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Biomedical subjects

O Niwa

Publications and source records attributed to O Niwa.

At least 91 records · Page 5Linked to original sources
PubMed1993

Heterogeneous expression of a novel MPC-1 antigen on myeloma cells: possible involvement of MPC-1 antigen in the adhesion of mature myeloma cells to bone marrow stromal cells.

Recent immunophenotypic analysis has shown that the heterogeneous expression of the adhesion molecule VLA-5 classifies myeloma cells into VLA-5+ mature and VLA-5- immature subpopulations. To further clarify the two myeloma subpopulations, we generated a monoclonal antibody, MPC-1, by immunizing mice with an adherent human myeloma cell line, KMS-5. The MPC-1 antibody recognized a 48-Kd surface antigen on KMS-5 but not on U-266, a nonadherent human myeloma cell line. Specificity characterization showed that MPC-1 antigen was expressed on mature myeloma cells, normal plasma cells, and mature B cells, whereas pre-B cells and germinal center B cells lacked its expression. Monocytes and a human bone marrow stromal cell line, KM102, also expressed this antigen. Two subclones of MPC-1+ VLA-5+ (KMS-5Ad) and MPC-1-VLA-5+ (KMS-5NAd) were separated from the KMS-5 cell line. The KMS-5NAd adhered to KM102 more tightly than did the KMS-5NAd, and the U-266 (MPC-1-VLA-5-) displayed almost no adherence to the KM102. The adhesion of the KMS-5Ad was partially inhibited by the MPC-1 antibody. These results, taken together, suggest that the MPC-1 antigen serves as a differentiation marker for B-lineage cells, including plasma cells, and may function as an adhesion molecule involved in the interaction of mature myeloma cells with bone marrow stromal cells.

Animals↗
PubMed1993

Transcriptional repressor ZF5 identifies a new conserved domain in zinc finger proteins.

We have cloned a cDNA encoding a new murine C2H2 zinc finger protein, ZF5. The 51.3 kD protein contains five GL1-Kruppel type zinc fingers at the C-terminus. At its N-terminus, ZF5 has a 41 amino acid region which was found to be homologous to the N-termini of several other zinc finger proteins. This region defines a new motif within zinc finger proteins which we have named the Zinc finger N-terminal (ZiN) domain. ZF5 binds to two sites in the c-myc promoter and to the -50 bp site of the herpes simplex thymidine kinase promoter. ZF5 is a transcriptional repressor and its repression domain is located N-terminal to the zinc finger domains. A single 4 kb ZF5 mRNA is expressed widely.

Amino Acid Sequence↗
PubMed1993

Small-volume voltammetric detection of 4-aminophenol with interdigitated array electrodes and its application to electrochemical enzyme immunoassay.

A small-volume voltammetric detection of 4-aminophenol (PAP) has been developed using an interdigitated array (IDA) microelectrode cell in order to apply the IDA to electrochemical enzyme immunoassay. The signal of PAP at the IDA was steady state, and its magnitude was amplified compared with that of the usual single electrode due to redox cycling of PAP between the two finger sets of the IDA. A linear relationship between PAP concentration and cathodic limiting current was obtained from 1 to 1000 microM, reproducibly. The minimum sample volume in the measurement was reduced to 800 nL. High sample throughput of less than 1-min detection time per sample was achieved on 2-10-microL PAP samples. This IDA cell was applied to the electrochemical enzyme immunoassay of mouse IgG. Alkaline phosphatase was used as the enzyme label. The mouse IgG concentration was evaluated by detecting the concentration of PAP, which is the product of enzymatic reaction of the substrate, 4-aminophenyl phosphate (PAPP). Anti-mouse IgG was covalently immobilized on the glass surface of the small-volume immunowells by carbodiimide coupling. The assay range was 10-1000 ng/mL using 10-microL sample and 20-microL substrate solutions.

Alkaline Phosphatase↗
PubMed1993

A 13 kb resolution cosmid map of the 14 Mb fission yeast genome by nonrandom sequence-tagged site mapping.

We present the application of a nonrandom sequence-tagged site (STS) content detection method in mapping an entire genome, that of fission yeast. The novelty of our strategy is in the use of STS probes made from both ends of cosmid clones, selected on the basis of "sample without replacement" (only library clones that show no previous positive hybridization are selected and made into probes). We developed powerful techniques, based on consistency analysis, for error detection and contig assembly. In addition, we probed our library with genetically mapped markers and Notl or Sfil linking clones, thereby anchoring contigs onto chromosomes. Our map contains more than 1000 sites, including genes (most were previously unmapped), occurrences of known repetitive elements, and Notl-Sfil restriction sites.

Chromosome Mapping↗
PubMed1993

Two mouse hypervariable minisatellites: chromosomal location and simultaneous mutation.

Pc-1 and Pc-2 hypervariable minisatellites of the mouse show a structural similarity and hence have been compared in terms of chromosomal location and mutability. Polymorphisms of the Pc-2 locus detected by analyses of Southern blotting and polymerase chain reaction were used for typing 61 backcross mice that were obtained by mating (C57BL/6 x MSM)F1 females to MSM males. Segregation patterns indicated that the Pc-2 locus maps to the 60 cM region of chromosome 6, while that of Pc-1 maps to chromosome 4. Simultaneous mutation of the two loci was examined in 101 F1 mice. Pc-1 and Pc-2 loci showed mutation rates of 14.4 and 5.0% per gamete, respectively. Of the nine mice with a Pc-2 mutation, four carried a Pc-1 mutation at the same maternally- or paternally-derived allele. The rate of comutation was 44% (4/9), higher than the 12.2% (24/197) of Pc-1 mutation alone. This suggests that the germline mutation of the two hypervariable minisatellites may result from a common mechanism.

Alleles↗
PubMed1993

Random process of metastasis and generation of heterogeneity in a mouse sarcoma line.

The process of metastasis was analyzed in 505-05-01 cells, an established line of a methyl-cholanthrene-induced mouse sarcoma, by tagging the cells with the pSV2neo plasmid. A dominant clone was identified which, upon transplantation together with other clones, overgrew tumors in the kidney capsule. However, when this clone was transplanted in a mouse collaterally with recessive clones, both clones grew at the same rate and metastasized to the lung at an equal frequency. This suggests that the process of metastasis in this particular sarcoma line is stochastic, the dominant population having a better chance to colonize to the lung. The dominant neomycin-resistant clone was transfected with another marker plasmid, pY3, which confers resistance to hygromycin. Results of mixed inoculation of 9 independently isolated clones revealed the hierarchy of dominance among clones. This indicated the existence of heterogeneity within the parental clone. Upon mixed inoculation with hygromycin-resistant clones, the parental clone overgrew in the tumors. This indicated that some clone had changed its phenotype to become less aggressive. Thus, the direction of phenotypic drift in vitro seems to be random in terms of behavior in vivo.

Animals↗
PubMed1993

Comparison of the age- and tumor-associated changes in the c-myc gene methylation in mouse liver.

Study of DNA methylation of the c-myc gene in liver tumors of BCF1 mice treated with various agents revealed frequent alteration from that in normal liver. The alteration, however, was complex and showed either an increase or a decrease of methylation to various degrees. On the other hand, the tumors obtained from middle-aged C3H/He mice, which develop liver tumors spontaneously at a high incidence rate starting in middle age, showed predominantly increases of methylation in the c-myc gene, while the large tumors found in older mice revealed decreases. The normal part of the liver showed a slight gradual age-dependent increase. These suggest that hypermethylation of the c-myc gene is common to aging and early tumor development in liver. Thus the alteration of DNA methylation seems to be a good clue to investigate why the tumor incidence rate increases rapidly as individuals grow older.

Aging↗
PubMed1992

Isolation of high affinity cellular targets of the embryonal LTR binding protein, an undifferentiated embryonal carcinoma cell-specific repressor of Moloney leukemia virus.

ELP, the embryonal LTR binding protein, is a member of the nuclear receptor superfamily and a mouse homologue of Drosophila FTZ-F1. ELP is expressed specifically in undifferentiated mouse embryonal carcinoma cells and participates in suppression of the Moloney murine leukemia virus genome. The zinc finger domain of the protein was fused with glutathione S-transferase and was successfully used for isolating genomic targets. Sixteen genomic fragments were isolated and twelve of them strongly interacted with ELP. Six of the ELP binding fragments were analyzed further. All of these contained the multiple binding sites for ELP, which matched well with the consensus binding sequence for FTZ-F1, YCAAGGYCR. Among these, three fragments functioned as negative regulatory elements in response to ELP, when placed upstream to the promoter region of the Moloney leukemia virus. These results indicate that ELP may function as a negative transcription factor for a variety of cellular sequences, in addition to suppressing expression of Moloney leukemia virus in early embryonal cells. It was also shown that the procedure employed here works well for isolation of genomic targets of transcription factors.

Animals↗
PubMed1992

Rat cellular mutants for expression of mRNA from the long terminal repeat of murine retrovirus.

Previously we isolated revertants from a rat cell line transformed by recombinant murine retrovirus containing the v-src gene. These mutant cell lines, R78 and R107, showed low src-kinase activity, but retained wild-type transforming retrovirus, suggesting that a cellular gene involved in viral gene expression was mutated. Southern and Northern hybridization analyses showed that the expression of viral mRNAs from the integrated proviral DNA was reduced in these mutant cells. DNA transfection experiments with various transforming genes and promoters revealed that the mutant cell lines were resistant to transformation by transforming genes expressed under the long terminal repeat (LTR) of Moloney murine leukemia virus (Mo-MuLV). In contrast, these cell lines could be efficiently transformed by the same transforming genes with human metallothionein promoter, polyomavirus promoter-enhancer, and c-H-ras promoter. Transient expression assays using plasmids containing the CAT gene under the LTR of Mo-MuLV also showed that CAT activity expressed under the LTR in these mutant cells was lower than that in the parental cell line, No. 7. These results suggest that cellular mutations of R78 and R107 cells affect specific transcription from the LTR of Mo-MuLV. Studies using various constructs of the LTR CAT indicated that the region responsible for the repression was located in a fragment (-328 to -150) of the LTR containing the 72-bp repeat enhancer. Somatic cell hybridization experiments showed that the mutant phenotype of these mutant cell lines is dominant to that of the parental cell line.

Animals↗
PubMed1992

A low copy number central sequence with strict symmetry and unusual chromatin structure in fission yeast centromere.

Fission yeast centromeres vary in size but are organized in a similar fashion. Each consists of two distinct domains, namely, the approximately 15-kilobase (kb) central region (cnt+imr), containing chromosome-specific low copy number sequences, and 20- to 100-kb outer surrounding sequences (otr) with highly repetitive motifs common to all centromeres. The central region consists of an inner asymmetric sequence flanked by inverted repeats that exhibit strict identity with each other. Nucleotide changes in the left repeat are always accompanied with the same changes in the right. The chromatin structure of the central region is unusual. A nucleosomal nuclease digestion pattern formed on unstable plasmids but not on stable chromosome. DNase I hypersensitive sites correlate with the location of tRNA genes in the central region. Autonomously replicating sequences are also present in the central region. The behavior of truncated minichromosomes suggested that the central region is essential, but not sufficient, to confer transmission stability. A portion of the outer repetitive region is also required. A larger outer region is necessary to ensure correct meiotic behavior. Fluorescence in situ hybridization identified individual cens. In the interphase, they cluster near the nuclear periphery. The central sequence (cnt+imr) may play a role in positioning individual chromosomes within the nucleus, whereas the outer regions (otr) may interact with each other to form the higher-order complex structure.

Base Sequence↗
PubMed1992

Somatic mutation during metastasis of a mouse fibrosarcoma line detected by DNA fingerprint analysis.

Metastatic nodules were examined by DNA fingerprint analysis. The probes used, Pc-1 and Pc-2, detect mutations as shifts in bands of the minisatellite loci which are dispersed among chromosomes. Four clonal lines of a fibrosarcoma from an F1 mouse (C57BL/Ka x C3H/He) were selected for various metastatic potentials upon inoculation into syngeneic mice. These four lines exhibited many extra bands resulting from recombination and/or DNA slippage, indicating accumulation of mutations during the successive passages in mice. One of the four, a 505 cell line which had been passaged extensively in vitro and consisted of a heterogenous population, was inoculated into thirteen syngeneic mice, and gave rise to six lung metastatic nodules in two mice. All the nodules showed band-patterns distinct from one another, although nodules within a given mouse tended to show similar patterns. When a genetically tagged 505-05-01 clone was analyzed, three of nine metastatic nodules obtained also revealed new bands. These results strongly suggest that somatic mutations occur at a high frequency during metastasis, providing direct evidence of genetic instability of the tumor cells.

Animals↗
PubMed1992

Lack of allelic preference in amplification and loss of the c-myc oncogene in methylcholanthrene-induced mouse sarcomas.

Sarcomas were induced in F1 mice between C57BL/6N and C3H/He strains by subcutaneous injection of methylcholanthrene. The c-myc oncogene was found to be amplified in 16 cases among 43 sarcomas of C57BL/6N x C3H/He mice and 1 case among 5 sarcomas of the reciprocal cross. The origin of the amplified allele was determined by the polymerase chain reaction single strand conformation polymorphism analysis. Among the 17 sarcomas, only one had both of the alleles amplified. The rest of the tumors carried the amplified c-myc allele coming either from C57BL/6N (9 cases) or from C3H/He (8 cases). These results indicate that the c-myc allele is amplified randomly in methylcholanthrene-induced mouse sarcomas irrespective of its origin, such as paternal or maternal allele and C57BL/6N or C3H/He allele. In addition to these changes, the unamplified c-myc oncogene was found to be lost in 12 cases out of the 17 sarcomas with the amplification.

Alleles↗
PubMed1992

Embryonal long terminal repeat-binding protein is a murine homolog of FTZ-F1, a member of the steroid receptor superfamily.

The embryonal long terminal repeat-binding protein, ELP, is present in undifferentiated mouse embryonal carcinoma cells. It binds to and suppresses transcription of the Moloney leukemia virus long terminal repeat in undifferentiated murine embryonal carcinoma cells. We report here that ELP is a mouse homolog of Drosophila FTZ-F1, which positively regulates transcription of the fushi tarazu gene in blastoderm-stage embryos of the fly. As members of the steroid receptor superfamily, ELP and FTZ-F1 have both DNA binding and putative ligand binding domains which are well conserved between the two. ELP and FTZ-F1 function in cells in the extremely early stage of development. A high degree of conservation between the two transcription factors during the evolution of these species indicates the importance of their functions in early-stage embryogenesis. In addition, the sequence elements they recognize do not contain repeat units, in contrast to other steroid receptors, which usually bind to either palindromic or direct repeat sequences.

Amino Acid Sequence↗
PubMed1991

A large number of tRNA genes are symmetrically located in fission yeast centromeres.

We report here that the fission yeast centromere regions in the three chromosomes contain no less than 36 symmetrically arranged tRNA-coding sequences, and many of them are located within the inner inverted regions that are thought to be essential for the centromere function. There are 11 different species of tRNA-coding sequences, and four of them are identical to those previously known in this organism. This high-density distribution of tRNA genes in the centromere regions is surprising, as the fission yeast centromeres were thought to form transcriptionally inactive structures.

Base Sequence↗
PubMed1991

Structure of the fission yeast centromere cen3: direct analysis of the reiterated inverted region.

We determined the structure of the Schizosaccharomyces pombe centromere cen3 using direct genomic mapping and cosmid walking. The repetitive region of cen3 is approximately 110 kb, much longer than that of the previously determined cen1 and cen2 regions. The approximately 30 kb long left and approximately 60 kb right repetitive sequences are arranged with an inverted symmetry and flank the 15 approximately 20 kb central domain. The repeat motifs in cen3, although they consist of the common centromeric repeat elements, are slightly different from those in cen1 and cen2. The cen3 repeat motifs appear to be reiterated four times in the left and nine times in the right side repetitive regions. We found that the central domain consists of the common approximately 5 kb core sequence associated with the pair of innermost inverted sequences, most of which are reiterated only twice in the genome. Although their sizes differ significantly, the general features of cen1, cen2 and cen3 are similar, and a prototype, consensus structure for the fission yeast centromere may be deduced.

Base Sequence↗
PubMed1991

Sexual difference and organ specificity of the effect of estradiol on carbonic anhydrase and Mg(2+)-HCO3(-)-ATPase activities isolated from duodenal mucosa and kidney cortex of male and female rats: preliminary study with crude enzyme samples.

Effects of the s.c. administration of various doses of estradiol propionate (E.P.; 25-500 micrograms/kg) on the activities of carbonic anhydrase (CA), Mg(2+)-dependent ATPase and Mg(2+)-dependent, HCO3(-)-stimulated ATPase (Mg(2+)-HCO3(-)-ATPase) in rat duodenal mucosa and kidney cortex, and on body weight, organ weight and serum concentrations of testosterone and estradiol-17 beta, were examined in adult male, female, testectomized and ovariectomized rats. In normal male rats, activities of cytosol CA and brush border Mg(2+)-HCO3(-)-ATPase in the kidney were increased in a dose-dependent manner and reached 1.6- and 2-fold of controls, respectively, after consecutive administration (daily for 7 days) of 500 micrograms E.P. with no changes in either enzyme activities in duodenal mucosa. The positive correlations (P less than 0.01) were observed by linear regression analysis between serum concentration of estradiol-17 beta and kidney cytosol CA or kidney brush border Mg(2+)-HCO3(-)-ATPase activities. In normal female rats, activities of cytosol CA and brush border Mg(2+)-HCO3(-)-ATPase in the duodenal mucosa, and brush border Mg(2+)-HCO3(-)-ATPase activity in the kidney were increased by E.P. administration (100 and 500 micrograms/kg, daily for 7 days), however, kidney cytosol CA activity did not change by any dosage. Behavior of a part of both enzymes to E.P. in testectomized rats was altered almost in the same way to that observed in normal female rats and vice versa in ovariectomized rats. Body weight was decreased, in general, by consecutive administration of E.P. in a dose-dependent manner, and kidney weight was increased by E.P. in both male and female rats.

Animals↗
PubMed1991

Induction of transplantable tumors by repeated subcutaneous injections of natural and synthetic vitamin E in mice and rats.

Natural vitamin E and synthetic vitamin E (dl-alpha-tocopheryl acetate) were tested for their tumorigenicity in rodents. Transplantable tumors, at the site of injection, were induced by repeated injections of these compounds in two strains of mice, NFS/N and C57BL/6N x C3H/He F1, and in a strain of rats, Fischer 344. Natural vitamin E was tumorigenic in both strains of female mice only when injected with soya oil. In contrast, dl-alpha-tocopheryl acetate alone was capable of inducing tumors in Fischer 344 rats. Only one out of 5 male NFS/N mice given dl-alpha-tocopheryl acetate developed a tumor. Therefore, Fischer 344 rats were more susceptible to tumor formation by dl-alpha-tocopheryl acetate than NFS/N mice. dl-alpha-Tocopheryl acetate with soya oil or with palm oil also resulted in the formation of transplantable tumors in NFS/N mice and Fischer 344 rats. There was no difference in the tumor incidence between mice treated with dl-alpha-tocopheryl acetate alone and dl-alpha-tocopheryl acetate plus soya oil or palm oil. However, in rats, the incidence was lower for a group treated with dl-alpha-tocopheryl acetate plus palm oil than for those with dl-alpha-tocopheryl acetate alone and with dl-alpha-tocopheryl acetate plus soya oil.

Animals↗