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Biomedical subjects

T Horiuchi

Publications and source records attributed to T Horiuchi.

At least 217 records · Page 12Linked to original sources
PubMed1996

Structure of a replication-terminator protein complexed with DNA.

The crystal structure of the Escherichia coli replication-terminator protein (Tus) bound to terminus-site (Ter) DNA has been determined at 2.7 A resolution. The Tus protein folds into a previously undescribed architecture divided into two domains by a central basic cleft. This cleft accommodates locally deformed B-form Ter DNA and makes extensive contacts with the major groove, mainly through two interdomain beta-strands. The unusual structural features of this complex may explain how the replication fork is halted in only one direction.

Amino Acid Sequence↗
PubMed1996

Cloning of a gene from Escherichia coli that confers resistance to fosmidomycin as a consequence of amplification.

A gene conferring resistance to fosmidomycin (Fs) was cloned from the gene pool of a wild-type strain of Escherichia coli. The cloned DNA fragment was sequenced and shown to encode a putative polypeptide of 406 amino acids (aa) with a molecular weight of 43303. The gene mapped at 10.9 min on the E. coli chromosome and was designated fsr (fosmidomycin resistance). Maxicell analysis revealed that the Fsr protein migrated in sodium dodecyl sulfate-polyacrylamide-gel electrophoresis as a broad band of 35 kDa. A comparison between the aa sequence of Fsr and sequences in a protein database revealed 18% homology to the bacterial drug-export proteins that mediate resistance to tetracycline and chloramphenicol. Hydropathy analysis of the Fsr protein revealed twelve putative transmembrane segments. The degree of FsR of transformants depended on the number of copies of the plasmid that contained fsr. The levels of ubiquinone-8 and undecaprenyl phosphate in cells that harbored a high-copy-number plasmid that included fsr were almost the same as those in the cells without the plasmid. These results suggest that Fsr does not have any direct effect on the biosynthesis of isoprenoid in E. coli, and that the mechanism for FsR involves the efflux of the drug by a process that is facilitated by Fsr.

Amino Acid Sequence↗
PubMed1996

LAPTM5: a novel lysosomal-associated multispanning membrane protein preferentially expressed in hematopoietic cells.

While a large body of knowledge about cell membrane proteins exists, much less is known about the repertoire and function of integral membrane proteins of intracellular organelles. In looking for novel classes of genes that are functionally important to hematopoietic cells, we have cloned the cDNA for a gene preferentially expressed in adult hematopoietic tissues. During embryonic development the gene is expressed in both hematopoietic and nonhematopoietic tissues. In cell lines the gene is expressed specifically in hematopoietic lineages, whereas in normal adult tissues the mRNA is preferentially detected at high levels in lymphoid and myeloid tissues. The predicted protein is a pentaspanner with no homology to known genes and conserved across evolution. Immunocytological and cell fractionation studies with a specific antibody revealed a protein localizing in lysosomes. The gene, provisionally named LAPTM5, maps to chromosome 1p34. The expression pattern of the gene together with preliminary evidence that the protein interacts with ubiquitin indicates that the protein may have a special functional role during embryogenesis and in adult hematopoietic cells.

Adult↗
PubMed1996

A 718-kb DNA sequence of the Escherichia coli K-12 genome corresponding to the 12.7-28.0 min region on the linkage map.

The 718,122 base pair sequence of the Escherichia coli K-12 genome corresponding to the region from 12.7 to 28.0 minutes on the genetic map is described. This region contains at least 681 potential open reading frames, of which 277 (41%) have been previously identified, 147 (22%) are homologous to other known genes, 139 (20%) are identical or similar to the hypothetical genes registered in databases, and the remaining 118 (17%) do not show a significant similarity to any other gene. In this region, we assigned a cluster of cit genes encoding multienzyme citrate lyase, two clusters of fimbrial genes and a set of lysogenic phage genes encoding integrase, excisionase and repressor in the e14 genetic element. In addition, a new valine tRNA gene, designated valZ, and a family of long directly repeated sequences, LDR-A, -B and -C, were found.

DNA, Bacterial↗
PubMed1996

Extent of tumor vascularization correlates with prognosis and hematogenous metastasis in gastric carcinomas.

To determine whether tumor angiogenesis correlates with prognosis and metastasis of patients with gastric carcinoma, we counted the microvessels within the primary carcinoma and compared their numbers with the patient's prognosis and mode of metastasis. Tumor specimens from 110 patients with gastric carcinoma, who had undergone curative resection more than 24 months before, were investigated. Intratumoral microvessels were stained with anti-CD34 and anti-von Willebrand factor monoclonal antibodies before being quantitated by light microscopy (x200). The antibody against von Willebrand factor often showed variability and stromal background staining, providing misleading low vessel counts. The data from three patients who died from nongastric carcinoma within 24 months after surgery were deleted. A total of 107 patients took part in the analysis examining the association between intratumoral microvessels and clinical outcomes. Vessel counts derived from CD34 expression were significantly higher in patients who experienced hematogenous or peritoneal metastasis after surgery than in patients with nonmetastatic tumors. No correlation between vessel counts and lymph node metastasis was found. The prevalence of hematogenous metastasis. but not peritoneal metastasis, increased as the vessel counts increased. Multivariate logistic regression and Cox hazards model analyses showed that the vessel counts obtained with CD34 staining correlated with the development of hematogenous recurrence but not peritoneal recurrence. It was the most important factor for predicting overall survival. These findings support the hypothesis that tumor angiogenesis is closely related to the development of hematogenous metastasis in human gastric carcinomas. Assessment of tumor vascularization may, therefore, prove valuable in identifying patients with gastric carcinoma at high risk for recurrence who would benefit from adjuvant therapy.

Adenocarcinoma↗
PubMed1996

Molecular bases for inherited human complement component C6 deficiency in two unrelated individuals.

Deficiency of the sixth component of complement (C6D) is frequently associated with recurrent neisserial infections, especially meningitis caused by Neisseria meningitidis. We here report the molecular bases of C6D in two unrelated subjects, one African American (case 1) and the other Japanese (case 2). Screening all 17 exons of the C6 gene and their boundaries by exon-specific PCR/single strand conformation polymorphism demonstrated aberrant single stranded DNA fragments in exon 12 of case 1 and exon 2 of case 2. Nucleotide sequencing of the amplified DNA fragments revealed a homozygous single-base deletion (G1936) in exon 12 case 1 and a heterozygous single base deletion (C291/C292/C293/C294) in exon 2 of case 2. Both mutations resulted in frame shifts and premature termination of the C6 polypeptide. Sequence-specific oligonucleotide probe hybridization and direct sequencing of exon 12 amplified from genomic DNA further supported the homozygosity of the mutation in case 1. Case 2 is apparently compound heterozygote, but the putative mutation in the other allele of the C6 gene remains unknown. Both case 1 and case 2 were homozygous for the C6A allotype. These data indicate that at least three distinct mutational events can cause C6D, single nucleotide deletions in exons 2 and 12, and a mutation yet unidentified. Thus, similar to other complement protein deficiencies, the pathogenesis of C6D appears to be heterogeneous.

Adult↗
PubMed1996

Partner switching mechanisms in inactivation and rejuvenation of Escherichia coli DNA gyrase by F plasmid proteins LetD (CcdB) and LetA (CcdA).

Escherichia coli DNA gyrase, as well as a free form of its A subunit (GyrA), exists in an inactivated form in cells that overproduce the F plasmid protein LetD (CcdB). We found that the inactivated DNA gyrase and GyrA protein can be rejuvenated in vitro by another F plasmid protein, LetA (CcdA). Using this rejuvenation as an assay, we purified the inactivated GyrA protein to near homogeneity and found it to be complexed with the LetD protein. The complex has a molecular mass of 230 kDa and was suggested to be a complex of two molecules each of GyrA and LetD proteins. The GyrA-LetD complex, in the presence of purified GyrB protein, does not cause DNA cleavage. Therefore, the LetD protein in the GyrA-LetD complex inhibits the gyrase action by a mechanism different from one that involves trapping a covalently linked gyrase-DNA complex. In as much as a free form of the LetD protein has been shown to induce DNA cleavage by gyrase, the LetD protein seems to have two distinct modes of action on DNA gyrase. Rejuvenation of the inactivated GyrA protein by the LetA protein was achieved in vitro, and mechanisms governing this process were examined using the purified proteins. The rejuvenated GyrA protein sediments through sucrose gradients as a single protein species of 190 kDa and is indistinguishable from a free form of GyrA protein. In the same sedimentation experiment, the LetD protein was seen to be complexed with the added LetA protein. Thus, the LetA protein apparently rejuvenates the GyrA protein by removing the bound LetD protein from the inactivated form, followed by formation of a LetA-LetD complex.

Bacterial Proteins↗
PubMed1996

Evidence for involvement of Escherichia coli genes pmbA, csrA and a previously unrecognized gene tldD, in the control of DNA gyrase by letD (ccdB) of sex factor F.

The letA (ccdA) and letD (ccdB) genes of the F plasmid, located just outside the sequence essential for replication, contribute to stable maintenance of the plasmid in Escherichia coli cells. The letD gene product acts to inhibit partitioning of chromosomal DNA and cell division by inhibiting DNA gyrase activity, whereas the letA gene product acts to reverse the inhibitory activity of the letD gene product. To identify the host factor(s) involved in this process, we analyzed the mutants that escaped letD expression and their suppressor, and found that the three E. coli genes tldD, tldE and zfiA participate in the process, in addition to the groE genes we reported previously. The tldD and tldE mutations made cells tolerant for letD expression, as did groES mutations, while the mutation in the zfiA gene made tldD, tldE and groES mutants LetD sensitive. We hypothesize that these gene products are factors that modulate activity of DNA gyrase along with the letD gene product; the zfiA gene product acts to inhibit interaction between the LetD protein and the A subunit of DNA gyrase, while the tldD, tldE and groE gene products act to suppress the inhibitory activity of the zfiA gene product. The tldD, tldE, and zfiA genes are located at 70.4, 96.0 and 58.2 minutes on the E. coli chromosome, respectively, and code for proteins with relative molecular masses of 51,000, 48,000 and 6800, respectively. tldD is a novel gene, but the tldE and zfiA genes proved to be the pmbA gene (production of Microcin B17) and the csrA gene (carbon storage regulator), respectively.

Amino Acid Sequence↗
PubMed1996

Biochemical bases in differentiation of a mouse cell line GSM06 to gastric surface cells.

A mouse gastric surface cell line GSM06 established from a transgenic mouse harboring temperature-sensitive simian virus 40 large T-antigen gene was subjected to the lipid and glycoprotein analysis. When GSM06 cells were cultured for a long time after formation of a confluent monolayer, they differentiated to resemble foveolar epithelial cells morphologically. Biochemical changes during culture were studied in cells harvested just when a monolayer had formed (day 0), on day 7, and on day 21. Content of total phospholipids, cholesterol, cholesterol sulfate, total sugar and sialic acid increased about 1.5-fold from day 0 to 7 and remained elevated till day 21. The fatty acid composition of phospholipids revealed increased relative levels of oleic acid in phosphatidylcholine and phosphatidylethanolamine, and an increased level of plasmenylethanolamine from day 0 to 7. The level of dolichylphosphate continued to increase in a time-dependent manner. Glycosylation of various proteins, detected with lectins, was enhanced from day 7. In addition, greater resistance to taurodeoxycholate and acetylsalicylic acid was observed on days 7 and 21 than on day 0. Thus, enhanced glycosylation of proteins and an overall increase in the area of cellular membranes were the major changes in GSM06 cells during culture, and they were accompanied by an enhancement of cytoprotective potential.

Animals↗
PubMed1996

Crystallization and preliminary X-ray analysis of the Escherichia coli replication terminator protein complexed with DNA.

Crystals of the Escherichia coli replication terminator protein (Tus) complexed with its binding site DNA were obtained by a microdialysis method using PEG 4000. They belong to the tetragonal space group P4(1)2(1)2 or P4(3)2(1)2 with the unit cell parameter: a = 68.1 A, c = 230.7 A and contain one protein-DNA complex in an asymmetric unit. The native data set has been collected to 2.7 A resolution.

Bacterial Proteins↗
PubMed1996

Deletion in the 190 kDa antigen gene repeat region of Rickettsia rickettsii.

The 190 kDa outer membrane protein of Rickettsia rickettsii is a major immunodominant protective antigen which contains 13 tandem nearly identical repeating sequences. We have identified a deletion in the 190 kDa antigen gene of R. rickettsii strain Smith maintained in the laboratory. The deletion occurred within the repeat region of this gene and the mutated repeat region corresponded to one repeating unit in size.

Bacterial Outer Membrane Proteins↗
PubMed1996

Extradural extension of glioblastoma multiforme into the oral cavity: case report.

A rare case of glioblastoma multiforme with oral extension is presented in a 41-year-old female. She underwent two surgical treatments and both radiotherapy and chemotherapy for the right temporoparietal glioblastoma multiforme. A follow-up computed tomographic scan and magnetic resonance imaging demonstrated destruction of the temporal base and extradural extension of the tumor into the orbital, nasal, and oral cavities. This is the first report of the oral extension of glioblastoma multiforme. The mechanism for the extradural extension is discussed.

Adult↗
PubMed1996

Comparative study of bone mineral density estimated by various methods of single- and dual-energy quantitative computed tomography: the capability of the four-equation four-unknown method.

A dual-energy (DE) quantitative computed tomography (QCT) method, the four-equation four-unknown method (DEQCT 4E-4U), was assessed and compared to single-energy (SE) QCT and standard DEQCT (two-line method). The results of this study indicate that bone mineral density (BMD) was more accurately estimated by the present method than by the SEQCT or standard DEQCT techniques on the basis of a phantom study when a large fat content was present. The results of both the phantom study and a human study also showed that the present method corrected for fat in estimating BMD in the presence of high-fat content. These findings suggest that use of this method for estimating BMD can provide useful information in studies assessing the metabolic state of bone. We propose that CT numbers estimated from excised vertebral bone marrow can serve as a soft-tissue correction for the present method.

Adult↗
PubMed1996

A yeast gene product, Fob1 protein, required for both replication fork blocking and recombinational hotspot activities.

BACKGROUND: In the rRNA gene cluster on the Saccharomyces cerevisiae chromosome XII, a unique site that blocks progression of the replication fork exists in a single unit of the repeats. The site RFB (replication fork blocking) is located within one of two cis-elements present in a nontranscribed region of each repeated unit, which are required for obtaining maximal activity of a hotspot (HOT1) dependent recombination. RESULTS: To investigate the correlation between replication fork blocking at RFB and homologous recombination at HOT1, we have isolated Hot1-defective mutants and examined their ability for fork blocking at RFB. Amongst 23 isolated mutants, four were found to be defective in both abilities. Genetic analysis of the mutants reveals that a single mutation, named fob1 (fork blocking less), is responsible for the defects in both abilities. The FOB1 gene is located on chromosome IV and has no homology with any other genes listed in DNA data banks. CONCLUSION: The pleiotropic effect of the fob1 mutation suggests that homologous recombination at HOT1 is closely linked with DNA replication fork blocking event at RFB.

Amino Acid Sequence↗
PubMed1996

Production of the third and fourth component of complement (C3, C4) by smooth muscle cells.

Production of the third and fourth components of complement (C3, C4) by smooth muscle cells was investigated by using normal human aortic smooth muscle cells (AoSMC), human smooth muscle cell line (G402) and vascular smooth muscle cells obtained from human umbilical cord vein (UVSMC). AoSMC spontaneously produced both C3 and C4 at 15 ng/10(6) cells/72 hr and 22 ng/10(6) cells/72 hr, respectively, and both were enhanced by interferon-gamma (IFN-gamma). Although phorbol 12-myristate 13-acetate (PMA) and tumour necrosis factor-alpha (TNF-alpha) enhanced C3 production, C4 production was reduced by these agents. On the other hand, G402 produced C4 but not C3 in a dose-dependent manner when cultured with IFN-gamma. UVSMC produced only a small amount of C3 and C4 compared with AoSMC or G402. C3 and C4 produced by AoSMC were confirmed to be identical with their human serum counterparts as determined by sodium dodecyl sulphate-polyacrylamide gel electrophoresis and measurement of haemolytic activity. Northern blotting analysis showed that the expression of mRNA of C3 and C4 was enhanced by TNF-alpha and IFN-gamma, respectively, in AoSMC. Our findings suggest the importance of smooth muscle cells as a source of components of complement in vascular diseases including vasculitis.

Aorta↗
PubMed1996

Diverse effects of tumor necrosis factor-alpha on three subclones from human myelomonocytic leukemia cell line ME-1 exhibiting different differentiation stages.

The effects of tumor necrosis factor-alpha (TNF-alpha) were examined in three subclone cells from human myelomonocytic leukemia cell line ME-1. These three subclone cells exhibit different differentiation stages of the myelomonocytic lineage. TNF-alpha exerted a growth-suppressive effect on the least mature subclone cells, ME-F2 cells. On the other hand, TNF-alpha induced the most mature ME-F1 cells and intermediate ME-F3 cells to differentiate along the monocytic pathway. TNF-alpha also enhanced interferon-gamma (IFN-gamma)-induced complement C2 production by ME-F1 and ME-F3 cells but did not affect production by differentiated ME-F1 and ME-F3 cells. These results suggest that the diversity of the effects of TNF on subclone cells from ME-1 depends on the stage of cell differentiation.

Cell Differentiation↗