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S Fujioka

Publications and source records attributed to S Fujioka.

At least 55 records · Page 3Linked to original sources
PubMed2000

Biosynthetic pathways of brassinolide in Arabidopsis.

Our previous studies on the endogenous brassinosteroids (BRs) in Arabidopsis have provided suggestive evidence for the operation of the early C6-oxidation and the late C6-oxidation pathways, leading to brassinolide (BL) in Arabidopsis. However, to date the in vivo operation of these pathways has not been fully confirmed in this species. This paper describes metabolic studies using deuterium-labeled BRs in wild-type and BR-insensitive mutant (bri1) seedlings to establish the intermediates of the biosynthetic pathway of BL in Arabidopsis. The first evidence for the conversion of campestanol to 6-deoxocathasterone and the conversion of 6-deoxocathasterone to 6-deoxoteasterone is provided. The later biosynthetic steps (6-deoxoteasterone --> 3-dehydro-6-deoxoteasterone --> 6-deoxotyphasterol --> 6-deoxocastasterone --> 6alpha-hydroxycastasterone --> castasterone --> BL) were demonstrated by stepwise metabolic experiments. Therefore, these studies complete the documentation of the late C6-oxidation pathway. The biosynthetic sequence involved in the early C6-oxidation pathway (teasterone --> 3-dehydroteasterone --> typhasterol --> castasterone --> BL) was also demonstrated. These results show that both the early and late C6-oxidation pathways are functional in Arabidopsis. In addition we report two new observations: the presence of a new branch in the pathway, C6 oxidation of 6-deoxotyphasterol to typhasterol, and increased metabolic flow in BR-insensitive mutants.

Arabidopsis↗
PubMed2000

Loss of function of a rice brassinosteroid insensitive1 homolog prevents internode elongation and bending of the lamina joint.

Brassinosteroids (BRs) are plant growth-promoting natural products required for plant growth and development. Physiological studies have demonstrated that exogenous BR, alone or in combination with auxin, enhance bending of the lamina joint of rice. However, little is known about the function of endogenous BR in rice or other grass species. We report here the phenotypical and molecular characterization of a rice dwarf mutant, d61, that is less sensitive to BR compared to the wild type. We cloned a rice gene, OsBRI1, with extensive sequence similarity to that of the Arabidopsis BRI gene, which encodes a putative BR receptor kinase. Linkage analysis showed that the OsBRI1 gene is closely linked to the d61 locus. Single nucleotide substitutions found at different sites of the d61 alleles would give rise to amino acid changes in the corresponding polypeptides. Furthermore, introduction of the entire OsBRI1 coding region, including the 5' and 3' flanking sequences, into d61 plants complemented the mutation to display the wild-type phenotype. Transgenic plants carrying the antisense strand of the OsBRI1 transcript showed similar or even more severe phenotypes than those of the d61 mutants. Our results show that OsBRI1 functions in various growth and developmental processes in rice, including (1) internode elongation, by inducing the formation of the intercalary meristem and the longitudinal elongation of internode cells; (2) bending of the lamina joint; and (3) skotomorphogenesis.

Amino Acid Sequence↗
PubMed2000

Nematicidal alkaloids and related compounds produced by the fungus Penicillium cf. simplicissimum.

A new nematicidal alkaloid, peniprequinolone (1), together with the known alkaloids penigequinolones A and B (2a, 2b), 3-methoxy-4-hydroxy-4-(4'-methoxyphenyl)quinolinone (3), and 3-methoxy-4,6-dihydroxy-4-(4'-methoxyphenyl)quinolinone (4), were isolated from Penicillium cf. simplicissimum (Oudemans) Thom. Cyclopenin (5) and a compound (6a/6b) structurally related to cyclopenin also were isolated from the fungus, and their structures were established by spectroscopic analysis. The biological activities of 1, 2, 3, 4, and 5 were examined by a bioassay with root-lesion nematodes.

Alkaloids↗
PubMed2000

Retinal changes after retinal translocation surgery with scleral imbrication in dog eyes.

PURPOSE: To examine retinal changes induced by scleral imbrication during retinal translocation surgery in dog eyes. METHODS: Fifteen dogs were anesthetized and underwent retinal translocation surgery. After lensectomy and vitrectomy, an intentional retinal detachment was created, and the upper temporal sclera around the equator was imbricated with five mattress sutures. Translocated distances were calculated by pre- and postoperative photographs. At 1, 2, and 4 weeks after the surgery, the retina was studied by TdT-dNTP terminal nick-end labeling (TUNEL) and immunohistochemistry of peanut agglutinin (PNA) lectin and glial fibrillary acidic protein (GFAP). RESULTS: The retina was translocated by a mean distance of 0.53 +/- 0.30 disc diameters or 959 +/- 543 micrometer. Retinal folds were created around the optic disc in all eyes. Histologic examination of the retinal folds 1 week after the surgery showed many TUNEL-positive cells in the outer nuclear layer, loss of photoreceptor cells, and shortening of the outer and inner segments. A strong immunoreactivity to GFAP was detected in the folds of the retina. CONCLUSIONS: . The results demonstrated that retinal translocation surgery by scleral imbrication inevitably caused retinal folds as a postoperative complication, and the retina within the folds showed extensive loss of photoreceptor cells. It is recommended that the foveal translocation surgery be planned to avoid involving the fovea in the retinal folds.

Animals↗
PubMed1999

BAS1: A gene regulating brassinosteroid levels and light responsiveness in Arabidopsis.

The Arabidopsis bas1-D mutation suppresses the long hypocotyl phenotype caused by mutations in the photoreceptor phytochrome B (phyB). The adult phenotype of bas1-D phyB-4 double mutants mimics that of brassinosteroid biosynthetic and response mutants. bas1-D phyB-4 has reduced levels of brassinosteroids and accumulates 26-hydroxybrassinolide in feeding experiments. The basis for the mutant phenotype is the enhanced expression of a cytochrome P450 (CYP72B1). bas1-D suppresses a phyB-null allele, but not a phyA-null mutation, and partially suppresses a cryptochrome-null mutation. Seedlings with reduced BAS1 expression are hyperresponsive to brassinosteroids in a light-dependent manner and display reduced sensitivity to light under a variety of conditions. Thus, BAS1 represents one of the control points between multiple photoreceptor systems and brassinosteroid signal transduction.

Arabidopsis↗
PubMed1999

Pulmonary delivery of insulin with nebulized DL-lactide/glycolide copolymer (PLGA) nanospheres to prolong hypoglycemic effect.

Insulin loaded PLGA nanospheres having weight mean diameters of 400 nm were prepared by the modified emulsion solvent diffusion method in water. The nanosphere recovery and the drug recovery in the nanospheres were 74.8%+/-4.71 and 46.8%+/-7.01, respectively. Eighty five percent of the drug was released from the nanospheres at the initial burst, followed by prolonged releasing of the remaining drug for a few hours in saline at 37 degrees C. The aqueous dispersions (6 mg/ml) of PLGA nanospheres were nebulized by a sieve type ultrasonic nebulizer to discrete droplets of 5 approximately 7 microm in mean diameters, 75% of which were successfully delivered into the alveolar fraction in a cascade impactor inhaled at 28.3 l/min. The nebulized PLGA nanospheres were administered via a spacer by using a constant volume respirator into the trachea of the fasted guinea pig for 20 min. After the administration of 3.9 I.U./kg insulin with the PLGA nanospheres, the blood glucose level was reduced significantly and the hypoglycemia was prolonged over 48 h, compared to the nebulized aqueous solution of insulin as a reference (6 h). This result could be attributed to the sustained releasing of insulin from the nanospheres deposited widely on to the whole of lung.

Administration, Inhalation↗
PubMed1999

Mouse semaphorin H induces PC12 cell neurite outgrowth activating Ras-mitogen-activated protein kinase signaling pathway via Ca(2+) influx.

We recently showed that mouse semaphorin H (MSH), a secreted semaphorin molecule, acts as a chemorepulsive factor on sensory neurites. In this study, we found for the first time that MSH induces neurite outgrowth in PC12 cells in a dose-dependent manner. Comparison of Ras-mitogen-activated protein kinase (MAPK) signaling pathways between MSH and nerve growth factor (NGF) revealed that these pathways are crucial for MSH action as well as NGF. K-252a, an inhibitor of tyrosine autophosphorylation of tyrosine kinase receptors (Trks), did not inhibit the action of MSH, suggesting that MSH action occurs via a different receptor than NGF. L- and N-types of voltage-dependent Ca(2+) channel blockers, diltiazem and omega-conotoxin, inhibited MSH-induced neurite outgrowth and MAPK phosphorylation in a Ca(2+)-dependent manner. A transient elevation in intracellular Ca(2+) level was observed upon MSH stimulation. These findings suggest that extracellular Ca(2+) influx, followed by activation of the Ras-MAPK signaling pathway, is required for MSH induced PC12 cell neurite outgrowth.

3T3 Cells↗
PubMed1999

The tomato DWARF enzyme catalyses C-6 oxidation in brassinosteroid biosynthesis.

Brassinosteroids (BRs) are steroidal plant hormones essential for normal plant growth and development. Mutants in the biosynthesis or perception of BRs are usually dwarf. The tomato Dwarf gene (D), which was predicted to encode a cytochrome P450 enzyme (P450) with homology to other P450s involved in BR biosynthesis, was cloned previously. Here, we show that DWARF catalyses the C-6 oxidation of 6-deoxocastasterone (6-deoxoCS) to castasterone (CS), the immediate precursor of brassinolide. To do this, we first confirmed that the D cDNA complemented the mutant light- and dark-grown phenotypes of the extreme dwarf (dx) allele of tomato. To identify a substrate for the DWARF enzyme, exogenous application of BR intermediates to dx plants was carried out. C-6 oxoBR intermediates enhanced hypocotyl elongation whereas the C-6 deoxoBR, 6-deoxoCS, had little effect. Quantitative analysis of endogenous BR levels in tomato showed mainly the presence of 6-deoxoBRs. Furthermore, dx plants were found to lack CS and had a high level of 6-deoxoCS in comparison to D plants that had CS and a lower level of 6-deoxoCS. Confirmation that DWARF catalyzed the C-6 oxidation of 6-deoxoCS to CS was obtained by functional expression of DWARF in yeast. In these experiments, the intermediate 6alpha-hydroxycastasterone was identified, indicating that DWARF catalyzes two steps in BR biosynthesis. These data show that DWARF is involved in the C-6 oxidation in BR biosynthesis.

Journal Article↗
PubMed1999

New lead compounds for brassinosteroid biosynthesis inhibitors.

The first brassinosteroid biosynthesis inhibitor is reported. Among newly synthesized triazole derivatives, 4-(4-chlorophenyl)-2-phenyl-3-(1,2,4-triazoyl)butan-2-ol (6) was found to inhibit the growth of cress seedlings, and this inhibition was recovered by the treatment of brassinolide, suggesting that compound 6 primarily inhibits brassinosteroid biosynthesis.

Butanols↗
PubMed1999

Lymphoepithelioma-like esophageal carcinoma: report of a case.

We herein report the rare case of a patient suffering from lymphoepithelioma-like poorly differentiated squamous cell carcinoma of the esophagus. The patient was a 74-year-old woman in whom an esophageal tumor was found during an operation for thyroid cancer. After performing a subtotal thyroidectomy and cervical esophagectomy, esophageal reconstruction was performed using a free jejunal graft. Based on the results of the pathological examination, the esophageal tumor was diagnosed to be primary lymphoepithelioma-like esophageal cancer, not metastasis of either unknown nasopharyngeal cancer or thyroid cancer. Since surgery, she has survived postoperatively for more than 4 years with no evidence of recurrent disease.

Aged↗
PubMed1999

Surgically induced degeneration and regeneration of the choriocapillaris in rabbit.

BACKGROUND: Retinal pigment epithelium (RPE) and choriocapillaris play an important role in maintaining the outer retina. Clinical and experimental studies have shown that there is a close relationship between them. To examine the relationship between RPE and choriocapillaris we developed an animal model in which degeneration and regeneration of the choriocapillaris can be created easily and reproducibly. METHODS: Using pigmented rabbits a retinal detachment (about 7 disk diameters) was created in the eye and the detached retina was surgically removed. A half area of the exposed RPE was mechanically debrided using a silicone brush. The other half area remained untouched. The eyes were morphologically examined at 3, 7, and 14 days and 4 and 8 weeks after the surgery using scanning or transmission electron microscopy. Changes in the choriocapillaris after the RPE debridement were also evaluated by corrosion vascular casts. RESULTS: The debrided area was resurfaced with elongated RPE in 7 days after surgery. The replaced RPE was dedifferentiated but it gradually recovered cell polarity by 4 weeks after surgery. The choriocapillaris beneath the debrided area underwent degeneration; however, it regenerated by 4 weeks after surgery, corresponding to the timing of the morphologic recovery of the replaced RPE. CONCLUSION: This animal model of surgically induced degeneration and regeneration of the choriocapillaris may be useful to clarify the relationship between RPE and choriocapillaris and to study potential treatments for choroidal vascular diseases.

Animals↗
PubMed1999

Surgery and long-term outcome for ruptured anterior circulation aneurysms in patients in their ninth decade of life.

BACKGROUND: The present pilot study was undertaken to analyze the long-term results of surgery for ruptured anterior circulation aneurysms in patients in their 9th decade of life. METHODS: Between 1992 and 1997, we treated 10 consecutive patients with ruptured anterior circulation aneurysms who were 80 years or older on admission and judged eligible for surgery based on stringent criteria. The outcomes at discharge and the latest outcomes (obtained at a median of 41.6 months) were assessed using the Glasgow Outcome Scale and the Barthel Score, respectively. RESULTS: Upon discharge, six of the 10 patients showed good recovery, two patients were moderately disabled, and two patients who suffered symptomatic vasospasm were severely disabled. As for the latest outcomes, assessed using Barthel Score, four patients had the maximum score of 100, two a score of 90, one a score of 35, two a score of 10, and one patient died. Of four patients who were in poor condition, two experienced deterioration attributable to unrelated causes 18 and 32 months after the ictus. CONCLUSION: Advanced age alone does not preclude successful surgery for ruptured anterior circulation aneurysms. Carefully selected patients over 80 years should also be considered for surgical treatment.

Aged↗
PubMed1999

Developmental localization of semaphorin H messenger RNA acting as a collapsing factor on sensory axons in the mouse brain.

Semaphorins/collapsins, a family of genes with a semaphorin domain conserved from insects through to mammals, are believed to be involved in axon guidance during neuronal development. We report the expression patterns of mouse semaphorin messenger RNAs. Among secreted semaphorins, mouse semaphorin H is structurally most similar to semaphorin III/D, the first semaphorin identified as a collapsing factor for sensory axons. However, its expression patterns apparently differ from those of semaphorin III/D. The messenger RNAs are distributed in the brain widely but unevenly during development, in particular, in the main olfactory bulb, hippocampus and pontine nucleus. In the trunk, the expression level is high in mesodermal tissues surrounding the dorsal root ganglia, while it is low in the spinal cord. Moreover, we examined whether this molecule has activity to collapse growth cones of sensory neurons, as well as semaphorin III/D. Mouse semaphorin H collapsed growth cones of sensory neurons of the dorsal root ganglion in a dose-dependent manner, and anti-neuropilin antibodies inhibited this activity. Taken together, these results suggest that mouse semaphorin H can function as a chemorepellent to guide sensory peripheral nerves, most likely via neuropilin as a receptor.

Amino Acid Sequence↗
PubMed1999

The sax1 mutation defines a new locus involved in the brassinosteroid biosynthesis pathway in Arabidopsis thaliana.

In this issue we described a dwarf mutant in Arabidopsis thaliana, sax1, which is affected in brassinosteroid biosynthesis. This primary defect is responsible for alterations in hormone sensitivity of sax1 plants characterized by the hypersensitivity of root elongation to abscisic acid and auxin and the insensitivity of hypocotyl growth to gibberellins and ethylene (Ephritikhine et al., 1999; Plant J. 18, 303-314). In this paper, we report the further characterization of the sax1 mutant aimed at identification of the mutated step in the brassinosteroid biosynthesis pathway. Rescue experiments with various intermediates of the pathway showed that the sax1 mutation alters a very early step catalyzing the oxidation and isomerization of 3 beta-hydroxyl, delta 5,6 precursors to 3-oxo, delta 4,5 steroids. The mapping of the mutation, the physiological properties of the mutant and the rescue experiments indicate that sax1 defines a new locus in the brassinosteroid biosynthesis pathway. The SAX1 protein is involved in brassinosteroid-dependent growth of seedlings in both light and dark conditions.

Arabidopsis↗
PubMed1999

The Arabidopsis dwarf1 mutant is defective in the conversion of 24-methylenecholesterol to campesterol in brassinosteroid biosynthesis.

Since the isolation and characterization of dwarf1-1 (dwf1-1) from a T-DNA insertion mutant population, phenotypically similar mutants, including deetiolated2 (det2), constitutive photomorphogenesis and dwarfism (cpd), brassinosteroid insensitive1 (bri1), and dwf4, have been reported to be defective in either the biosynthesis or the perception of brassinosteroids. We present further characterization of dwf1-1 and additional dwf1 alleles. Feeding tests with brassinosteroid-biosynthetic intermediates revealed that dwf1 can be rescued by 22alpha-hydroxycampesterol and downstream intermediates in the brassinosteroid pathway. Analysis of the endogenous levels of brassinosteroid intermediates showed that 24-methylenecholesterol in dwf1 accumulates to 12 times the level of the wild type, whereas the level of campesterol is greatly diminished, indicating that the defective step is in C-24 reduction. Furthermore, the deduced amino acid sequence of DWF1 shows significant similarity to a flavin adenine dinucleotide-binding domain conserved in various oxidoreductases, suggesting an enzymatic role for DWF1. In support of this, 7 of 10 dwf1 mutations directly affected the flavin adenine dinucleotide-binding domain. Our molecular characterization of dwf1 alleles, together with our biochemical data, suggest that the biosynthetic defect in dwf1 results in reduced synthesis of bioactive brassinosteroids, causing dwarfism.

Alleles↗
PubMed1999

Arabidopsis det2 is defective in the conversion of (24R)-24-methylcholest-4-En-3-one to (24R)-24-methyl-5alpha-cholestan-3-one in brassinosteroid biosynthesis.

Previously, we have shown that the Arabidopsis det2 (deetiolated2) mutant is defective in the biosynthesis of brassinosteroids (BR) and that DET2 (a steroid 5alpha-reductase) acts early in the proposed BR biosynthetic pathway. In this paper we present further biochemical characterization of det2. We have undertaken metabolic experiments with 2H-labeled substrates of intermediates involved in the formation of campestanol from campesterol, and quantitative analysis of intermediates in Arabidopsis wild type and det2. The results of these studies indicate the early operating steps of BR biosynthesis as: campesterol --> 4-en-3beta-ol --> 4-en-3-one --> 3-one --> campestanol in Arabidopsis, with det2 deficient in the conversion of 4-en-3-one to 3-one. We have also detected these intermediates in the formation of campestanol from campesterol and their metabolic conversions using cultured cells of Catharanthus roseus. These studies confirmed the biosynthetic sequence of events from campesterol to campestanol as was found in Arabidopsis. As such, the originally proposed biosynthetic pathway should be modified.

Arabidopsis↗
PubMed1999

Brassinosteroid-insensitive dwarf mutants of Arabidopsis accumulate brassinosteroids.

Seven dwarf mutants resembling brassinosteroid (BR)-biosynthetic dwarfs were isolated that did not respond significantly to the application of exogenous BRs. Genetic and molecular analyses revealed that these were novel alleles of BRI1 (Brassinosteroid-Insensitive 1), which encodes a receptor kinase that may act as a receptor for BRs or be involved in downstream signaling. The results of morphological and molecular analyses indicated that these represent a range of alleles from weak to null. The endogenous BRs were examined from 5-week-old plants of a null allele (bri1-4) and two weak alleles (bri1-5 and bri1-6). Previous analysis of endogenous BRs in several BR-biosynthetic dwarf mutants revealed that active BRs are deficient in these mutants. However, bri1-4 plants accumulated very high levels of brassinolide, castasterone, and typhasterol (57-, 128-, and 33-fold higher, respectively, than those of wild-type plants). Weaker alleles (bri1-5 and bri1-6) also accumulated considerable levels of brassinolide, castasterone, and typhasterol, but less than the null allele (bri1-4). The levels of 6-deoxoBRs in bri1 mutants were comparable to that of wild type. The accumulation of biologically active BRs may result from the inability to utilize these active BRs, the inability to regulate BR biosynthesis in bri1 mutants, or both. Therefore, BRI1 is required for the homeostasis of endogenous BR levels.

Alleles↗