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Komei Shirabe

Publications and source records attributed to Komei Shirabe.

9 recordsLinked to original sources
PubMed2006

Molecular cloning and expression of two small leucine-rich proteoglycan (SLRP) genes, dspg3l and optcl, in zebrafish.

Epiphycan (DSPG3) and opticin are two class III small leucine-rich proteoglycans (SLRP). We isolated two zebrafish cDNAs, dspg3l and optcl, that encode proteins homologous to epiphycan and opticin in other vertebrates. Like epiphycans in other species, dspg3l is exclusively expressed in the developing notochord and cartilage. optcl is expressed transiently in the developing nervous system, eyes and somites much like opticin. The zebrafish dspg3l and optcl genes are located in linkage group 4 and 11, respectively. The genomic locations for both genes in zebrafish are syntenic with the genomic locations of dspg3 and opticin (optc) in human and mouse. Synteny and the expression patterns of these genes suggest that the dspg3l and optcl are the orthologs to the mammalian dspg3 and optc genes, respectively.

Amino Acid Sequence↗
PubMed2006

Characterization and expression of serotonin transporter genes in zebrafish.

To understand the development of serotonergic neurons in vertebrates, we used zebrafish as a model system. In this study we cloned two cDNAs (complementary DNAs) coding for serotonin transporter (SERT) from the zebrafish, named serta and sertb. The serta cDNA encodes a protein of 693 amino acids and showed high level of sequence identity with rat and human SERTs. In situ hybridization showed serta to be expressed in raphe nuclei, ventral posterior tuberculum and pineal organ. The expression of serta in raphe and ventral posterior tuberculum overlapped with the location of serotonin and expression of tryptophan hydroxylase, which is a key enzyme for serotonin synthesis. In the pineal organ serta is expressed in the cells in the vicinity of tryptophan hydroxylase-positive cells. We also cloned another zebrafish serotonin transporter, sertb, and found to be expressed in the medulla oblongata and in the inner nuclear layer of retina. The existence of two sert genes in the zebrafish genome indicates the gene was duplicated in the process of evolution as can be seen in other genes in the teleosts including zebrafish. The expression of the serta cDNA in cultured cells conferred a serotonin transport activity, thus indicating the validity of the cloned cDNA. We have established the expression system of zebrafish serotonin transporter in the cell culture in the present study, which is useful for the pharmacological analysis to determine the important residues for the interaction with serotonin and inhibitors. The expression system in the cell culture can be used to determine the effective concentration of inhibitors and addictive drugs. These information might be useful to evaluate the effect of those chemicals on serotonin neuron development and behavior of the animal.

Amino Acid Sequence↗
PubMed2006

Multifactor complex containing B element binding factor, BBF, and repressors regulate the human alpha 1(III) collagen gene (COL3A1).

Type III collagen is found in fetal skin and blood vessels. Previously, we characterized the proximal promoter of the human alpha1(III) collagen gene (COL3A1) using the human rhabdomyosarcoma cell line, A204, and NIH3T3 cells (Yoshino et al., Biochim Biophys Acta, 2005). In the present study, we further analyzed this promoter using additional cell lines, namely a human embryonal rhabdomyosarcoma cell line (RD) and bovine vascular smooth muscle cells (vSMCs), both of which show high expression of type III collagen. Using a luciferase assay, electrophoretic mobility shift assays (EMSA), and DNase footprinting assay, 2 types of multifactor complexes were shown to bind to the DNA region in the vicinity of the B element (- 80 to - 58), depending on the cell type. Next, we used cells stably transfected with a GFP-linked type III collagen promoter fragment for analysis of promoter expression. Usually, transfected cells retained the characteristics of the original cells. However, in several clones derived from RD cells, promoter expression as well as cell shape changed to patterns characteristic of the A204 cell line. Nuclear factors expressed by these clones were also characteristic of the A204 line.

Animals↗
PubMed2005

MN-cadherin and its novel variant are transiently expressed in chick embryo spinal cord.

To isolate cDNAs that are involved in limb-motoneuron development, we compared mRNAs of lumbar and thoracic motoneurons purified from spinal cord of E4 chick embryo by differential display. In situ hybridization demonstrated that one of cDNAs is expressed exclusively in lateral motor column in spinal cord from E4 to E10. We identified two mRNA variants for the cDNA by library screening. The long form (788 amino acids) was identical to chick MN-cadherin. The short variant (543 amino acids) lacks the first two of five extracellular domains of MN-cadherin, which commonly exist in classical cadherins. The amino acid sequence of the short form is identical to that of the carboxyl terminal MN-cadherin, except for the distinct signal sequence. The ratio of mRNA of short form to long form was 1-20. cDNA transfection study revealed that the long form but not the short form MN-cadherin had cell adhesion activity.

Amino Acid Sequence↗
PubMed2005

Chemokine signaling guides axons within the retina in zebrafish.

Chemokines are a large family of secreted proteins that play an important role in the migration of leukocytes during hematopoiesis and inflammation. Chemokines and their receptors are also widely distributed in the CNS. Although recent investigations are beginning to elucidate chemokine function within the CNS, relatively little is known about the CNS function of this important class of molecules. To better appreciate the CNS function of chemokines, the role of signaling by stromal cell-derived factor-1 (SDF-1) through its receptor, chemokine (CXC motif) receptor 4 (CXCR4), was analyzed in zebrafish embryos. The SDF-1/CXCR4 expression pattern suggested that SDF-1/CXCR4 signaling was important for guiding retinal ganglion cell axons within the retina to the optic stalk to exit the retina. Antisense knockdown of the ligand and/or receptor and a genetic CXCR4 mutation both induced retinal axons to follow aberrant pathways within the retina. Furthermore, retinal axons deviated from their normal pathway and extended to cells ectopically expressing SDF-1 within the retina. These data suggest that chemokine signaling is both necessary and sufficient for directing retinal growth cones within the retina.

Animals↗
PubMed2004

Structure of human erythrocyte NADH-cytochrome b5 reductase.

Erythrocyte NADH-cytochrome b(5) reductase reduces methaemoglobin to functional haemoglobin. In order to examine the function of the enzyme, the structure of NADH-cytochrome b(5) reductase from human erythrocytes has been determined and refined by X-ray crystallography. At 1.75 A resolution, the root-mean-square deviations (r.m.s.d.) from standard bond lengths and angles are 0.006 A and 1.03 degrees , respectively. The molecular structure was compared with those of rat NADH-cytochrome b(5) reductase and corn nitrate reductase. The human reductase resembles the rat reductase in overall structure as well as in many side chains. Nevertheless, there is a large main-chain shift from the human reductase to the rat reductase or the corn reductase caused by a single-residue replacement from proline to threonine. A model of the complex between cytochrome b(5) and the human reductase has been built and compared with that of the haem-containing domain of the nitrate reductase molecule. The interaction between cytochrome b(5) and the human reductase differs from that of the nitrate reductase because of differences in the amino-acid sequences. The structures around 15 mutation sites of the human reductase have been examined for the influence of residue substitutions using the program ROTAMER. Five mutations in the FAD-binding domain seem to be related to cytochrome b(5).

Amino Acid Sequence↗
PubMed2004

Chemokine signaling regulates sensory cell migration in zebrafish.

Chemokines play an important role in the migration of a variety of cells during development. Recent investigations have begun to elucidate the importance of chemokine signaling within the developing nervous system. To better appreciate the neural function of chemokines in vivo, the role of signaling by SDF-1 through its CXCR4 receptor was analyzed in zebrafish. The SDF-1-CXCR4 expression pattern suggested that SDF-1-CXCR4 signaling was important for guiding migration by sensory cells known as the migrating primordium of the posterior lateral line. Ubiquitous induction of the ligand in transgenic embryos, antisense knockdown of the ligand or receptor, and a genetic receptor mutation all disrupted migration by the primordium. Furthermore, in embryos in which endogenous SDF-1 was knocked down, the primordium migrated towards exogenous sources of SDF-1. These data demonstrate that SDF-1 signaling mediated via CXCR4 functions as a chemoattractant for the migrating primordium and that chemokine signaling is both necessary and sufficient for directing primordium migration.

Animals↗
PubMed2002

Identification and characterization of a soluble cadherin-7 isoform produced by alternative splicing.

We identified an alternative mRNA encoding a novel cadherin-7 isoform by reverse transcriptase-PCR of RNA from day 12 chicken embryos. The alternative mRNA contains 49 bases of insertion in the premembrane region, leading to the substitution of 14 amino acids and the introduction of a premature stop codon. Identification of a 49-bp insertion sequence in the genomic DNA corresponding to the intron of the cadherin-7 gene suggests that alternative splicing is the cause of the alternative mRNA. Transient expression of the variant form in COS-7 or 293 cells produced a soluble protein. Aggregation assays and immunoprecipitation showed that the variant protein interacts with full-length cadherin-7 in vitro and in vivo and inhibits full-length cadherin-7-mediated cell adhesion. Immunohistochemistry revealed that the variant form was strongly expressed in dermomyotomes rather than in migrating neural crest cells, in contrast to the full-length cadherin-7, suggesting differential regulation of splicing and possible roles of variant cadherin-7 in the development of dermomyotomes and other tissues.

Alternative Splicing↗