Biomedical subjects

Shigeo Yoshida

Publications and source records attributed to Shigeo Yoshida.

At least 37 records · Page 2Linked to original sources

PubMed2006

[What can we learn from molecular genetic analyses of inherited eye diseases?].

Over the past decade, the pace of gene identification of the causes of inherited eye diseases has increased dramatically as the complete human genome information becoming available. Molecular genetic analysis establishes reliable clinical diagnostic criteria and improves the accuracy of diagnosis. We encountered two atypical cases, a patient who had only bilateral band-shaped opacities without any gelatinous prominences, and a patient who had diffuse central corneal stromal opacity without any lattice lines. The diagnosis of gelatinous drop-like dystrophy and lattice corneal dystrophy I was confirmed by molecular genetic analysis of TACSTD2 and TGFBI, respectively. We confirmed the existence of a predominantly ocular type of stickler syndrome by identifying the mutation involving exon 2 of the COL2A1 gene in a Japanese patient who had received a diagnosis of rhegmatogenous retinal detachment. We surmise that in Japan in the past, this diagnosis may have been overlooked or misdiagnosed as Wagner disease. Molecular genetic analysis is also useful for gaining a better understanding of diseases. We detected a novel FZD4 mutation in a patient with familial exudative vitreoretinopathy who exhibited peripheral avascular areas bilaterally, a dragged disk, and retinal holes unilaterally, suggesting that FZD4 may be involved in the angiogenesis of the human peripheral retina. Molecular genetic analysis of a Japanese patient with pseudoxanthoma elasticum with choroidal neovascularization revealed a homozygous nonsense mutation in the ABCC6, a member of the ABC transporter family, indicating that angioid streaks may be caused by a primary metabolic disorder. The development of rapid and comprehensive genotyping systems using state-of-the-art technology such as genotyping microarray may eventually offer unique and reliable diagnostic tools. This should then accelerate our understanding of the basic mechanisms underlying inherited eye diseases and their phenotypic variability, thus facilitating prospective diagnosis.

Antigens, Neoplasm↗

PubMed2005

Erect leaves caused by brassinosteroid deficiency increase biomass production and grain yield in rice.

New cultivars with very erect leaves, which increase light capture for photosynthesis and nitrogen storage for grain filling, may have increased grain yields. Here we show that the erect leaf phenotype of a rice brassinosteroid-deficient mutant, osdwarf4-1, is associated with enhanced grain yields under conditions of dense planting, even without extra fertilizer. Molecular and biochemical studies reveal that two different cytochrome P450s, CYP90B2/OsDWARF4 and CYP724B1/D11, function redundantly in C-22 hydroxylation, the rate-limiting step of brassinosteroid biosynthesis. Therefore, despite the central role of brassinosteroids in plant growth and development, mutation of OsDWARF4 alone causes only limited defects in brassinosteroid biosynthesis and plant morphology. These results suggest that regulated genetic modulation of brassinosteroid biosynthesis can improve crops without the negative environmental effects of fertilizers.

Biomass↗

PubMed2005

The UGT73C5 of Arabidopsis thaliana glucosylates brassinosteroids.

Steroid hormones are essential for development, and the precise control of their homeostasis is a prerequisite for normal growth. UDP-glycosyltransferases (UGTs) are considered to play an important regulatory role in the activity of steroids in mammals and insects. This study provides an indication that a UGT accepting plant steroids as substrates functions in brassinosteroid (BR) homeostasis. The UGT73C5 of Arabidopsis thaliana catalyses 23-O-glucosylation of the BRs brassinolide (BL) and castasterone. Transgenic plants overexpressing UGT73C5 displayed BR-deficient phenotypes and contained reduced amounts of BRs. The phenotype, which was already apparent in seedlings, could be rescued by application of BR. In feeding experiments with BL, wild-type seedlings converted BL to the 23-O-glucoside; in the transgenic lines silenced in UGT73C5 expression, no 23-O-glucoside was detected, implying that this UGT is the only enzyme that catalyzes BL-23-O-glucosylation in seedlings. Plant lines in which UGT73C5 expression was altered also displayed hypocotyl phenotypes previously described for seedlings in which BR inactivation by hydroxylation was changed. These data support the hypothesis that 23-O-glucosylation of BL is a function of UGT73C5 in planta, and that glucosylation regulates BR activity.

Animals↗

PubMed2005

DNA polymorphisms in the tetrahydrocannabinolic acid (THCA) synthase gene in "drug-type" and "fiber-type" Cannabis sativa L.

The cannabinoid content of 13 different strains of cannabis plant (Cannabis sativa L.) was analyzed. Six strains fell into the "drug-type" class, with high Delta-9-tetrahydrocannabinolic acid (THCA) content, and seven strains into the "fiber-type" class, with low THCA using HPLC analysis. Genomic DNA sequence polymorphisms in the THCA synthase gene from each strain were studied. A single PCR fragment of the THCA synthase gene was detected from six strains of "drug-type" plants. We could also detect the fragment from seven strains of "fiber-type" plants, although no or very low content of THCA were detected in these samples. These were 1638 bp from all 13 strains and no intron among the sequences obtained. There were two variants of the THCA synthase gene in the "drug-type" and "fiber-type" cannabis plants, respectively. Thirty-seven major substitutions were detected in the alignment of the deduced amino acid sequences from these variants. Furthermore, we identified a specific PCR marker for the THCA synthase gene for the "drug-type" strains. This PCR marker was not detected in the "fiber-type" strains.

Base Sequence↗

PubMed2005

TANMEI/EMB2757 encodes a WD repeat protein required for embryo development in Arabidopsis.

We identified the Arabidopsis (Arabidopsis thaliana) tanmei/emb2757 (tan) mutation that causes defects in both embryo and seedling development. tan mutant embryos share many characteristics with the leafy cotyledon (lec) class of mutants in that they accumulate anthocyanin, are intolerant of desiccation, form trichomes on cotyledons, and have reduced accumulation of storage proteins and lipids. Thus, TAN functions both in the early and late phases of embryo development. Moreover, the TAN and LEC genes interact synergistically, suggesting that they do not act in series in the same genetic pathway but, rather, that they have overlapping roles during embryogenesis. tan mutants die as embryos, but immature mutant seeds can be germinated in culture. However, tan mutant seedlings are defective in shoot and root development, their hypocotyls fail to elongate in the dark, and they die as seedlings. We isolated the TAN gene and showed that the predicted polypeptide has seven WD repeat motifs, suggesting that TAN forms complexes with other proteins. Together, these results suggest that TAN interacts with other proteins to control many aspects of embryo development.

Amino Acid Motifs↗

PubMed2005

Chemical regulation of abscisic acid catabolism in plants by cytochrome P450 inhibitors.

Plant hormone abscisic acid (ABA) is an important factor for conferring drought stress resistance on plants. Therefore, small molecules that regulate ABA levels in plants can be useful both for investigating functions of ABA and for developing new plant growth regulators. Abscisic acid (ABA) catabolism in plants is primarily regulated by ABA 8'-hydroxylase, which is a cytochrome P450 (P450). We tested known P450 inhibitors containing a triazole group and found that uniconazole-P inhibited ABA catabolism in cultured tobacco Bright Yellow-2 cells. In a structure-activity study of uniconazole, we found a more effective ABA catabolic inhibitor (diniconazole) than uniconazole-P. Diniconazole, a fungicide, acted as a potent competitive inhibitor of recombinant Arabidopsis ABA 8'-hydroxylase, CYP707A3, in an in vitro assay. Diniconazole-treated plants retained a higher ABA content and higher transcription levels of ABA response genes during rehydration than did untreated plants and were more drought stress tolerant than untreated plants. These results strongly suggest that ABA catabolic inhibitors that target ABA 8'-hydroxylase can regulate the ABA content of plants and conferred drought stress resistance on plants. The optical resolution of diniconazole revealed that the S-form isomer, which is a weak fungicidal isomer, was more active as an ABA catabolic inhibitor than was the R-form isomer.

Abscisic Acid↗

PubMed2005

The Rice brassinosteroid-deficient dwarf2 mutant, defective in the rice homolog of Arabidopsis DIMINUTO/DWARF1, is rescued by the endogenously accumulated alternative bioactive brassinosteroid, dolichosterone.

We have identified a rice (Oryza sativa) brassinosteroid (BR)-deficient mutant, BR-deficient dwarf2 (brd2). The brd2 locus contains a single base deletion in the coding region of Dim/dwf1, a homolog of Arabidopsis thaliana DIMINUTO/DWARF1 (DIM/DWF1). Introduction of the wild-type Dim/dwf1 gene into brd2 restored the normal phenotype. Overproduction and repression of Dim/dwf1 resulted in contrasting phenotypes, with repressors mimicking the brd2 phenotype and overproducers having large stature with increased numbers of flowers and seeds. Although brd2 contains low levels of common 6-oxo-type BRs, the severity of the brd2 phenotype is much milder than brd1 mutants and most similar to d2 and d11, which show a semidwarf phenotype at the young seedling stage. Quantitative analysis suggested that in brd2, the 24-methylene BR biosynthesis pathway is activated and the uncommon BR, dolichosterone (DS), is produced. DS enhances the rice lamina joint bending angle, rescues the brd1 dwarf phenotype, and inhibits root elongation, indicating that DS is a bioactive BR in rice. Based on these observations, we discuss an alternative BR biosynthetic pathway that produces DS when Dim/dwf1 is defective.

Amino Acid Sequence↗

PubMed2005

Brassinosteroid regulates fiber development on cultured cotton ovules.

Our current understanding of the role of phytohormones in the development of cotton fibers is derived largely from an amenable culture system in which cotton ovules, collected on the day of anthesis, are floated on liquid media. Under these conditions, supplemental auxin and gibberellin were found to promote fiber initiation and elongation. More recently, addition of low concentrations of the brassinosteroid brassinolide (BL) were also found to promote fiber elongation while a brassinosteroid biosynthesis inhibitor brassinazole2001 (Brz) inhibited fiber development. In order to elucidate the role of brassinosteroid in cotton fiber development further, we have performed a more detailed analysis of the effects of these chemicals on cultured cotton ovules. Our results confirm that exogenous BL promotes fiber elongation while treatment with Brz inhibits it. Furthermore, treatment of cotton floral buds with Brz results in the complete absence of fiber differentiation, indicating that BR is required for fiber initiation as well as elongation. Expression of fiber genes associated with cell elongation increased in ovules treated with BL and was suppressed by Brz treatment, establishing a correlation between brassinosteroid-regulated gene expression and fiber elongation. These results establish a clear connection between brassinosteroid and fiber development and open the door for genetic analysis of cotton development through direct modification of the brassinosteroid signal transduction pathway.

Base Sequence↗

PubMed2005

Mechanisms of aging in senescence-accelerated mice.

BACKGROUND: Progressive neurological dysfunction is a key aspect of human aging. Because of underlying differences in the aging of mice and humans, useful mouse models have been difficult to obtain and study. We have used gene-expression analysis and polymorphism screening to study molecular senescence of the retina and hippocampus in two rare inbred mouse models of accelerated neurological senescence (SAMP8 and SAMP10) that closely mimic human neurological aging, and in a related normal strain (SAMR1) and an unrelated normal strain (C57BL/6J). RESULTS: The majority of age-related gene expression changes were strain-specific, with only a few common pathways found for normal and accelerated neurological aging. Polymorphism screening led to the identification of mutations that could have a direct impact on important disease processes, including a mutation in a fibroblast growth factor gene, Fgf1, and a mutation in and ectopic expression of the gene for the chemokine CCL19, which is involved in the inflammatory response. CONCLUSION: We show that combining the study of inbred mouse strains with interesting traits and gene-expression profiling can lead to the discovery of genes important for complex phenotypes. Furthermore, full-genome polymorphism detection, sequencing and gene-expression profiling of inbred mouse strains with interesting phenotypic differences may provide unique insights into the molecular genetics of late-manifesting complex diseases.

Aging↗

PubMed2005

Brassinosteroid homeostasis in Arabidopsis is ensured by feedback expressions of multiple genes involved in its metabolism.

Homeostasis of brassinosteroids (BRs) is essential for normal growth and development in higher plants. We examined responsiveness of 11 BR metabolic gene expressions to the decrease or increase of endogenous BR contents in Arabidopsis (Arabidopsis thaliana) to expand our knowledge of molecular mechanisms underlying BR homeostasis. Five BR-specific biosynthesis genes (DET2, DWF4, CPD, BR6ox1, and ROT3) and two sterol biosynthesis genes (FK and DWF5) were up-regulated in BR-depleted wild-type plants grown under brassinazole, a BR biosynthesis inhibitor. On the other hand, in BR-excessive wild-type plants that were fed with brassinolide, four BR-specific synthesis genes (DWF4, CPD, BR6ox1, and ROT3) and a sterol synthesis gene (DWF7) were down-regulated and a BR inactivation gene (BAS1) was up-regulated. However, their response to fluctuation of BR levels was highly reduced (DWF4) or nullified (the other eight genes) in a bri1 mutant. Taken together, our results imply that BR homeostasis is maintained through feedback expressions of multiple genes, each of which is involved not only in BR-specific biosynthesis and inactivation, but also in sterol biosynthesis. Our results also indicate that their feedback expressions are under the control of a BRI1-mediated signaling pathway. Moreover, a weak response in the mutant suggests that DWF4 alone is likely to be regulated in other way(s) in addition to BRI1 mediation.

Arabidopsis↗

PubMed2005

Biotin-labeled abscisic acid as a probe for investigating abscisic acid binding sites on plasma membranes of barley aleurone protoplasts.

Plant hormone abscisic acid (ABA) plays important roles in dormancy and stress responses, but its binding sites have not yet been fully elucidated. In this report, we suggest the utility of biotin-labeled abscisic acid (bioABA) as a probe to investigate ABA-binding sites on the plasma membrane of barley aleurone protoplasts. BioABA was approximately 100 times less effective than ABA in inhibiting expression of gibberellin-inducible alpha-amylase and in inducing expression of a reporter gene fused to the dehydrin promoter. To ascertain that bioABA could bind to ABA-binding sites on the plasma membrane, we used fluorescence flow cytometry to measure the fluorescence intensity of aleurone protoplasts treated with a combination of bioABA and fluorescence-labeled streptavidin. Addition of bioABA increased the fluorescence of aleurone protoplasts in a concentration-dependent manner, but addition of non-active bioABA derivatives did not. Furthermore, the increase in fluorescence intensity observed upon addition of bioABA was eliminated by co-treatment with excess ABA, but it was not eliminated by co-treatment with other plant hormones. These results suggest that bioABA binds to ABA-binding sites, and that bioABA should be a valuable probe for investigating ABA-binding sites on the plasma membrane.

Abscisic Acid↗

PubMed2005

Identification and functional analysis of in vivo phosphorylation sites of the Arabidopsis BRASSINOSTEROID-INSENSITIVE1 receptor kinase.

Brassinosteroids (BRs) regulate multiple aspects of plant growth and development and require an active BRASSINOSTEROID-INSENSITIVE1 (BRI1) and BRI1-ASSOCIATED RECEPTOR KINASE1 (BAK1) for hormone perception and signal transduction. Many animal receptor kinases exhibit ligand-dependent oligomerization followed by autophosphorylation and activation of the intracellular kinase domain. To determine if early events in BR signaling share this mechanism, we used coimmunoprecipitation of epitope-tagged proteins to show that in vivo association of BRI1 and BAK1 was affected by endogenous and exogenous BR levels and that phosphorylation of both BRI1 and BAK1 on Thr residues was BR dependent. Immunoprecipitation of epitope-tagged BRI1 from Arabidopsis thaliana followed by liquid chromatography-tandem mass spectrometry (LC/MS/MS) identified S-838, S-858, T-872, and T-880 in the juxtamembrane region, T-982 in the kinase domain, and S-1168 in C-terminal region as in vivo phosphorylation sites of BRI1. MS analysis also strongly suggested that an additional two residues in the juxtamembrane region and three sites in the activation loop of kinase subdomain VII/VIII were phosphorylated in vivo. We also identified four specific BAK1 autophosphorylation sites in vitro using LC/MS/MS. Site-directed mutagenesis of identified and predicted BRI1 phosphorylation sites revealed that the highly conserved activation loop residue T-1049 and either S-1044 or T-1045 were essential for kinase function in vitro and normal BRI1 signaling in planta. Mutations in the juxtamembrane or C-terminal regions had only small observable effects on autophosphorylation and in planta signaling but dramatically affected phosphorylation of a peptide substrate in vitro. These findings are consistent with many aspects of the animal receptor kinase model in which ligand-dependent autophosphorylation of the activation loop generates a functional kinase, whereas phosphorylation of noncatalytic intracellular domains is required for recognition and/or phosphorylation of downstream substrates.

Arabidopsis↗

PubMed2005

Rapid genotyping for most common TGFBI mutations with real-time polymerase chain reaction.

Recent studies of the corneal dystrophies (CDs) have shown that most cases of granular CD, Avellino CD, and lattice CD type I are caused by mutations in the human transforming growth factor beta-induced (TGFBI) gene. The aim of this study was to develop a rapid diagnostic assay to detect mutations in the TGFBI gene. Sixty-six patients from 64 families with TGFBI-associated CD were studied. A primer probe set was designed to examine the genome from exons 4 and 12 of the TGFBI gene in order to identify mutant and wild-type alleles. A region spanning the mutations was amplified by the polymerase chain reaction (PCR) in a commercial cycler. Mutations were then identified by melting curve analysis of the hybrid formed between the PCR product and a specific fluorescent probe. Using this system, we clearly distinguished each CD genotype (homozygous and heterozygous 418G-->A, heterozygous 417C-->T, heterozygous 1710C-->T, and wild-type) of all the patients by means of the clearly distinct melting peaks at different temperatures. One thermal cycling took approximately 54 min, and all results were completely in concordance with the genotypes determined by conventional DNA sequencing. Thus, the technique is accurate and can be used for routine clinical diagnosis. We expect that our new method will help in making precise diagnoses of patients with atypical CDs and aid the revision of the clinical classification of inherited corneal diseases based on the genetic pathogenesis.

Adolescent↗

PubMed2005

Ectopic endoreduplication caused by sterol alteration results in serrated petals in Arabidopsis.

The Arabidopsis frill1 (frl1) mutant, that has serrated petals and sepals but no other large changes in plant morphology, was studied. The frl1 had a mutation in STEROL METHYLTRANSFERASE 2 and an altered sterol composition. It was found that the frl1 mutation causes ectopic endoreduplication in petal tips that do not normally endoreduplicate. The rosette leaves of frl1 also showed an enhanced level of endoreduplication, but their morphology was hardly affected. These facts suggest that the suppression of endoreduplication is important for petal morphogenesis and the normal sterol composition is required for this suppression.

Arabidopsis↗

PubMed2005

A novel cytochrome P450 is implicated in brassinosteroid biosynthesis via the characterization of a rice dwarf mutant, dwarf11, with reduced seed length.

We have characterized a rice (Oryza sativa) dwarf mutant, dwarf11 (d11), that bears seeds of reduced length. To understand the mechanism by which seed length is regulated, the D11 gene was isolated by a map-based cloning method. The gene was found to encode a novel cytochrome P450 (CYP724B1), which showed homology to enzymes involved in brassinosteroid (BR) biosynthesis. The dwarf phenotype of d11 mutants was restored by the application of the brassinolide (BL). Compared with wild-type plants, the aberrant D11 mRNA accumulated at higher levels in d11 mutants and was dramatically reduced by treatment with BL, implying that the gene is feedback-regulated by BL. Precise determination of the defective step(s) in BR synthesis in d11 mutants proved intractable because of tissue specificity and the complex control of BR accumulation in plants. However, 6-deoxotyphasterol (6-DeoxoTY) and typhasterol (TY), but not any upstream intermediates before these compounds, effectively restored BR response in d11 mutants in a lamina joint bending assay. Multiple lines of evidence together suggest that the D11/CYP724B1 gene plays a role in BR synthesis and may be involved in the supply of 6-DeoxoTY and TY in the BR biosynthesis network in rice.

Amino Acid Sequence↗

PubMed2005

Identification of a novel factor, vanillyl benzyl ether, which inhibits somatic embryogenesis of Japanese larch (Larix leptolepis Gordon).

In contrast to angiosperms, some gymnosperms form well-developed suspensors in somatic embryogenesis. This characteristic makes it easy to study suspensor biology. In cultures with high cell densities, somatic embryogenesis of Japanese larch, especially the suspensor development, is strongly inhibited due to factor(s) that are released by the cells into the culture medium. In this study, we purified and identified one of the inhibitory factors present in high-cell-density conditioned medium (HCM) of larch cells. The factor with the strongest inhibitory activity was purified by dialysis, extraction by ethyl acetate, octadecylsilyl (ODS) column chromatography and high-performance liquid chromatography (HPLC). The inhibitory factor was identified as vanillyl benzyl ether (VBE) by physicochemical analysis. This compound was first isolated from natural resources. Authentic VBE inhibited somatic embryo formation in Japanese larch, and the inhibitory effect in the suspensor was stronger than in the embryo proper. Furthermore, quantification of VBE by HPLC demonstrated that VBE accumulates at high concentrations in HCM. These results suggest that VBE is a novel negative regulator of somatic embryogenesis.

Benzaldehydes↗

PubMed2005

A new class of transcription factors mediates brassinosteroid-regulated gene expression in Arabidopsis.

Brassinosteroids (BRs) signal through a plasma membrane-localized receptor kinase to regulate plant growth and development. We showed previously that a novel protein, BES1, accumulates in the nucleus in response to BRs, where it plays a role in BR-regulated gene expression; however, the mechanism by which BES1 regulates gene expression is unknown. In this study, we dissect BES1 subdomains and establish that BES1 is a transcription factor that binds to and activates BR target gene promoters both in vitro and in vivo. BES1 interacts with a basic helix-loop-helix protein, BIM1, to synergistically bind to E box (CANNTG) sequences present in many BR-induced promoters. Loss-of-function and gain-of-function mutants of BIM1 and its close family members display BR response phenotypes. Thus, BES1 defines a new class of plant-specific transcription factors that cooperate with transcription factors such as BIM1 to regulate BR-induced genes.

Amino Acid Sequence↗

PubMed2005

Binding of brassinosteroids to the extracellular domain of plant receptor kinase BRI1.

Both animals and plants use steroids as signalling molecules during growth and development. Animal steroids are principally recognized by members of the nuclear receptor superfamily of transcription factors. In plants, BRI1, a leucine-rich repeat (LRR) receptor kinase localized to the plasma membrane, is a critical component of a receptor complex for brassinosteroids. Here, we present the first evidence for direct binding of active brassinosteroids to BRI1 using a biotin-tagged photoaffinity castasterone (BPCS), a biosynthetic precursor of brassinolide (the most active of the brassinosteroids). Binding studies using BPCS, (3)H-labelled brassinolide and recombinant BRI1 fragments show that the minimal binding domain for brassinosteroids consists of a 70-amino acid island domain (ID) located between LRR21 and LRR22 in the extracellular domain of BRI1, together with the carboxy-terminal flanking LRR (ID-LRR22). Our results demonstrate that brassinosteroids bind directly to the 94 amino acids comprising ID-LRR22 in the extracellular domain of BRI1, and define a new binding domain for steroid hormones.

Amino Acid Sequence↗