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S Kato

Publications and source records attributed to S Kato.

At least 361 records · Page 20Linked to original sources
PubMed2000

Functional evaluation of p53 and PTEN gene mutations in gliomas.

We screened mutations of two major tumor suppressor genes, p53 and PTEN, in 66 human brain tumors using a yeast-based functional assay and cDNA-based direct sequencing, respectively. The frequency of p53 mutations was 28.8% (19 of 66) and was higher in anaplastic astrocytoma (9 of 14, 64.3%,) than in glioblastoma multiforme (GBM; 7 of 27, 25.9%,), supporting previous speculation that there are at least two genetic pathways leading to GBM, a de novo pathway without p53 mutation and a "progressive" pathway with p53 mutation. PTEN mutation was observed in 8 of 64 tumors (12.5%), mainly GBMs (7 of 26, 26.9%), both with and without p53 mutation. These results suggest that mutation of the PTEN gene is a later event than that of the p53 gene in glioma progression and is associated with both the genetic pathways. All of the detected PTEN missense mutations and an in-frame small deletion inactivated PTEN phosphoinositide phosphatase activity in vitro. Because the tumors containing PTEN mutations also showed loss of heterozygosity in the chromosome 10q23 region flanking the PTEN gene, our data clearly indicate that inactivation of both PTEN alleles occurs in a subset of high-grade gliomas, therefore confirming the previous idea that PTEN acts as a tumor suppressor gene.

Adolescent↗
PubMed2000

[Transcriptional control by nuclear steroid hormone receptors and endocrine disruptants].

Endocrine disruptant is supposed to act as a ligand to modulate function of nuclear steroid hormone receptors. The nuclear receptors form a superfamily compromising nearly hundred members in human including steroid/thyroid hormones, vitamin A and D, other fat-soluble compounds nuclear receptors, and a subfamily of orphan receptors. The nuclear receptors act as ligand-inducible transcription factors in concert with co-activators in nuclear complexes. In this review, the functions of nuclear receptors are described in terms of transcriptional controls and co-activator complexes.

Animals↗
PubMed1999

Association of two nuclear proteins, Npw38 and NpwBP, via the interaction between the WW domain and a novel proline-rich motif containing glycine and arginine.

We have previously reported a nuclear protein possessing a WW domain, Npw38 (Komuro, A., Saeki, M., and Kato, S. (1999) Nucleic Acids Res. 27, 1957-1965). Here we report a Npw38-binding protein, NpwBP, isolated from HeLa cell nuclear extracts and its characterization using a cloned cDNA. NpwBP contains two proline-rich regions that are capable of binding to the WW domain of Npw38. The binding analysis using an oligopeptide-immobilized membrane revealed that the WW domain of Npw38 preferentially recognizes a short proline-rich sequence, PPGPPP, surrounded by an arginine residue, so we named it a PGR motif. Localization analysis using green fluorescent protein fusion protein and immunostaining showed that Npw38 and NpwBP are colocalized in the same subnuclear region. Coimmunoprecipitation experiments confirmed the association between Npw38 and NpwBP, which were expressed as epitope-tagged forms in COS7 cells. Furthermore, the N-terminal region of NpwBP has binding ability for poly(rG) and G-rich single-stranded DNA. These results suggest that NpwBP is a physiological ligand of Npw38 and that the Npw38-NpwBP complex may function as a component of an mRNA factory in the nucleus.

Amino Acid Sequence↗
PubMed1999

Ex vivo generation of CD34(+) cells from CD34(-) hematopoietic cells.

The human Lin(-)CD34(-) cell population contains a newly defined class of hematopoietic stem cells that reconstitute hematopoiesis in xenogeneic transplantation systems. We therefore developed a culture condition in which these cells were maintained and then acquired CD34 expression and the ability to produce colony-forming cells (CFC) and SCID-repopulating cells (SRCs). A murine bone marrow stromal cell line, HESS-5, supports the survival and proliferation of Lin(-)CD34(-) cells in the presence of fetal calf serum and human cytokines thrombopoietin, Flk-2/Flt-3 ligand, stem cell factor, granulocyte colony-stimulating factor, interleukin-3, and interleukin-6. Although Lin(-)CD34(-) cells do not initially form any hematopoietic colonies in methylcellulose, they do acquire the colony-forming ability during 7 days of culture, which coincides with their conversion to a CD34(+) phenotype. From 2.2% to 12.1% of the cells became positive for CD34 after culture. The long-term multilineage repopulating ability of these cultured cells was also confirmed by transplantation into irradiated NOD/SCID mice. These results represent the first in vitro demonstration of the precursor of CD34(+) cells in the human CD34(-) cell population. Furthermore, the in vitro system we reported here is expected to open the way to the precise characterization and ex vivo manipulation of Lin(-)CD34(-) hematopoietic stem cells.

Animals↗
PubMed1999

Effects of p51/p63 missense mutations on transcriptional activities of p53 downstream gene promoters.

The p51/p63 gene is a homologue of p53, the product of which acts as a transcriptional activator by binding to p53-responsive elements in the promoter regions of several p53 downstream genes. Recently, we identified four distinct mutations in the p51/p63 gene after screening >200 human tumors and cell lines. Because all of the detected p51/p63 mutations were missense mutations, the pathogenic effect of these mutations is difficult to determine without performing a functional analysis. In this study, we examined the transcriptional activity of tumor-derived p51/p63 missense mutations using a yeast-based assay and compared the data with that of artificial p51/p63 missense mutations at residues corresponding to the positions and substituted residues of p53 mutation "hotspots." Although most of the p51/p63 missense mutations at the p53 hotspot residues were unable to transactivate the promoters used in this study, the tumor-derived p51/p63 missense mutations retained their ability to transactivate the MDM2 and/or the BAX promoter but not the p21/WAF1 promoter. These results suggest that the p51/p63 mutation might be involved in an unknown tumor suppression pathway distinct from that of p53.

Amino Acid Sequence↗
PubMed1999

Covalent modification of all members of human cullin family proteins by NEDD8.

Recently we found that NEDD8, a ubiquitin-like protein, was linked covalently to human cullin-4A (abbreviated Cul-4A) by a new ubiquitin-related pathway that is analogous to but distinct from the ligating system for SUMO1, another ubiquitin-like protein. However, it remained unknown whether the other five members of the family of human cullin/Cdc53 proteins are modified by NEDD8. Here we report that all Hs-Cul family proteins, such as Cul-1, Cul-2, Cul-3, Cul-4B, and Cul-5, in addition to Cul-4A, were modified by covalent attachment of NEDD8 in rabbit reticulocyte lysates. Moreover, by comprehensive Northern-blot analyses, we examined multiple tissue distributions of the messages for all Cul-family proteins, NEDD8, and the NEDD8-ligating system consisting of APP-BP1/hUba3, and hUbc12, which function as E1- and E2-like enzymes, respectively. The expressions of Cul-1, Cul-2, and Cul-3 resembled each other and were apparently correlated to those of NEDD8 and the NEDD8-ligating system in various human cells and tissues. However, the mRNA levels of Cul-4A, Cul-4B, and Cul-5 differed considerably from each other as well as from other Cul-family proteins. The enhanced expression of all Cul-family proteins except Cul-5 was observed in a variety of tumor cell lines.

Cell Cycle Proteins↗
PubMed1999

Genetic mutation in the human 25-hydroxyvitamin D3 1alpha-hydroxylase gene causes vitamin D-dependent rickets type I.

Vitamin D is deeply involved in a wide variety of biological events such as calcium homeostasis, bone formation and cellular differentiation. An active form of vitamin D, 1alpha,25(OH)2D3, serves as a vitamin D receptor (VDR)-specific ligand to activate the expression of a particular set of target genes. 1Alpha,25(OH)2D3, is biosynthesized from cholesterol, and at the final biosynthesis step, 25-hydroxyvitamin D3 1alpha-hydroxylase [1alpha(OH)ase] in kidney conducts 1alpha-hydroxylation of 25(OH)2D3. This enzymatic activity is under multihormonal regulation and critical for the biosynthesis. Molecular cloning of 1alpha(OH)ase from several species has revealed that this enzyme belongs to a member of the cytochrome P450 enzyme superfamily, with highest homologies to the P450 hydroxylases for vitamin D derivatives. The renal gene expression is strictly regulated at the transcriptional level through its gene promoter by PTH and calcitonine (positive) and 1alpha,25(OH)2D3 (negative). Most importantly in clinical aspects, genetic mutations in this gene to abolish the enzymatic activity are now shown to cause the one of three kinds of hereditary rickets, vitamin D-dependent rickets type I.

Amino Acid Substitution↗
PubMed1999

Analysis of MEG signals of spreading cortical depression with propagation constrained to a rectangular cortical strip. II. Gyrencephalic swine model.

Currents produced during spreading cortical depression (SCD) in a gyrencephalic species (swine) were studied with magnetoencephalography (MEG) and electrocorticography (ECoG). SCD, initiated using electrical stimulation of the cortex, was constrained to propagate within a rectangular cortical strip in order to simplify the interpretation of the underlying currents. The ECoG signals monitored along the strip revealed that SCD propagated from an initiation site on the gyrus at a rate of 7.9+/-3.2 mm/min (n=23), entered the deep coronal sulcus and in most cases emerged from the other side of the sulcus, continuing to propagate across the next gyrus at a rate of 5.9+/-2.7 mm/min (n=22). The apparent propagation velocity within the sulcus was reduced to 1.7+/-0.8 mm/min (n=21). Strong MEG signals were observed as SCD entered the sulcus. The direction of magnetic field was opposite for SCD's on opposite banks of the sulcus. The currents were directed from a superficial layer to deeper layers of the cortex. The characteristics of SCD and associated MEG patterns from a gyrencephalic species may be similar to those in human patients during migraine aura.

Animals↗
PubMed1999

A heterozygous mutation of beta-actin associated with neutrophil dysfunction and recurrent infection.

A human disorder caused by mutation in nonmuscle actin has not been reported. We report here a variant of nonmuscle actin in a female patient with recurrent infections, photosensitivity, and mental retardation. She also had abnormalities in neutrophil chemotaxis, superoxide production, and membrane potential response. Two-dimensional PAGE analysis of proteins from neutrophils and other cell types from this patient demonstrated a unique protein spot migrating at 42 kDa with pI shifted slightly to neutral relative to normal beta- and gamma-actin. Digestion peptide mapping and Western blotting showed this spot to be an abnormal actin. A full-length cDNA library was constructed by using mRNA from patient's cells and cDNA encoding the mutant beta-actin molecule was identified by an in vitro translation method. Sequencing of the clones demonstrated a G-1174 to A substitution, predicting a glutamic acid-364 to lysine substitution in beta-actin and eliminating a HinfI DNase restriction site found in normal beta-actin sequence. By HinfI digestion and by sequencing, the mutation in one allele of patient's genomic DNA was confirmed. Though no defect in cell-free polymerization of actin was detected, this defect lies in a domain important for binding to profilin and other actin-regulatory molecules. In fact, the mutant actin bound to profilin less efficiently than normal actin did. Heterozygous expression of mutant beta-actin in neutrophils and other cells of this patient may act in a dominant-negative fashion to adversely affect cellular activities dependent on the function of nonmuscle actin.

Actins↗
PubMed1999

Mutational analysis of p51A/TAp63gamma, a p53 homolog, in non-small cell lung cancer and breast cancer.

p51, a novel family member of human p53, is a recently identified candidate tumor suppressor gene mapped at chromosome 3q28. Like p53, p51 was found to activate p21Waf1/Cip1 and to induce apoptosis. Since the DNA loss at 3q is reported in several cancers including non-small cell lung cancer (NSCLC), we screened for mutations in p51A (TAp63gamma), an isoform of p51 with short C-terminal region, in 80 NSCLCs as well as 85 breast cancers by RT-PCR single strand conformation polymorphism (SSCP) analysis and DNA sequencing. In NSCLCs, p51 was expressed in most tumors at variable levels and we found three missense and one silent mutations: Gln31His (transactivation domain) in two tumors, Ala148Pro (DNA-binding domain) and Leu248Leu (DNA-binding domain). In the tumor with Ala148Pro or the silent mutation, only the mutant gene appeared to be expressed. The modified FASAY method to test the ability of yeast expressing p51A cDNA to grow in medium lacking histidine has revealed that Ala148Pro results in a loss of function, while Gln31His does not. In contrast to NSCLC, no mutation was observed in all 85 breast cancers by the similar method. Our results suggest that, because of infrequent mutation, p51 may not be a Knudson type tumor suppressor in most NSCLCs and breast cancers. Nevertheless, in at least a part of NSCLC, p51 may play a certain role in carcinogenesis in a tissue-specific manner.

Adult↗
PubMed1999

Inhibitory effect of lipopolysaccharide on apoptotic cell death in macrophages infected with Actinobacillus actinomycetemcomitans.

We have reported that the macrophage-like cell line J774.1, when infected with the periodontopathic bacterium Actinobacillus actinomycetemcomitans, undergoes apoptosis. In this study, we examined whether stimulation of J774.1 cells with lipopolysaccharide (LPS) before the infection affects the subsequent apoptosis. Cytotoxicity on the LPS-stimulated cells was about half of the unstimulated control cells. DNA fragmentation in the LPS-stimulated cells was also significantly lower than in the control cells, whereas it was increased to a level similar to that of the control cells by addition of a nitric oxide (NO) inhibitor. In addition, significantly smaller numbers of live A. actinomycetemcomitans were recovered from the LPS-stimulated macrophages at 8 h after the infection as compared with the control cells. These findings suggest that the inhibitory effect of LPS on apoptosis results from an enhanced NO-mediated bactericidal activity.

Aggregatibacter actinomycetemcomitans↗
PubMed1999

The transcriptional activities of p53 and its homologue p51/p63: similarities and differences.

p51/p63 is a novel p53 homologue that has been shown to act as a transcriptional activator through the p53-binding sequence of the p21/WAF1 promoter and to induce apoptosis when it is expressed transiently in a human tumor cell line. We developed transcription assay systems for these two related genes in both Saccharomyces cerevisiae and mammalian cells and used them to investigate the functional similarities and differences of these genes. We found that p51/p63 trans-activated the previously identified p53 target genes, but the degree of the transactivation by p51/p63 differed from that by p53. These results suggest that the cellular signal on p51/p63 cross-talks partially but not completely with that of the p53 pathway.

Cyclin-Dependent Kinase Inhibitor p21↗
PubMed1999

Exploitation of the continuum between early ischemia/reperfusion injury and host alloresponsiveness: indefinite kidney allograft survival by treatment with a soluble P-selectin ligand and low-dose cyclosporine in combination.

BACKGROUND: We have shown previously that sPSGL, a soluble glycoprotein ligand for P and E selectins, reduces the events associated with ischemia/reperfusion injury of the kidney. In the present study, we have attempted to modulate differentially early inflammatory influences and later host alloresponsiveness in an LBNF1-Lewis renal graft model by treatment with sPSGL in combination with a marginally effective dose of cyclosporine (CsA). METHODS: Four experimental groups were studied: group 1=control animals receiving vehicle only; group 2=sPSGL monotherapy alone; group 3=low-dose CsA; group 4=sPSGL plus low-dose CsA. Grafts were removed at 1, 3, 5, and 7 days (n=3/time point) and assessed by histology, immunohistology, and reverse transcriptase-polymerase chain reaction. Long-surviving grafts in recipients of groups 3 and 4 were followed functionally for more than 28 weeks. RESULTS: Graft function was prolonged indefinitely in recipients in group 4, all of which survived for more than 200 days. In contrast, survival of animals in groups 1 and 2 was not increased substantially, whereas only 4 of 17 animals in group 3 (23.5%) survived more than 24 days (P<0.01). Five days after engraftment, necrosis was relatively minimal in group 4 organs but pronounced in those of the other groups. By immunohistology, numbers of infiltrating CD4+ and CD8+ T cells and ED1+ macrophages were significantly diminished in group 4 allografts compared with those of the other groups. Serial assessment of chemokine and cytokine mRNA expression confirmed these findings. The long-term effects of CsA treatment alone were compared with those of sPSGL in combination with CsA. Proteinuria remained virtually absent in group 4 recipients. Morphologically, the few long-surviving grafts in group 3 showed signs of chronic rejection; those in group 4 remained relatively normal. CONCLUSIONS: Although treatment with sPSGL alone showed no apparent influence on the acutely rejecting transplants, at least by the parameters examined in this study, it produced indefinite survival of kidney grafts when used in combination with low-dose CsA. The data support the influence of early nonspecific injury on later immunological rejection.

Animals↗
PubMed1999

Positive and negative modulation of vitamin D receptor function by transforming growth factor-beta signaling through smad proteins.

Several lines of experiments demonstrated the interplay between the transforming growth factor-beta (TGF-beta) and vitamin D signaling pathways. Recently, we found that Smad3, a downstream component of the TGF-beta signaling pathway, potentiates ligand-induced transactivation of vitamin D receptor (VDR) as a coactivator of VDR (Yanagisawa, J., Yanagi, Y., Masuhiro, Y., Suzawa, M., Watanabe, M., Kashiwagi, K., Toriyabe, T., Kawabata, M., Miyazono, K., and Kato, S. (1999) Science 283, 1317-1321). Here, we investigated the roles of inhibitory Smads, Smad6 and Smad7, which are negative regulators of the TGF-beta/bone morphogenetic protein signaling pathway, on the Smad3-mediated potentiation of VDR function. We found that Smad7, but not Smad6, abrogates the Smad3-mediated VDR potentiation. Interaction studies in vivo and in vitro showed that Smad7 inhibited the formation of the VDR-Smad3 complex, whereas Smad6 had no effect. Taken together, our results strongly suggest that the interplay between the TGF-beta and vitamin D signaling pathways is, at least in part, mediated by the two classes of Smad proteins, which modulate VDR transactivation function both positively and negatively.

Animals↗
PubMed1999

Npw38, a novel nuclear protein possessing a WW domain capable of activating basal transcription.

We have found a novel cDNA encoding a 265 amino acid protein possessing a WW domain in our full-length cDNA bank. The WW domain was sandwiched between an acidic region and an acidic-basic amino acid repetitive region. In vitro transcription/translation of the cDNA produced a 38 kDa product that was also found in the cell lysate by western blot analysis. Thus this protein is named the nuclear protein containing a WW domain with a molecular mass of 38 kDa, Npw38. Immunofluorescence studies and expression of a fusion protein to a green fluorescent protein revealed that this protein is localized in the nucleus. Npw38 was shown to be capable of binding to a poly(rG) resin. Interestingly, the WW domain of Npw38 was found to function as a transcriptional activator in CHO cells using the GAL4 DNA-binding fusion system. Furthermore, the WW domains of human YAP and Pin1 were demonstrated to have a similar transcription-promoting activity. Combined mutation of the conserved first and second Trp residues and a hydrophobic triplet of TyrTyrTrp in the WW domain of Npw38 abolished the transcription-promoting activity, but single mutations of these sites did not. These results suggest that some WW domains potentially possess transcription-promoting activity in mammalian cells.

Amino Acid Sequence↗
PubMed1999

Structural Change in Dextran: Mechanism of Insolubilization by Adsorption on the Air-Liquid Interface.

To clarify the insolubilization mechanism of water-soluble dextran, the association of dextran in water was studied by dynamic light scattering measurements and a surface chemical approach. Dynamic light scattering measurements indicated that insolubilization of dextran is accompanied by a structural change in dextran. Surface tension data for dextran molecules revealed a structural change in dextran molecules at the air-liquid interface. These results suggest that insolubilization of dextran molecules occurred through an adsorption process at the air-liquid interface. Insolubilization of dextran molecules can be reduced by inhibition of this structural change in dextran molecules. The presence of boron as an impurity was found to trigger precipitation based on inductively coupled plasma mass spectrometry measurements and precipitation tests. Copyright 1999 Academic Press.

Journal Article↗