
[Bronchiolitis obliterans organizing pneumonia (BOOP) after allogeneic bone marrow transplantation].
We report a patient who underwent allogeneic bone marrow transplantation for ALL. The patient presented low grade fever, cough and dyspnea at day 3 after after bone marrow transplantation. Imaging studies showed bilateral patchy infiltrates on chest X-ray and chest CT. Though treated by antibiotics, chest auscultation recognized marked bilateral inspiratory crackles. Transbronchial lung biopsy performed on day 34 showed bronchiolitis obliterans and an organizing interstitial pneumonia. The patient was treated with methylprednisolone and follow up computed tomography findings improved.
Administration, Oral↗
Structure and distribution of specific cis-elements for transcriptional regulation of PHO84 in Saccharomyces cerevisiae.
Transcription of the PHO84 gene encoding a Pi transporter in Saccharomyces cerevisiae is regulated by the Pi concentration in the medium. The promoter region of PHO84 bears five copies of the motif 5'-CACGT(G/T)-3', a candidate for the upstream activation site (UAS) that binds the transcriptional activator protein of the phosphatase regulon, Pho4p. These motifs are found at nucleotides -880 (site A), -587 (B), -436 (C), -414 (D), and -262 (E) relative to the putative ATG codon of PHO84. The Pho4p binds to all five 6-bp motifs with various affinities. Deletion analysis of the PHO84 promoter using a PHO84-lacZ fusion gene and base substitutions in the 6-bp motif revealed that two copies of the 6-bp motif, either C or D, and E, are necessary and sufficient for full regulation of the PHO84 gene. Results of expression studies with a CYC1-lacZ fusion gene with various 36-bp oligonucleotides including the 30-bp sequences around site D or E, or with modified sequences, inserted in the CYC1 promoter region indicated that the 6-bps motif flanked by a thymine nucleotide at its 5' end is much less effective as a UAS site for Pho4p in vivo than other versions. Thus, the consensus sequences for phosphatase regulation are 5'-GCACGTGGG-3' and 5'-GCACGTTTT-3' which differ from the binding sequences for the Cpflp protein required for transcription of the genes in methionine biosynthesis and for centromere function. However, Pho4p binding in vitro was unaffected by modification of the 5' or 3' flanking sites of the 6-bp motif, while modification inside the 6-bp motif affected it severely. The UAS function of the GCACGTTTT motif with respect to the Pi signal depends on its orientation in the promoter sequence.
Base Sequence↗
Mutations causing high basal level transcription that is independent of transcriptional activators but dependent on chromosomal position in Saccharomyces cerevisiae.
Two single (bel2 and bel4) and two double (bel3 bel7 and bel5 be16) mutations causing enhanced transcription of a gene fusion, consisting of the open reading frame of PHO5 connected to the HIS5 promoter (HIS5p) integrated at the ura3 or leu2 locus, were isolated from a gcn4-disrupted mutant of Saccharomyces cerevisiae. The PHO5 gene, encoding repressible acid phosphatase, in the HIS5p-PHO5 construct was derepressed under amino acid starved conditions by the action of the transcriptional activator Gcn4p. The bel mutants showed temperature-sensitive cell growth and/or cell aggregation. All the mutants except bel4 also showed high levels of transcription of an intact PHO5 DNA integrated at the URA3 locus in the absence of the cognate transcriptional activator, Pho4p, and in the absence of upstream activating sequences of PHO5. The HIS5 and PHO5 genes at their original chromosomal positions were, however, not affected by the bel2 mutation. The BEL2 gene was found to be identical with SIN4/TSF3, mutations in which cause high levels of transcription of the HO and GAL genes in the absence of their respective transcriptional activators, Swi5p and Gal4p. The effect of the bel2/sin4/tsf3 mutation on PHO5 transcription was additive with the Pho4p function. Thus the effect of the bel2/sin4/tsf3 mutation is dependent on the position of PHO5 in the chromosome and independent of Pho4p and Gen4p activation.
Alleles↗
Determination of a new fluoroquinolone antimicrobial agent, (S)-10-[(S)-(8-amino-6-azaspiro d3,4]octan-6-yl)]-9-fluoro-2, 3-dihydro-3-methyl-7-oxo-7H-pyrido[1,2,3-de][1,4]benzoxazine-6- carboxylic acid hemihydrate, DV-7751a, in human serum and urine using solid-phase extraction and high-performance liquid chromatography with fluorescence detection.
A high-performance liquid chromatographic method for the determination of a new fluoroquinolone antimicrobial agent, (S)-10-[(S)-(8-amino-6-azaspiro[3,4]octan-6-yl)]-9-fluoro-2,3- dihydro-3-methyl-7-oxo-7H-pyrido[1,2,3-de][1,4]benzoxazine-6-carboxylic acid hemihydrate (DV-7751a, I) in human serum and urine has been developed. Compound I and the internal standard were extracted from serum and urine by means of Bond Elut C8 LRC column. The extracts were chromatographed on a reversed-phase Inertsil ODS-2 column using tetrahydrofuran-50 mM KH2PO4 (pH2)-1 M ammonium acetate (19:81:1, v/v) as the mobile phase at a flow-rate of 1.0 ml/min. Fluorescence detection at an excitation wavelength of 305 nm and an emission wavelength of 530 nm resulted in a limit of quantitation of 0.0098 microgram/ml for serum and 0.098 microgram/ml for urine. The method showed satisfactory sensitivity, precision, accuracy, recovery and selectivity. Stability studies showed that I was stable in serum and urine for at least 1 month at -20 degrees C and for at least 48 h at room temperature.
Anti-Infective Agents↗
Powerful hepatoprotective and hepatotoxic plant oligostilbenes, isolated from the Oriental medicinal plant Vitis coignetiae (Vitaceae).
The methanol extract of the Oriental medicinal plant Vitis coignetiae (Vitaceae) showed hepatoprotective activity in the in vitro assay method using primary cultured rat hepatocytes. Activity-guided fractionation of the extract afforded epsilon-viniferin as an active principle. The protective effect of epsilon-viniferin against mice carbon tetrachloride-induced hepatic injury in mice was shown by serum enzyme assay as well as by pathological examination. In addition to epsilon-viniferin, plant oligostilbenes, ampelopsins A, C, F and the mixture of vitisin A and cis-vitisin A were also present in the extract. Among them, ampelopsin C and the mixture of vitisin A and cis-vitisin A were found to be powerful hepatotoxins.
Animals↗
Functional domains of Pho81p, an inhibitor of Pho85p protein kinase, in the transduction pathway of Pi signals in Saccharomyces cerevisiae.
The PHO81 gene is thought to encode an inhibitor of the negative regulators (Pho80p and Pho85p) in the phosphatase (PHO) regulon. Transcription of PHO81 is regulated by Pi signals through the same PHO regulatory system. Elimination of the PHO81 promoter or its substitution by the GAL1 promoter revealed that stimulation of the PHO regulatory system requires both increased transcription of PHO81 and a Pi starvation signal. The predicted Pho81p protein contains 1,179 amino acids (aa) and has six repeats of an ankyrin-like sequence in its central region. The minimum amino acid sequence required for Pho81p function was narrowed down to a 141-aa segment (aa 584 to 724), which contains the fifth and sixth repeats of the ankyrin-like motif. The third to sixth repeats of the ankyrin-like motif of Pho81p have significant similarities to that of p16INK4, which inhibits activity of the human cyclin D-CDK4 kinase complex. Deletion analyses revealed that the N- and C-terminal regions of Pho81p behave as negative and positive regulatory domains, respectively, for the minimal 141-aa region. The negative regulatory activity of the N-terminal domain was antagonized by a C-terminal segment of Pho81p supplied in trans. All four known classes of PHO81c mutations that show repressible acid phosphatase activity in high-Pi medium affect the N-terminal half of Pho81p. An in vitro assay showed that a glutathione S-transferase-Pho81p fusion protein inhibits the Pho85p protein kinase. Association of Pho81p with Pho85p or with the Pho80p-Pho85p complex was demonstrated by the two-hybrid system.
Amino Acid Sequence↗
Selective induction of interleukin-4- and interferon-y-producing T cells from cord blood naive T cells. Effects of costimulatory signaling through CD28.
We investigated the effect of costimulation through CD28 and CD11a on the differentiation of human naive CD4+ T cells with restricted cytokine production profiles. Interleukin (IL)-4 and interferon-gamma (IFN-gamma) were measured by ELISA and IL-2 was detected by a bioassay. Naive CD4+ T cells proliferated and produced IL-2 upon cross-linking of CD3, and costimulation through CD28 enhanced IL-2 production. After repeated stimulation, CD4+ T cells which were stimulated in the absence of costimulation through CD28 lost their ability to secrete IL-2 and started secreting IL-4 and IFN-gamma. Instead in the presence of costimulation through CD28, they secreted IL-2, IL-4 and IFN-gamma. Blocking of endogenous IL-4 activity with anti-IL-4 Ab suppressed the IL-4 secretion and proliferation of T cells.
CD28 Antigens↗
Antidepressant principles of Valeriana fauriei roots.
A methanol extract of the roots of Valeriana fauriei (Valerianaceae), exhibited antidepressant activity in mice. The extract was fractioned, monitored by the activity, to afford alpha-kessyl alcohol as an active principle. The antidepressant activity of some guaiane and valerane types of sesquiterpenoids in the active fraction was also evaluated.
Animals↗
[A case of Ebstein's anomaly treated by tricuspid valve replacement].
A 8-year-old girl with Ebstein's anomaly was operated on because of progressive heart failure caused by severe tricuspid regurgitation. At the operation, the tricuspid leaflets were found to be destroyed, and the annulus was very enlarged. The enlarged true tricuspid annulus was constricted using the DeVega method, and the tricuspid valve was replaced on the true annulus using a porcine valve. To avoid injury to the auriculoventricular bundle, the suture line was shifted to the false annulus at the portion close to the course of His' bundle. Postoperative course was uneventful in satisfactory condition. The role of valve replacement in the treatment of Ebstein's anomaly of the tricuspid valve is not only to improve valve formation, but also to secure the function of the right ventricle. It is ideal to replace the tricuspid valve on the true annulus provided that the injury to the His' bundle is safely avoided.
Child↗
[Targeted toxin therapy in the treatment of leukemia].
A chimeric toxin in which the cell surface binding domain of Pseudomonas exotoxin was replaced with mature human granulocyte colony-stimulating factor (G-CSF) was produced in Escherichia coli, partially purified and tested for its biological activity on a G-CSF-dependent murine myeloid leukemia cell line, NFS60. This fusion protein, termed as G-CSF-PE40, can displace [125I] G-CSF binding to its receptor. After 48 hrs incubation in the presence of G-CSF, G-CSF-PE40 inhibited protein synthesis and revealed cytotoxicity in NFS60 cells in a concentration dependent manner. The half maximal dose of G-CSF-PE40 for protein synthesis inhibition (ID50) in NFS60 cells was estimated at around 100pM. Additionally, relatively low concentration of G-CSF-PE40 stimulated DNA synthesis in NFS60 cells after 16 hrs' incubation in the absence of G-CSF, suggesting that G-CSF-PE40 also can transduce a transient mitogenic signal. Thus, G-CSF-PE40 may be useful in the selective elimination of myeloid cells expressing G-CSF receptors, especially in combination with chemotherapeutic agents like cytosine arabinoside.
ADP Ribose Transferases↗
[Multiple myeloma developing myelodysplastic syndrome with thrombocytosis].
A 64-year-old woman with multiple myeloma, IgG lambda type Durie-Salmon Stage II, was admitted because of gradually developing anemia and increased blasts with abnormal karyotype in her bone marrow after 10 years of treatment. The chromosomal analysis showed 44, XX, del(5q), del(7q), -9, add(12p), -21, typical of secondary MDS due to the cumulative alkylating agents. Thrombocytosis concomitantly occurred with emergence of chromosomal abnormality, but the serum interleukin 6 level was not elevated, which suggested that it was related to development of secondary MDS.
Antineoplastic Agents↗
[A diabetic patient with bilateral carotid stenosis who developed neovascular glaucoma following cataract surgery].
A 67-year-old diabetic man suffered from right neovascular glaucoma following the ipsilateral cataract surgery. Three years later, he underwent left cataract surgery and again developed left neovascular glaucoma after the operation. Fluorescein angiogram showed a marked delay in retinal circulation. Moreover, severe stenosis of bilateral carotid origins and reflux of bilateral ophthalmic arteries were ascertained by neurosonographical examination such as duplex cervical echography and transcranial Doppler, as well as an angiogram. Brain imaging demonstrated asymptomatic watershed infarction in the left parieto-occipital cortex. Chronic ocular ischemia caused by carotid stenosis is one of the decisive risk factors for secondary glaucoma after cataract surgery. Preoperative neurosonographical screening tests are required to decrease ocular surgery complications, especially in the aged, and diabetic patients.
Aged↗
Characteristics of paralytic shellfish poisoning toxins derived from short-necked clams (Tapes japonica) in Mikawa Bay.
A "red tide" bloom of Alexandrium tamarense occurred in Mikawa Bay in April 1991. In association with this, paralytic shellfish poisoning toxin was detected in short-necked clams by mouse bioassay. High performance liquid chromatography analysis demonstrated large quantities of the N-sulfocarbamoyl toxins, C1, and C2 (approximately 90% of the total toxins), whose latent potency was confirmed by treatment with boiling at low pH.
Animals↗
Differences in the drug-metabolizing enzyme activities among fish and bivalves living in waters near industrial and non-industrial areas.
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Animals↗
Protective effect of heparin on renal glomerular anionic sites of streptozotocin-injected rats.
Albuminuria at the rate of 500 micrograms/24 h was observed in rats treated with 50 mg/kg body wt. i.v. streptozotocin (STZ). When STZ was combined with heparin, administered twice daily (250 IU/kg subcutaneous injection) after 24 h following STZ injection, the daily urinary albumin excretion (U-AE) was less than half the amount found in non-heparinized rats (280.3 micrograms/24 h). The observation period in both instances was 8 weeks. When animals were permitted to develop albuminuria over the first 4 weeks, subsequent heparin administration for another 4 weeks lowered U-AE from 500 micrograms/24 h to less than half the amount (227.8 micrograms/24 h). A significantly negative correlation (P < 0.001) existed between U-AE and the number of anionic sites (AS) in the lamina rara externa of glomerular basement membranes, as visualized by electron microscopy. The number of AS per 1000 nm in STZ-injected rats (15.5 +/- 0.2) was lower than that in control rats (23.1 +/- 0.6); however, in heparinized animals, regardless of STZ, the values were not significantly different from normal. Heparin by itself had no effect on the number of AS in normal rats. All STZ-injected animals became hyperglycemic (400-550 mg/dl), but received no insulin. Heparin had no effect on plasma glucose levels. From these results, it is concluded that heparin suppressed both of an increase in U-AE and a decrease in AS count of glomerular basement membranes in STZ-injected rats.
Albuminuria↗