Biomedical subjects

Shoshi Kikuchi

Publications and source records attributed to Shoshi Kikuchi.

9 recordsLinked to original sources

PubMed2003

Collection, mapping, and annotation of over 28,000 cDNA clones from japonica rice.

We collected and completely sequenced 28,469 full-length complementary DNA clones from Oryza sativa L. ssp. japonica cv. Nipponbare. Through homology searches of publicly available sequence data, we assigned tentative protein functions to 21,596 clones (75.86%). Mapping of the cDNA clones to genomic DNA revealed that there are 19,000 to 20,500 transcription units in the rice genome. Protein informatics analysis against the InterPro database revealed the existence of proteins presented in rice but not in Arabidopsis. Sixty-four percent of our cDNAs are homologous to Arabidopsis proteins.

Alternative Splicing↗

PubMed2003

Levels of active oxygen species are controlled by ascorbic acid and anthocyanin in Arabidopsis.

Stabilization of the levels of active oxygen species (AOS) is important to the survival of organisms. To clarify the system controlling levels of AOS in plants, this study used an electron spin resonance (ESR) method to directly measure superoxide radical (O(2)(.-)) scavenging activities in the wild-type Arabidopsis thaliana (Col and Ler ecotypes), two anthocyanin mutants (tt3 and ttg1), and an ascorbic acid mutant (vtc1). Under ordinary growth conditions, Arabidopsis contained superoxide-scavenging activity (SOSA) of approximately 300-500 SOD units/g of fresh weight. The ESR pattern indicated that most (40-50%) of this activity was due to ascorbic acid. For the analysis of SOSA under conditions of oxidative stress, synthesis of AOS was induced by gamma-irradiation. The radical scavenging activity in irradiated plants increased approximately 10-fold following an associated increase in the accumulation of ascorbic acid and anthocyanin. The accumulation of ascorbic acid and anthocyanin was suppressed by treatment with an antioxidant before irradiation and was induced by treatment with a radical-generating reagent. The contributions of ascorbic acid and anthocyanin to the total superoxide radical scavenging activity differed among ecotypes. In the Ler ecotype, ascorbic acid accumulated at twice the level of that in the Col ecotype, and induction of anthocyanin was half that in Col. To confirm the activity of ascorbic acid and anthocyanin against AOS stress, the viability of the wild type and mutants (tt2, tt3,tt5, ttg1, and vtc1) was examined after gamma-irradiation. Only the plants in which ascorbic acid and anthocyanin were induced had the ability to grow and flower.

Anthocyanins↗

PubMed2003

Rice gene expression in response to N-acetylchitooligosaccharide elicitor: comprehensive analysis by DNA microarray with randomly selected ESTs.

N-acetylchitooligosaccharides are potent elicitors to suspension-cultured rice cells, inducing a set of defense reactions. Expression of defense-related genes is considered to play an important role in defense reactions, and we employed microarray analysis of 8987 randomly selected expressed sequence tags to analyze the changes in gene expression caused by N-acetylchitooctaose. In this experiment, 166 genes were significantly induced and 93 genes were repressed. RNA gel blot analysis of 16 of these genes confirmed the microarray results. Of the 259 ESTs identified as responsive to N-acetylchytooctaose, 18 genes are related to signal transduction, including five calcium-dependent protein kinases (CDPKs). Among these, three novel CDPKs responsive to N-acetylchitooctaose were isolated.

Amino Acid Sequence↗

PubMed2003

Novel gene encoding a Ca2+-binding protein and under hexokinase-dependent sugar regulation.

A cDNA encoding a predicted 15-kDa protein was earlier isolated from sugar-induced genes in rice embryos (Oryza sativa L.) by cDNA microarray analysis. Here we report that this cDNA encodes a novel Ca2+-binding protein, named OsSUR1 (for Oryza sativa sugar-up-regulated-1). The recombinant OsSUR1 protein expressed in Escherichia coli had 45Ca2+-binding activity. Northern analysis showed that the OsSUR1 gene was expressed mainly in the internodes of mature plants and in embryos at an early stage of germination. Expression of the OsSUR1 gene was induced by sugars that could serve as substrates of hexokinase, but expression was not repressed by Ca2+ signaling inhibitors, calmodulin antagonists and inhibitors of protein kinase or protein phosphatase. These results suggested that Os-SUR1 gene expression was stimulated by a hexokinase-dependent pathway not mediated by Ca2+.

Amino Acid Motifs↗

PubMed2002

Visualization by comprehensive microarray analysis of gene expression programs during transdifferentiation of mesophyll cells into xylem cells.

Plants have a unique transdifferentiation mechanism by which differentiated cells can initiate a new program of differentiation. We used a comprehensive analysis of gene expression in an in vitro zinnia (Zinnia elegans L.) culture model system to gather fundamental information about the gene regulation underlying the transdifferentiation of plant cells. In this model, photosynthetic mesophyll cells isolated from zinnia leaves transdifferentiate into xylem cells in a morphogenic process characterized by features such as secondary-wall formation and programmed cell death. More than 8,000 zinnia cDNA clones were isolated from an equalized cDNA library prepared from cultured cells transdifferentiating into xylem cells. Microarray analysis using these cDNAs revealed several types of unique gene regulation patterns, including: the transient expression of a set of genes during cell isolation, presumably induced by wounding; a rapid reduction in the expression of photosynthetic genes and the rapid induction of protein synthesis-associated genes during the first stage; the preferential induction of auxin-related genes during the subsequent stage; and the transient induction of genes closely associated with particular morphogenetic events, including cell-wall formation and degradation and programmed cell death during the final stage. This analysis also revealed a number of previously uncharacterized genes encoding proteins that function in signal transduction, such as protein kinases and transcription factors that are expressed in a stage-specific manner. These findings provide new clues to the molecular mechanisms of both plant transdifferentiation and wood formation.

Asteraceae↗

PubMed2002

cDNA microarray analysis of gene expression during Fe-deficiency stress in barley suggests that polar transport of vesicles is implicated in phytosiderophore secretion in Fe-deficient barley roots.

To acquire Fe from soil, graminaceous plants secrete mugineic acid family phytosiderophores (MAs) from their roots. The secretion of MAs increases in response to Fe deficiency, and shows a distinct diurnal rhythm. We used a microarray that included 8987 cDNAs of rice EST clones to examine gene expression profiles in barley roots during Fe-deficiency stress. Approximately 200 clones were identified as Fe-deficiency-inducible genes, of which seven had been identified previously. In order to meet the increased demand for methionine to produce MAs, Fe-deficiency enhances the expression of genes that participate in methionine synthesis, as well as recycling methionine through the Yang cycle. Of these 200 genes, approximately 50 exhibited different transcription levels in Fe-deficient roots at noon and at night. Northern blot analysis of time course experiments confirmed that five of these genes exhibited a diurnal change in their level of expression. The diurnal changes in the expression of these genes suggest that polar vesicle transport is involved in the diurnal secretion of MAs.

Azetidinecarboxylic Acid↗

PubMed2002

Construction of a specialized cDNA library from plant cells isolated by laser capture microdissection: toward comprehensive analysis of the genes expressed in the rice phloem.

Laser capture microdissection (LCM) is a powerful system which allows the isolation of selectively targeted cells from a tissue section for the analysis of gene-expression profiles of individual cells. The technique has been successfully used for the isolation of specific mammalian cells, mainly cancer cells. However, LCM has never been reported to be applied to the gene expression analysis of plant cells. We used a modified LCM system and successfully applied it to target and isolate phloem cells of rice leaf tissue whose morphology is apparently different from the surrounding cells. Total RNA was extracted from microdissected (approximately 150) phloem cells and the isolated RNA was used for the construction of a cDNA library following the T7 RNA polymerase amplification. Sequence analysis of 413 randomly chosen clones from the library revealed that there was a high level of redundancy in the population and the clones could be subclassified into 124 different groups that contained related sequences. Approximately 37% of both the redundant population and the non-redundant subgroups had novel components while approximately 63% were either homologues to the known genes reported to be localized in phloem of different plant species, or were homologues to other known genes. In situ hybridization revealed that putative amino acid permease, one of the non-redundant clones, was specifically expressed in the phloem. The results proved the effectiveness of construction of a specialized cDNA library from the specific plant cells.

Amino Acid Transport Systems↗

PubMed2002

A computer-based method of selecting clones for a full-length cDNA project: simultaneous collection of negligibly redundant and variant cDNAs.

We describe a computer-based method that selects representative clones for full-length sequencing in a full-length cDNA project. Our method classifies end sequences using two kinds of criteria, grouping, and clustering. Grouping places together variant cDNAs, family genes, and cDNAs with sequencing errors. Clustering separates those cDNA clones into distinct clusters. The full-length sequences of the clones selected by grouping are determined preferentially, and then the sequences selected by clustering are determined. Grouping reduced the number of rice cDNA clones for full-length sequencing to 21% and mouse cDNA clones to 25%. Rice full-length sequences selected by grouping showed a 1.07-fold redundancy. Mouse full-length sequences showed a 1.04-fold redundancy, which can be reduced by approximately 30% from the selection using our previous method. To estimate the coverage of unique genes, we used FANTOM (Functional Annotation of RIKEN Mouse cDNA Clones) clusters (). Grouping covered almost all unique genes (93% of FANTOM clusters), and clustering covered all genes. Therefore, our method is useful for the selection of appropriate representative clones for full-length sequencing, thereby greatly reducing the cost, labor, and time necessary for this process.

Animals↗

PubMed2002

Isolation and characterization of a rice dwarf mutant with a defect in brassinosteroid biosynthesis.

We have isolated a new recessive dwarf mutant of rice (Oryza sativa L. cv Nipponbare). Under normal growth conditions, the mutant has very short leaf sheaths; has short, curled, and frizzled leaf blades; has few tillers; and is sterile. Longitudinal sections of the leaf sheaths revealed that the cell length along the longitudinal axis is reduced, which explains the short leaf sheaths. Transverse sections of the leaf blades revealed enlargement of the motor cells along the dorsal-ventral axis, which explains the curled and frizzled leaf blades. In addition, the number of crown roots was smaller and the growth of branch roots was weaker than those in the wild-type plant. Because exogenously supplied brassinolide considerably restored the normal phenotypes, we designated the mutant brassinosteroid-dependent 1 (brd1). Further, under darkness, brd1 showed constitutive photomorphogenesis. Quantitative analyses of endogenous sterols and brassinosteroids (BRs) indicated that BR-6-oxidase, a BR biosynthesis enzyme, would be defective. In fact, a 0.2-kb deletion was detected in the genomic region of OsBR6ox (a rice BR-6-oxidase gene) in the brd1 mutant. These results indicate that BRs are involved in many morphological and physiological processes in rice, including the elongation and unrolling of leaves, development of tillers, skotomorphogenesis, root differentiation, and reproductive growth, and that the defect of BR-6-oxidase caused the brd1 phenotype.

Amino Acid Sequence↗