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Kunio Inoue

Publications and source records attributed to Kunio Inoue.

12 recordsLinked to original sources
PubMed2006

Zebrafish MiR-430 promotes deadenylation and clearance of maternal mRNAs.

MicroRNAs (miRNAs) comprise 1 to 3% of all vertebrate genes, but their in vivo functions and mechanisms of action remain largely unknown. Zebrafish miR-430 is expressed at the onset of zygotic transcription and regulates morphogenesis during early development. By using a microarray approach and in vivo target validation, we find that miR-430 directly regulates several hundred target messenger RNA molecules (mRNAs). Most targets are maternally expressed mRNAs that accumulate in the absence of miR-430. We also show that miR-430 accelerates the deadenylation of target mRNAs. These results suggest that miR-430 facilitates the deadenylation and clearance of maternal mRNAs during early embryogenesis.

3' Untranslated Regions↗
PubMed2005

Bruno-like protein is localized to zebrafish germ plasm during the early cleavage stages.

Maternally supplied germ plasm is essential for germ lineage establishment in many species, but the molecular details are still largely unknown, especially in vertebrates, and identification of novel factors that localize to germ plasm is desirable. We previously reported that one of the components of zebrafish germ plasm is mRNA of the bruno-like (brul) gene, a homologue of bruno, which, in Drosophila, is known to participate in germ lineage establishment. Here, we show that not only mRNA but also protein of brul is localized to the zebrafish germ plasm at the ends of the cleavage furrows. In 4- and 8-cell stage embryos, Brul protein is localized to the periphery of the blastomeres, as well as to the ends of the cleavage furrows, forming numerous minute particles. These particles appear at the cortex of the fertilized egg within 10 min after fertilization. Surprisingly, these distinctive localizations, as well as the minute particles, completely disappeared by the 16-cell stage, although relatively weak expression was detected ubiquitously throughout embryogenesis. This is the first report of a protein that localizes to the germ plasm in zebrafish.

Animals↗
PubMed2005

Splicing potentiation by growth factor signals via estrogen receptor phosphorylation.

Mitogen-activated protein kinase-mediated growth factor signals are known to augment the ligand-induced transactivation function of nuclear estrogen receptor alpha (ERalpha) through phosphorylation of Ser-118 within the ERalpha N-terminal transactivation (activation function-1) domain. We identified the spliceosome component splicing factor (SF)3a p120 as a coactivator specific for human ERalpha (hERalpha) activation function-1 that physically associated with ERalpha dependent on the phosphorylation state of Ser-118. SF3a p120 potentiated hERalpha-mediated RNA splicing, and notably, the potentiation of RNA splicing by SF3a p120 depended on hER Ser-118 phosphorylation. Thus, our findings suggest a mechanism by which growth factor signaling can regulate gene expression through the modulation of RNA splicing efficiency via phosphorylation of sequence-specific activators, after association between such activators and the spliceosome.

Animals↗
PubMed2005

The formation of primordial germ cells from germline cells in spherical embryos derived from the blastodisc of 2-cell embryos in goldfish, Carassius auratus.

The property of primordial germ cells (PGCs) in fragmented goldfish embryos was investigated. When 1- and 2- cell embryos were cut at several perpendicular levels at the animal-vegetal axis, cells expressing vas mRNA were observed in the resultant embryos derived from all kinds of animal fragments. Blastodisc fragments from the 1- to 2-cell stage developed to spherical embryos containing yolk body with a yolk syncytial layer (YSL). Germ ring and no tail expression were not observed in the spherical embryo. When the spherical embryo labeled with tracer dye or GFP-nos1 3'UTR mRNA was transplanted onto the animal part of the blastoderm in a host embryo at the blastula stage, PGCs of spherical embryo origin were detected around the gonadal ridges in the resultant embryos which developed normally. These results suggest that small animal fragments should contain factors sufficient for PGC differentiation and that PGCs differentiate without mesoderm induction, since mesoderm is not induced in a spherical embryo.

Animals↗
PubMed2004

TAP/NXF1, the primary mRNA export receptor, specifically interacts with a neuronal RNA-binding protein HuD.

Hu proteins are RNA-binding proteins that are implicated in the control of stabilization, nuclear export, and/or translation of specific mRNAs with AU-rich elements (AREs) in the 3'-untranslated region. Three neuron-specific Hu proteins (HuD, HuB, and HuC), but not a ubiquitously expressed Hu protein HuR, have an activity to induce neurite outgrowth when they are overexpressed in a rat neuronal cell line PC12. Here we show that TAP/NXF1, the primary export receptor for the bulk mRNA, is a specific binding partner for HuD. In vitro binding experiments using recombinant proteins revealed that the interaction between TAP and HuD is direct and that HuD can form a ternary complex together with both TAP and RNA. Interestingly, HuR does not interact with TAP. These results suggest that HuD acts as a novel adaptor protein to recruit TAP for efficient export of ARE-containing mRNAs in neuronal cells.

Animals↗
PubMed2004

Localized maternal factors are required for zebrafish germ cell formation.

Maternally supplied factors in fertilized eggs play essential roles in the establishment of primordial germ cells. In zebrafish, cytoplasm at the distal ends of the first and second cleavage furrows has been assumed to contain germ lineage determinants, since maternal transcripts of germ lineage-specific genes are localized to ends of the cleavage furrows. To investigate whether these parts of cytoplasm are required for germ cell formation, we removed all four regions of the cytoplasm by glass capillary at the 4-cell stage. Histological analysis revealed that the ablation of cytoplasm at the ends of the cleavage planes resulted in a severe reduction in the number of germ cells. In addition, the expression of germ lineage markers was eliminated by cytoplasmic ablation. These results demonstrated that cytoplasm at the distal ends of cleavage furrows is essential for germ cell formation. We also found novel localization patterns for zDazl and brul mRNAs along the cleavage planes. Our findings provide the first direct evidence that localized cytoplasmic factors are indispensable for germ cell establishment in zebrafish.

Animals↗
PubMed2004

RBD-1, a nucleolar RNA-binding protein, is essential for Caenorhabditis elegans early development through 18S ribosomal RNA processing.

RBD-1 is the Caenorhabditis elegans homolog of Mrd1p, which was recently shown to be required for 18S ribosomal RNA (rRNA) processing in yeast. To gain insights into the relationship between ribosome biogenesis and the development of multicellular organisms, we examined the expression and function of RBD-1. Maternal RBD-1 in the fertilized egg disappears immediately after cleavage starts, whereas zygotic RBD-1 first appears in late embryos and is localized in the nucleolus in most cells, although zygotic transcription of pre-rRNA is known to be initiated as early as the one-cell stage. RNA interference of the rbd-1 gene severely inhibits the processing of 18S rRNA in association with various developmental abnormalities, indicating its essential role in pre-rRNA processing and development in C.elegans. These results provide evidence for the linkage between ribosome biogenesis and the control of development and imply unexpected uncoupling of transcription and processing of pre-rRNA in early C.elegans embryos.

Animals↗
PubMed2003

A vertebrate RNA-binding protein Fox-1 regulates tissue-specific splicing via the pentanucleotide GCAUG.

Alternative splicing is one of the central mechanisms that regulate eukaryotic gene expression. Here we report a tissue-specific RNA-binding protein, Fox-1, which regulates alternative splicing in vertebrates. Fox-1 bound specifically to a pentanucleotide GCAUG in vitro. In zebrafish and mouse, fox-1 is expressed in heart and skeletal muscles. As candidates for muscle-specific targets of Fox-1, we considered two genes, the human mitochondrial ATP synthase gamma-subunit gene (F1gamma) and the rat alpha-actinin gene, because their primary transcripts contain several copies of GCAUG. In transfection experiments, Fox-1 induced muscle-specific exon skipping of the F1gamma gene via binding to GCAUG sequences upstream of the regulated exon. Fox-1 also regulated mutually exclusive splicing of the alpha-actinin gene, antagonizing the repressive effect of polypyrimidine tract-binding protein (PTB). It has been reported that GCAUG is essential for the alternative splicing regulation of several genes including fibronectin. We found that Fox-1 promoted inclusion of the fibronectin EIIIB exon. Thus, we conclude that Fox-1 plays key roles in both positive and negative regulation of tissue-specific splicing via GCAUG.

Actinin↗
PubMed2002

Regulation of alternative splicing of alpha-actinin transcript by Bruno-like proteins.

BACKGROUND: The Bruno-like or CELF proteins, such as mammalian CUGBP1 and Etr-3, Xenopus EDEN-BP, and Drosophila Bruno (Bru), are regulators of gene expression at the post-transcriptional level, and contain three RNA-recognition motifs (RRMs). It has been shown that mammalian CUGBP1 and Etr-3 regulate alternative splicing of cardiac troponin T pre-mRNA via binding to CUG-triplet repeats. RESULTS: Using in vitro selection and UV-crosslinking experiments, we found that zebrafish Bruno-like proteins bound to repeat elements of uridine and purine (termed UREs). It is known that non-muscle (NM) and smooth muscle (SM) exons of the rat alpha-actinin gene are used in a mutually exclusive manner. Transfection experiments in mammalian cells showed that zebrafish Brul and Etr-3 induced the muscle-specific splicing of rat alpha-actinin pre-mRNA via binding to the URE at the branch point upstream of the NM exon. In contrast, zebrafish Etr-1 promoted skipping of both the NM and SM exons in a manner which was not dependent on URE-binding. CONCLUSIONS: Our results showed that Bruno-like proteins bind to UREs and regulate the alternative splicing of alpha-actinin pre-mRNA. Members of the Bruno family play multiple roles in splicing regulation.

Actinin↗
PubMed2002

Zebrafish DAZ-like protein controls translation via the sequence 'GUUC'.

BACKGROUND: In many species, DAZ homologous genes encode RNA-binding proteins containing two conserved motifs, namely the RNA-recognition motif (RRM) and the DAZ motif. Genetic analysis and gene disruption studies have demonstrated that DAZ family proteins play important roles in gametogenesis. However, little is known about the biochemical functions of DAZ family proteins. RESULTS: Using in vitro selection and UV-crosslinking experiments, we identified the sequence 'GUUC' as the target RNA sequence of zebrafish DAZ-like protein (zDAZL). In transfection experiments, zDAZL protein activated translation in a manner dependent on the binding sequence in the 3'UTR of the Drosophila twine gene or zDazl gene. Moreover, it is highly likely that the zDAZL protein associates with polysomes through the DAZ motif in vivo, and that the association with polysomes is indispensable for translational activation. CONCLUSIONS: This is the first report that the DAZ family protein directly promotes the translation of the target mRNAs in vertebrates. This study provides important insights into the molecular mechanisms underlying the post-transcriptional regulation of DAZ family proteins in gametogenesis.

3' Untranslated Regions↗
PubMed2002

The germ cell lineage identified by vas-mRNA during the embryogenesis in goldfish.

vas RNA has been identified in germ-line cells and its precursors in zebrafish, with the result that the germ-line lineage can be traced throughout embryogenesis. In the present study, we described vas localization and the migration of vas-positive cells in goldfish, using whole mount in situ hybridization. The signals of vas mRNA localization appeared at the marginal part of the first to third cleavage planes. The eight signals were detected during the period from the 8- cells to the 512-cell stage. At the late-blastula stage, additional numbers of vas-positive cells were observed, suggesting the proliferation of these cells. At the segmentation period, vas-positive cells showed a long extended distribution along the embryonic axis, but did not form any clusters. vas-positive cells were occasionally distributed at the head region, especially around the future otic vesicle. These signals were inherited to the primordial germ cells, suggesting that vas-positive cells were primordial germ cells (PGCs) in goldfish.

Animals↗