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Y Sakaki

Publications and source records attributed to Y Sakaki.

At least 73 records · Page 4Linked to original sources
PubMed1999

Spatiotemporal, allelic, and enforced expression of Ximpact, the Xenopus homolog of mouse imprinted gene impact.

Mouse Impact is an imprinted gene encoding an evolutionarily conserved protein of unknown function. We isolated cDNA for the Xenopus homolog of Impact (Ximpact), since the clawed frog not only provides an excellent model for the study of gene function in early development but also allows the generation of interspecific F1 hybrids required for the examination of allelic expression status. The predicted product of Ximpact shows an extreme sequence similarity to those of mouse Impact and its homologs in nematoda, fission yeast, and budding yeast. The transcript of Ximpact is present in oocytes as well as in early embryos, and its spatial distribution is ubiquitous in both embryonic and adult stages. An RT-PCR-RFLP assay using the reciprocal interspecific F1 hybrids and a DNA polymorphism between X. laevis and X. borealis showed that Ximpact is expressed biallelically when analyzed as a whole embryo. Overexpression of Ximpact by RNA microinjection resulted in a higher than normal rate of gastrulation defects, suggesting the need for tight control of its dosage in early development.

Alleles↗
PubMed1999

Inhibition of light- or glutamate-induced mPer1 expression represses the phase shifts into the mouse circadian locomotor and suprachiasmatic firing rhythms.

mPer1, a mouse gene, is a homolog of the Drosophila clock gene period and has been shown to be closely associated with the light-induced resetting of a mammalian circadian clock. To investigate whether the rapid induction of mPer1 after light exposure is necessary for light-induced phase shifting, we injected an antisense phosphotioate oligonucleotide (ODN) to mPer1 mRNA into the cerebral ventricle. Light-induced phase delay of locomotor activity at CT16 was significantly inhibited when the mice were pretreated with mPer1 antisense ODN 1 hr before light exposure. mPer1 sense ODN or random ODN treatment had little effect on phase delay induced by light pulses. In addition, glutamate-induced phase delay of suprachiasmatic nucleus (SCN) firing rhythm was attenuated by pretreatment with mPer1 antisense ODN, but not by random ODN. The present results demonstrate that induction of mPer1 mRNA is required for light- or glutamate-induced phase shifting, suggesting that the acute induction of mPer1 mRNA in the SCN after light exposure is involved in light-induced phase shifting of the overt rhythm.

Animals↗
PubMed1999

Inhibition of the NGF and IL-1beta-induced expression of Alzheimer's amyloid precursor protein by antisense oligonucleotides.

Alzheimer's disease (AD) is a neurodegenerative disorder of the brain characterized by the extracellular deposition of amyloid in senile plaques and along the walls of the cerebral vasculature. The principal constituent of amyloid deposit is amyloid beta peptide (Abeta) derived from its larger precursor protein, amyloid precursor protein (APP). The overexpression of APP is known to be a risk factor for Abeta deposit in AD and in Down syndrome (DS). The inhibition of APP expression has been thought to be beneficial to patients with AD and DS. In this study, we investigated the effects of antisense oligonucleotide (AO) on the overexpression of APP induced by IL-1beta and NGF. Using phosphorothioate-oligonucleotides against initiation codon significantly reduced the protein levels of APP induced by NGF and IL-1beta to basal level in PC12 cell culture systems. These results showed that these antisense oligonucleotides may have a potential to be a therapeutic agent for some patients with AD and DS.

Amyloid beta-Protein Precursor↗
PubMed1998

Identification of the mammalian homologues of the Drosophila timeless gene, Timeless1.

We have identified novel mammalian homologues of a Drosophila clock gene, timeless, and designated them as human TIMELESS1 (hTIM1) and mouse Timeless1 (mTim1), respectively. These genes were mapped by FISH to chromosomal regions 12q12-13 in human and 10D3 in mouse. The deduced amino acid sequences of hTim1 and mTim1 proteins were 1208 and 1197 amino acids in length and shared 83% identity. Northern blot analysis identified a single transcript of 4.5 kb expressed widely in many tissues examined. Unlike the Drosophila counterpart, the levels of the mTim1 transcript exhibited no prominent circadian oscillation in the mouse brain.

Animals↗
PubMed1998

Fluorescent differential display analysis of gene expression in apoptotic neuroblastoma cells.

Identification of differentially expressed genes will provide leads in the elucidation of the molecular mechanisms underlying neuronal cell death associated with neurodegenerative disorders. Using a high-throughput fluorescent differential display (FDD) system based on an automated DNA sequencer, we analyzed global patterns of gene expression during the apoptosis of neuroblastoma SH-SY5Y cells induced by a neurotoxin, colchicine. Initial screening of approximately 24000 cDNA bands displayed with 320 primer combinations has revealed 263 fragments showing differential expression patterns, suggesting that approximately 1% of transcripts are modulated in their expression level. Of these differentially displayed bands, we cloned 18 fragments composed of 17 distinct species and confirmed differential expression of each species by reverse transcription-PCR or Northern blot hybridization, thereby proving the reliability of the approach. These include eight derived from seven known genes, five homologous to expressed sequence tags (ESTs), and five totally lacking any homology to those deposited in the database. Among these, a novel transcript SAI1 induced prominently was characterized further and revealed to encode a putative RNA-binding protein NAPOR (neuroblastoma apoptosis-related RNA-binding protein), containing three copies of evolutionarily conserved RNA recognition motif. Since several RNA-binding proteins have been known to play crucial roles in other apoptosis systems, it is conceivable that NAPOR is also involved in the process of neuronal cell death.

Amino Acid Sequence↗
PubMed1998

Cloning and mapping of ZNF231, a novel brain-specific gene encoding neuronal double zinc finger protein whose expression is enhanced in a neurodegenerative disorder, multiple system atrophy (MSA).

A novel brain-specific gene, neuronal double zinc finger protein (ZNF231), was cloned and mapped. We used the high-density cDNA filter method to analyze the gene-expression profile in brains with multiple system atrophy (MSA). MSA is a sporadic progressive neurodegenerative disease characterized clinically by cerebellar symptoms, parkinsonism, autonomic dysfunction, or their various combinations, but its pathogenesis has yet to be clarified. In total, 8300 cDNA clones were screened, and a novel gene, ZNF231, was identified, whose expression was elevated in cerebella of patients with MSA. Its transcript is approximately 16 kb long and encodes an open reading frame of 3926 amino acid residues that has several interesting motifs; two glycine-proline dipeptide repeats (aa 22-32 and aa 61-74), a pair of homologous C8 double zinc finger motifs (aa 169-226 and aa 465-521), a leucine zipper motif (aa 561-582), a SH3 domain-binding motif (aa 825-831), two nuclear targeting signals (aa 1011-1028 and aa 1071-1091), two glutamine-rich domains (aa 2428-2473 and aa 3775-3804), and a histidine-rich domain (aa 3597-3682). These features suggest that the new gene encodes a nuclear protein or transcription regulator. Northern blot and RT-PCR analyses showed that its expression is specific to the brain and apparently restricted to the neurons. Elevation of ZNF231 expression may be involved in the pathogenesis of multiple system atrophy. The gene for ZNF231 is located on chromosome 3p21.

Amino Acid Sequence↗
PubMed1998

Involvement of dendritic cell response to resistance of stomach carcinogenesis caused by N-methyl-N'-nitro-N-nitrosoguanidine in rats.

The involvement of immune response in the resistance of chemically induced stomach cancer was studied in a resistant rat strain (Buffalo) and a sensitive rat strain (ACI). Groups of 10 male Buffalo and ACI rats, 6 weeks of age, were given drinking water with or without N-methyl-N'-nitro-N-nitrosoguanidine (MNNG; 100 mg/l) for 14 days. Total RNA was isolated from the stomach pyloric mucosa from five rats, and cDNA was prepared with reverse transcriptase. Tissue sections of the stomach pyloric mucosa from five rats were stained with antibodies recognizing molecules expressed by various immune cells. Reverse transcription-PCR (RT-PCR), competitive RT-PCR, and Northern blot demonstrated that the expression of MHC class II group genes [MHC class II, MHC class II-associated invariant chain (Ii), CD4 and IgM (B cell marker)], MHC class I group genes (MHC class I and CD8), B7-1 (costimulator on dendritic cells), and CD28 (receptor to B7 on T cells) in the pyloric mucosa was elevated by MNNG in both rat strains but was elevated to a 4-7-fold greater extent in Buffalo rats than in ACI rats. These genes were scarcely expressed in control rats. Histochemical antibody staining after MNNG exposure showed a greater number of cells stained with monoclonal antibody to Ii, OX-62 (dendritic cell marker), and ED-1 (dendritic cell and macrophage common marker) in the interstitial tissue of the pyloric mucosa of Buffalo rats compared with ACI rats. Cell proliferation, as measured by 5-bromo-2-deoxyuridine (BrdUrd)-labeling indices, revealed the presence of BrdUrd-labeled cells only among epithelial cells in the proliferative zone; cells in the interstitial tissue were not labeled with BrdUrd. The results suggest the involvement of dendritic cell response in the resistance to the MNNG induction of stomach carcinogenesis in rats.

Animals↗
PubMed1998

Cataloging altered gene expression during rat hippocampal long-term potentiation by means of differential display.

We have employed mRNA differential display (DD) to generate a catalog of cDNAs whose expression in the hippocampus was regulated during long-term potentiation (LTP) in dentate gyrus of anesthetized rats. DD with 459 combinations of primer pairs revealed that 80 out of approximately 70000 bands displayed showed a reproducible change in their expression level. These cDNAs were categorized into seven groups according to changes in their temporal expression pattern. Some of these cDNAs were induced rapidly, but transiently, after the LTP induction, some induced rapidly and persistently, some induced slowly, and some down-regulated following LTP. This suggests that a complex molecular hierarchy underlies the maintenance of hippocampal LTP.

Animals↗
PubMed1998

Functional modules and expression of mouse p40(phox) and p67(phox), SH3-domain-containing proteins involved in the phagocyte NADPH oxidase complex.

The phagocyte NADPH oxidase is activated during phagocytosis to produce superoxide, a precursor of microbicidal oxidants. The formation of the active oxidase complex at the membrane requires translocation of the Rac GTPase and two specialized cytosolic proteins that harbor SH3 domains, p67phox and p47phox. Another SH3-domain-containing protein p40phox, which is constitutively associated with p67phox in phagocytes, also enters the complex upon cell stimulation. Here we describe how we cloned mouse cDNAs encoding p40phox and its partner in phagocytes, p67phox. Both p40phox and p67phox comprise several protein-binding modules that are structurally and functionally well conserved between mouse and human, indicating their nature as adaptor proteins. We have also systematically investigated expression of the gene for p40phox in comparison with those for p67phox and p47phox. Distributions of the mRNAs for the three proteins among tissues are similar, with the most abundant expression in the spleen. The messages are abundant not only in phagocytic cells, but also in B cell lineage. The p40phox gene, but not the other two, is expressed in some types of cells such as plasma cells and T lymphocytes. Furthermore, in situ hybridization analysis shows that the p40phox mRNA is distributed in neuronal cells of mouse brain, providing evidence that one of the genes for the specialized oxidase factors is expressed in neurons. These observations raise the possibility that the adaptor protein p40phox plays a heretofore unsuspected role via interacting with other proteins in the cells that do not express p67phox or p47phox.

Amino Acid Sequence↗
PubMed1998

The PC motif: a novel and evolutionarily conserved sequence involved in interaction between p40phox and p67phox, SH3 domain-containing cytosolic factors of the phagocyte NADPH oxidase.

The superoxide-generating NADPH oxidase, dormant in resting phagocytes, is activated during phagocytosis following assembly of the membrane-integrated protein cytochrome b558 and cytosolic factors. Among the latter are the three proteins containing Src homology 3 (SH3) domains, p67phox, p47phox and p40phox. While the first two factors are indispensable for the activity, p40phox is tightly associated with p67phox in resting cells and is suggested to have some modulatory role. Here we describe a systematic analysis of the interaction between p40phox and p67phox using the yeast two-hybrid system and in vitro binding assays with recombinant proteins. Both methods unequivocally showed that the minimum requirements for stable interaction are the C-terminal region of p40phox and the region between the two SH3 domains of p67phox. This interaction is maintained even in the presence of anionic amphiphiles used for the activation of the NADPH oxidase, raising a possibility that it mediates constitutive association of the two factors in both resting and activated cells. The C-terminal region of p40phox responsible for the interaction contains a characteristic stretch of amino acids designated as the PC motif, that also exists in other signal-transducing proteins from yeast to human. Intensive site-directed mutagenesis to the motif in p40phox revealed that it plays a critical role in the binding to p67phox. Thus the PC motif appears to represent a novel module for protein-protein interaction used in a variety of signaling pathways.

Amino Acid Sequence↗
PubMed1998

Presenilin 1 mutations linked to familial Alzheimer's disease increase the intracellular levels of amyloid beta-protein 1-42 and its N-terminally truncated variant(s) which are generated at distinct sites.

Mutations in the presenilin genes PS1 and PS2 cause the most common form of early-onset familial Alzheimer's disease. The influence of PS1 mutations on the generation of endogenous intracellular amyloid beta-protein (A beta) species was assessed using a highly sensitive immunoblotting technique with inducible mouse neuroblastoma (Neuro 2a) cell lines expressing the human wild-type (wt) or mutated PS1 (M146L or delta exon 10). The induction of mutated PS1 increased the intracellular levels of two distinct A beta species ending at residue 42 that were likely to be A beta1-42 and its N-terminally truncated variant(s) A beta x-42. The induction of mutated PS1 resulted in a higher level of intracellular A beta1-42 than of intracellular A beta x-42, whereas extracellular levels of A beta1-42 and A beta x-42 were increased proportionally. In addition, the intracellular generation of these A beta42 species in wt and mutated PS1-induced cells was completely blocked by brefeldin A, whereas it exhibited differential sensitivities to monensin: the increased accumulation of intracellular A beta x-42 versus inhibition of intracellular A beta1-42 generation. These data strongly suggest that A beta x-42 is generated in a proximal Golgi, whereas A beta1-42 is generated in a distal Golgi and/or a post-Golgi compartment. Thus, it appears that PS1 mutations enhance the degree of 42-specific gamma-secretase cleavage that occurs in the normal beta-amyloid precursor protein processing pathway (a) in the endoplasmic reticulum or the early Golgi apparatus prior to beta-secretase cleavage or (b) in the distinct sites where A beta x-42 and A beta1-42 are generated.

Alzheimer Disease↗
PubMed1998

Molecular characterization of the mouse mtprd gene, a homologue of human TPRD: unique gene expression suggesting its critical role in the pathophysiology of Down syndrome.

We and others recently isolated a human TPRD gene, possessing a motif of the tetratricopeptide repeat (TPR), from the Down syndrome-critical region (DCR) of chromosome 21q22.2. In this study, we isolated a mouse homologue of TPRD cDNA, mtprd, and examined its expression profile in mouse embryos. The gene was mapped to mouse chromosome 16C3.3-4, consistent with the location of DCR, and encodes 1,979 amino acid residues with 76% identity to TPRD. The mtprd protein has three units of the TPR motif with 91% homology to TPRD. The protein also has two regions homologous to several matrix proteins with 86 and 70% identities to those of TPRD. Several splicing variants of the 5' portion of the open reading frame of mtprd were identified by RT-PCR and sequencing of mRNAs. In situ hybridization showed that mtprd is ubiquitously expressed in mouse embryos but predominantly in the central nervous system, including the telencephalon, mesencephalon, and metencephalon. These results suggest that the TPRD gene is one of the genes responsible for not only the morphological anomalies but also the neurological abnormalities observed in Down syndrome. The presence of splicing variants indicates that the protein may also have several isoforms in mice.

Amino Acid Sequence↗
PubMed1998

Genomic structure, sequence, and refined mapping of the human intersectin gene (ITSN), which encompasses 250 kb on chromosome 21q22.1-->q22.2.

The ubiquitously expressed and brain-specific human intersectin (ITSN) isoforms are scaffold proteins probably involved in general endocytosis and synaptic vesicle recycling, respectively. Here, analysis of 21q22.1-->q22.2 genomic sequence revealed that ITSN consists of 41 exons spanning approximately 250 kb and maps between GART and D21S325. The probable function of the ITSN isoforms and mapping position of ITSN suggest that disproportionate expression of this gene may be implicated in the phenotypic characteristics of Down syndrome.

Adaptor Proteins, Vesicular Transport↗
PubMed1997

A novel gene isolated from human placenta located in Down syndrome critical region on chromosome 21.

Down syndrome is the most common birth defect, which is caused by trisomy 21. We identified a novel gene in the so-called Down syndrome critical region by EST mapping to genomic DNA and following cDNA cloning. The gene, designated DCRB (Down syndrome Critical Region gene B), consisted of three exons of 1095 bp in total and encoded a large open reading frame of 118 amino acid residues. The amino acids sequence of DCRB showed no significant homology to any known protein. Northern blot analysis showed that DCRB is mainly expressed in the placenta, in which a major 1.1-kb band and a minor 2.0-kb band were detected. Minor bands of 1.4 kb and 2.2 kb were also detected in adult heart and skeletal muscle.

Adult↗
PubMed1997

Circadian oscillation of a mammalian homologue of the Drosophila period gene.

Many biochemical, physiological and behavioural processes in organisms ranging from microorganisms to vertebrates exhibit circadian rhythms. In Drosophila, the gene period (per) is required for the circadian rhythms of locomotor activity and eclosion behaviour. Oscillation in the levels of per mRNA and Period (dPer) protein in the fly brain is thought to be responsible for the rhythmicity. However, no per homologues in animals other than insects have been identified. Here we identify the human and mouse genes (hPER and mPer, respectively) encoding PAS-domain (PAS, a dimerization domain present in Per, Amt and Sim)-containing polypeptides that are highly homologous to dPer. Besides this structural resemblance, mPer shows autonomous circadian oscillation in its expression in the suprachiasmaticnucleus, which is the primary circadian pacemaker in the mammalian brain. Clock, a mammalian clock gene encoding a PAS-containing polypeptide, has now been cloned: it is likely that the Per homologues dimerize with other molecule(s) such as Clock through PAS-PAS interaction in the circadian clock system.

Amino Acid Sequence↗
PubMed1997

Isolation and expression study of a maternally expressed novel Xenopus gene Xem1 encoding a putative evolutionarily conserved membrane protein.

A novel Xenopus maternally expressed gene, Xem1, was isolated by differential display PCR and 5'-RACE. Xem1 coded for a putative transmembrane protein of 172 amino acids. Sequence analysis, including the clustering and reconstruction of ESTs (Expressed Sequence Tags), revealed that homologs of Xem1 are widely distributed in eukaryotic phyla, suggesting that Xem1 is a member of evolutionarily conserved proteins. Expression of Xem1 mRNA occurred from the previtellogenic stage and its level increased during oogenesis, maintained throughout oocyte maturation to blastula stage and then decreased in post gastrula stages. In cleavage stage, Xem1 RNA was distributed uniformly, and in adult, occurred predominantly in ovary and testis. We assume that Xenopus Xem1 may have its function in gametogenesis and in early phase of embryogenesis, whose function may be related to transport mechanism of small molecular weight substances like metal ions, from analogy to the function of its homologs in other organisms.

Amino Acid Sequence↗