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F Shimizu

Publications and source records attributed to F Shimizu.

At least 73 records · Page 4Linked to original sources

Cloning and characterization of two novel human cDNAs (NELL1 and NELL2) encoding proteins with six EGF-like repeats.

From a human fetal-brain cDNA library we isolated two novel genes encoding peptides containing six EGF-like repeats. Both showed significant homologies with nel, a gene strongly expressed in neural tissues of chicken. The cDNAs, designated NELL1 (nel-like, type 1) and NELL2 (nel-like, type 2), contained open reading frames encoding 810 and 816 amino acids, respectively. NELL2 is strongly expressed in brain of adult and fetus but only weakly in fetal kidney. NELL1 and NELL2 were mapped by FISH to chromosomal bands 11p15.1-p15.2 and 12q13.11-q13.12, respectively.

Adult↗

Calcium-bound recoverin targets rhodopsin kinase to membranes to inhibit rhodopsin phosphorylation.

In rod photoreceptor cells, Ca2+-bound recoverin associates with disk membranes and inhibits light-dependent phosphorylation of rhodopsin. However, the functional significance of Ca2+-induced membrane association of recoverin has not been fully evaluated. We found that Ca2+-bound recoverin forms a complex with rhodopsin kinase preferentially at the membrane surface. Addition of increasing amounts of membranes promoted the membrane association of recoverin, and remarkably suppressed rhodopsin kinase activity. It was concluded that the Ca2+-recoverin-rhodopsin kinase complex is stabilized by membrane association, leading to effective suppression of the kinase activity.

Animals↗

Transforming growth factor-beta 1 is the predominant paracrine inhibitor of macrophage cytokine synthesis produced by glomerular mesangial cells .

Cross-communication between glomerular cells and infiltrating mononuclear cells plays an important role in the generation of or recovery from glomerular diseases. We found that cultured mesangial cells secrete a factor that inhibits production of proinflammatory cytokines by activated macrophages. Treatment of J774.2 macrophages with conditioned media from rat mesangial cells blunted the transcriptional induction of IL-1 beta, IL-6, and TNF-alpha by LPS. None of the media conditioned by other fibroblastic, epithelial, or endothelial cell lines exhibited the inhibitory effect. Media conditioned by normal rat glomeruli contained a similar inhibitory activity, which was enhanced in an acute model of mesangial proliferative glomerulonephritis. To identify the active component involved, we examined the expression of known macrophage deactivators IL-10, IL-13, and TGF-beta 1 in mesangial cells. Under the basal culture conditions, strong expression of TGF-beta 1 mRNA was observed, whereas expression of neither IL-10 nor IL-13 was detected. Immunoblot analysis and a specific bioassay detected the active form of TGF-beta 1 exclusively in the mesangial cell conditioned media. The inhibitory activity was enhanced by heat treatment, consistent with the known property of TGF-beta. A specific anti-TGF-beta 1 neutralizing Ab abolished the inhibitory effect exerted by the mesangial cell media, and exogenously added TGF-beta1 suppressed macrophage cytokine expression in a dose-dependent manner. These findings demonstrate that mesangial cells and isolated glomeruli secrete a factor which suppresses cytokine expression by activated macrophages, the active entity being identified as TGF-beta 1.

Animals↗

Thy-1-mediated phosphatidylinositol turnover in cultured rat glomerular mesangial cell.

Thy-1 glycoprotein is expressed in rat glomerular mesangial cells, and anti-Thy-1 nephritis induced by anti-Thy-1 antibodies is a model of human renal diseases. In this study, we examined Thy-1-mediated biological reactions in cultured rat glomerular mesangial cells utilizing two anti-Thy-1 monoclonal antibodies (mAbs), 1-22-3 and OX-7. Incubation of the cells with these mAbs resulted in increased inositol trisphosphate (IP3) levels. The rise in IP3 produced by mAb 1-22-3 was greater than that produced by mAb OX-7 at the same dose. Incubation of mesangial cells with these mAbs resulted in an increase in the intracellular free calcium concentration ([Ca2+]i). mAb 1-22-3 induced a sustained increase in [Ca2+]i, while that induced by mAb OX-7 lasted 1-2 min, then decreased to the basal level. An transient increase in [Ca2+]i was also observed in Ca(2+)-free medium, indicating that these [Ca2+]i increases are due to release of Ca2+ from internal stores by IP3 without calcium flux across cell membrane. When cells were pretreated with protein tyrosine kinase (PTK) inhibitors (herbimycin A or genistein), Thy-1-mediated increases in [Ca2+]i were inhibited. These data suggest that Thy-1 induces the production of IP3 (including inositol 1,4,5-triphosphate, an intracellular Ca(2+)-releasing factor) and that PTKs may contribute to the Thy-1-mediated elevation of [Ca2+]i which presumably results from phospholipase C activation following Thy-1-mediated signaling in rat mesangial cells.

Animals↗

Polyreactive antigen-binding B cells in the peripheral circulation are IgD+ and B7-.

Polyreactive antibodies are naturally occurring antibodies, primarily of the IgM isotype, that are capable of reacting with a wide variety of different self and non-self antigens. Previously, we reported that a B cell capable of making polyreactive antibody has Ig receptors on its surface that can bind different antigens. The present investigation was initiated to characterize these polyreactive antigen-binding B cells further. A panel of fluorescein isothiocyanate-labeled antigens (insulin, IgG Fc fragment or beta-galactosidase) served as probes to select polyreactive antigen-binding B cells by cell sorting. Our experiment revealed that these polyreactive antigen-binding B cells were mainly of the IgD isotype. They expressed high levels of CD40 and major histocompatibility complex class II molecules, but little or no B7-1, B7-2, or Fas. In contrast to the binding of antigens to monoreactive receptors (usually high affinity), the binding of antigens to polyreactive receptors (usually moderate or low affinity) did not up-regulate the expression of B7-1 or B7-2. Antigens that bound to polyreactive receptors, however, were internalized and degraded, although not as efficiently as antigens that bound to monoreactive receptors. Despite the ability of these B7- cells to process antigens, they were not able to activate T cells in a mixed leukocyte reaction. It is concluded that polyreactive antigen-binding B cells have properties that are consistent with the ability to induce immunological tolerance.

Antibody-Producing Cells↗

Structural continuity of filtration slit (slit diaphragm) to plasma membrane of podocyte.

Murine monoclonal antibody 5-1-6 was reported to bind to the slit membrane and closely related structures in rat renal glomeruli; it induced heavy, reversible proteinuria and appeared to redistribute onto the plasma membrane of epithelial cells after binding at the original target sites. This phenomenon of antigenic movement has not been analyzed in detail to date. In addition to normal kidneys we also studied localization of the antigen recognized by monoclonal antibody 5-1-6 in protamine sulfate-perfused rat kidneys, in which slit diaphragms are known to be functionally modified. Isolated glomeruli as well as ultrathin kidney cryosections were labeled by the immunogold technique to clarify the relation between this antigen and the slit diaphragm. Sequential localization of injected monoclonal antibody was visualized using a post-embedding immunogold method in rats 2 hours to 12 days after injection of antibody. Ultrastructural immunogold labeling demonstrated that under normal conditions antigenic molecules were expressed mainly in the area beneath the slit diaphragms. Occasionally labeling was found at the base of the foot process, facing the glomerular basement membrane. After protamine sulfate treatment antigenic sites were dislocated due to the lifting and disruption of slit diaphragms, indicating that this antigen is associated with slit diaphragms. Injected antibody was localized at the filtration slits at 2 hours, and by 12 hours it had moved onto the apical plasma cell membrane of foot process. In addition, from 3 days onwards patch or cap-like formation on the plasma cell membrane of podocytes was seen. Possible shedding of antibody from podocyte cell surface membrane was occasionally encountered, but internalization of antibody was a minor event. Elution experiments in isolated glomeruli at day 3 indicated that antigen and antibody were both localized on the podocyte cell surface membrane, suggesting redistribution of immune complexes. In conclusion, filtration slits (slit diaphragms) and the apical membrane of foot process of podocytes demonstrate structural continuity, as revealed by the movement of the antigen recognized by monoclonal antibody 5-1-6 as antigen-antibody complexes.

Animals↗

Anti-DNA antibody derived from a systemic lupus erythematosus (SLE) patient forms histone-DNA-anti-DNA complexes that bind to rat glomeruli in vivo.

Histone can mediate the binding of both free DNA and DNA complexed to anti-DNA antibody to the glomerular capillary wall. We tested whether performed histone-DNA-anti-DNA immune complexes (IC) could bind to the glomerular capillary wall. The immune complex, generated with anti-DNA antibody derived from an SLE patient and excess of 125I-DNA followed by digestion with DNase, was mixed with histones. The complex containing 4 micrograms DNA was injected via the aorta into the left kidney of rats. At 15 min, 1-3% of the histone-DNA-anti-DNA antibody complex bound (measured as 125I-DNA), when histone was omitted less than 0-1% of the DNA-anti-DNA antibody complex bound. By immunofluorescence human immunoglobulins and histones, representing the IC, could be observed in a capillary pattern; but no complement deposition was detected. Electron microscopy revealed discrete, electron dense deposits in a subendothelial, subepithelial and mesangial localization at 15 min. These results provide direct evidence that antibodies from serum of SLE patients can form soluble histone-DNA-anti-DNA immune complexes that bind to the glomerular capillary wall in vivo.

Animals↗

Thy-1 molecule associates with protein tyrosine kinase(s) in rat mesangial cells.

Glycophosphatidylinositol (GPI)-linked Thy-1 molecules, well known cell surface markers of murine T cells, are present on the glomerular mesangial cells of the rat kidney. The administration of anti-Thy-1.1 MoAbs 1-22-3 and OX-7 to rats induces severe and mild complement-dependent mesangial proliferative glomerulonephritis, respectively. In order to determine whether protein-tyrosine kinase (PTK) activity is associated with Thy-1 molecules on rat mesangial cell surface, we performed an immune complex kinase assay, using anti-Thy-1 MoAbs 1-22-3 and OX-7, followed by reimmunoprecipitation with anti-phosphotyrosine, anti-fyn, anti-lck and anti-lyn antibodies. Physical association of PTK, p59fyn and p56/53lyn with Thy-1 molecules was demonstrated in cultured rat mesangial cells. The activities of these kinases detected in MoAb 1-22-3 precipitates were higher than those in MoAb OX-7 precipitates. These results suggest that Thy-1 molecule transduces some signals also in rat mesangial cells.

Animals↗

Cloning, expression, and mapping of UBE2I, a novel gene encoding a human homologue of yeast ubiquitin-conjugating enzymes which are critical for regulating the cell cycle.

From a human fetal-brain cDNA library we isolated a novel gene sharing significant homology with two yeast genes, UBC9 and hus5, which encode ubiquitin-conjugating enzyme 9 (UBC9). In yeast this protein is critical for normal mitosis, and seems to be closely involved in progression of G2 to M phase of the cell cycle. The human UBC9 (h-UBC9) cDNA, (gene symbol UBE2I), contained an open reading frame of 474 nucleotides encoding 158 amino acids. Its predicted peptide showed respectively 56% and 66% identity (75% and 82% similarity) with the products of UBC9 and hus5. Northern-blot analysis revealed expression of three transcripts, 6.4 kb, 3.3 kb, and 1.35 kb, in all human tissues examined. This gene, UBE2I, was mapped to chromosome band 16p13.3 by FISH.

Amino Acid Sequence↗

Cloning, expression, and mapping of CKAPI, which encodes a putative cytoskeleton-associated protein containing a CAP-GLY domain.

From a human fetal-brain cDNA library we isolated a novel cDNA clone encoding a protein containing a CAP-GLY domain that is highly conserved among several cytoskeleton-associated proteins. The CAP-GLY domain is thought to be essential for their association with microtubules. The cDNA, designated CKAPI (for cytoskeleton-associated protein I, glycine motif) contained an open reading frame of 579 nucleotides encoding 193 amino acids. Northern-blot analysis revealed expression of three transcripts, 1.0, 3.4, and 4.6 kb in size, in all tissues examined. The 1.0-kb transcript was significantly higher in brain and heart than in other tissues. This gene was mapped by FISH to chromosome bands 19q13.11-->q13.12.

Amino Acid Sequence↗

Cloning, expression and chromosomal mapping of a novel cyclophilin-related gene (PPIL1) from human fetal brain.

We isolated a human cDNA clone encoding a novel protein homologous to cyclophilins, specific cellular targets of cyclosporin A, which are conserved in species ranging from human to prokaryotes. This cDNA, designated hCyPX, contained an open reading frame of 498 nucleotides encoding 166 amino acids. Computer analysis indicated that its predicted amino acid sequence had 41.6%, 40.4%, and 39.2% homology to those of human, bovine, and Drosophila cyclophilins, respectively. Northern blot analysis indicated ubiquitous expression in adult human tissues, but most abundant expression in heart. Fluorescence in situ hybridization to human metaphase chromosomes localized this gene (PPIL1, peptidylprolyl isomerase [cyclophilin]-like 1) to chromosome bands 2p23.3-->p23.1.

Amino Acid Isomerases↗

Cloning, expression, and mapping of TCTEL1, a putative human homologue of murine Tcte1, to 6q.

From a human fetal-brain cDNA library we isolated a putative human homologue of the murine Tcte1 gene. The cDNA, designated TCTEL1, contained an open reading frame of 339 nucleotides encoding 113 amino acids. The predicted peptides of TCTEL1 showed 94% and 55% identity (100% and 94% similarity) with those of murine Tcte1 and human RP3. Northern-blot analysis revealed a 0.9-kb transcript in all tissues examined. This gene was mapped by FISH to chromosome bands 6q25.2 --> q25.3, the syntenic region of the murine t-complex locus of chromosome 17.

Animals↗

Cloning, expression pattern and mapping to 12p 13.2 --> p13.1 of CLAPS3, a gene encoding a novel clathrin-adaptor small chain.

From a human fetal-brain cDNA library we isolated a novel gene encoding a peptide homologous to clathrin-adaptor small chains in rat, mouse, and yeast. The cDNA, designated CLAPS3 (clathrin-associated/assembly/adaptor protein, small 3, 22 kDa), contained an open reading frame of 579 nucleotides encoding 193 amino acids. Northern-blot analysis revealed expression of a 1.35-kb transcript in all human tissues examined. This gene was mapped to chromosome bands 12p13.2 --> p13.1 by FISH.

Adaptor Protein Complex 3↗

Cloning, expression and chromosome mapping of adducin-like 70 (ADDL), a human cDNA highly homologous to human erythrocyte adducin.

From a human fetal-brain cDNA library we isolated a novel human cDNA, termed human adducin-like 70 (gene symbol ADDL), whose predicted amino acid sequence showed a high degree of homology to adducins. This cDNA clone (ADDL), which contained an open reading frame of 2,022 nucleotides encoding 674 amino acids, revealed 54%, 53%, and 59% identity in predicted amino acid sequence with alpha and beta components of human adducin and rat adducin 63, respectively. Human adducin-like 70 is likely to play an important role in the skeletal organization of the cell membrane. Northern blot analysis indicated ubiquitous expression of this gene in adult human tissues. We localized the gene to chromosome bands 10q24.2-->q24.3 by fluorescence in situ hybridization (FISH).

Adult↗

Isolation and mapping of the human glycoprotein M6 gene (GPM6A) to 4q33-->q34.

We have isolated a human cDNA that is highly homologous to the murine gene (Gpm6) encoding a membrane glycoprotein, M6. The human gene (GPM6A) contains an open reading frame of 834 nucleotides encoding a peptide of 278 amino acids. Northern-blot analysis revealed specific expression in human brain. We assigned the GPM6A locus to chromosome bands 4q33-->q34 by radiation hybrid mapping.

Amino Acid Sequence↗