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H Shida

Publications and source records attributed to H Shida.

At least 73 records · Page 4Linked to original sources
PubMed1990

Hemadsorption and fusion inhibition activities of hemagglutinin analyzed by vaccinia virus mutants.

Vaccinia virus IHD-J strain induces hemagglutinin (HA) on the surface membrane of infected cells and does not elicit cell-cell fusion (F-). We isolated 21 independent hemadsorption-negative (HAD-) mutant viruses from IHD-J and five HAD+ revertants from one of these mutants. Of the 21 mutants, 19 that synthesized either no or little HA at the cell surface caused cell-cell fusion (F+), whereas none of the five revertants that synthesized HA at the cell surface induced cell-cell fusion. Furthermore, anti-HA monoclonal antibody B2D10 induced extensive polykaryocytosis of IHD-J-infected cells and suppressed the ability of the IHD-J-infected cell extract to inhibit the polykaryocytosis induced by IHD-W. The other 2 of the 21 HAD- mutants, B1 and A2, which induced HAs at the cell surface, showed F- and F+ phenotype, respectively. The HA molecule of mutant B1 had a single amino acid substitution of Lys for Glu-121 in its extracellular domain, whereas that of mutant A2 had a single substitution mutation of Tyr for Cys-103. We conclude that the vaccinia HA is a fusion inhibition protein, that the active sites for the two activities reside separately in its extracellular domain, and that cysteine-103 is important in forming the proper tertiary structure of the protein to exert both activities.

Animals↗
PubMed1990

Characterization of antibody and cytotoxic T lymphocyte responses to human influenza virus H3 haemagglutinin expressed from the haemagglutinin locus of vaccinia virus.

Antibody and cytotoxic T lymphocyte (CTL) responses to the haemagglutinin (HA) of human H3N2 influenza virus were analysed, using recombinant vaccinia viruses containing the influenza HA gene inserted into the HA gene locus of vaccinia virus. The recombinant vaccinia viruses elicited a high haemagglutination inhibiting (HI) antibody response to the homologous influenza virus in mice. In addition, HI antibody generated by the recombinant vaccinia virus reacted with antigenic variants of human H3N2 influenza virus in a manner similar to that elicited by the HA vaccine. Mice with a high response to influenza virus HA vaccine were highly responsive to the HA expressed from the recombinant vaccinia virus, as measured by HI antibody production. The immunogenicity of the influenza virus HA expressed by the recombinant seems to be attributable to the intrinsic immunogenicity of the HA molecule. The recombinants primed mice for an influenza virus H3-specific CTL response and primed CTLs recognized the target cells in a subtype-specific manner. The results indicate that a recombinant vaccinia virus derived by the insertion of a foreign gene into its HA gene locus is a potent live vaccine not only for eliciting a high antibody response but also for priming a specific CTL response.

Animals↗
PubMed1990

Biological and immunological characterization of influenza virus haemagglutinin expressed from the haemagglutinin locus of vaccinia virus.

Plasmid vectors were constructed to facilitate the insertion and expression of a foreign gene in the haemagglutinin (HA) gene locus of vaccinia virus. Five unique cloning sites adjacent to the P7.5 promoter of vaccinia virus permit the rapid insertion of a foreign sequence coding for a protein into these plasmids. This vector system provides a simple procedure to select recombinant viruses because they can be readily identified on the basis of their HA-defective phenotype. Recombinant vaccinia viruses expressing influenza virus HA were constructed to characterize the possible use of this system. The recombinant viruses did express the influenza HA through the authentic pathway of biosynthesis. In addition to having immunological characteristics similar to the authentic influenza HA, the expressed HA was found to possess haemagglutinating, haemadsorption and acid-inducible fusion activities. These findings demonstrate the usefulness of this eukaryotic vector system.

Animals↗
PubMed1990

Functional comparison of transactivation by human retrovirus rev and rex genes.

The effect of rev-responsive element deletion on human immunodeficiency virus type 1 (HIV-1) and type 2 (HIV-2) gene expression was examined. The phenotypes of HIV-1 and HIV-2 provirus DNAs lacking the rev-responsive element, as determined by transfection experiments, were indistinguishable from those of virus DNAs carrying rev gene mutations. By using rev-response elements derived from these two viruses, we developed two monitoring systems to evaluate the functionality of HIV-1 rev, HIV-2 rev, and human T-lymphotropic virus type I rex. In both systems, HIV-1 rev and human T-lymphotropic virus type I rex transactivated HIV-2 very efficiently. On the contrary, HIV-2 rev and human T-lymphotropic virus type I rex were poor activators of HIV-1. No functional replacement of rex by HIV-2 rev was observed.

Blotting, Northern↗
PubMed1990

Effects of a highly basic region of human immunodeficiency virus Tat protein on nucleolar localization.

Human immunodeficiency virus type 1 encodes a positive trans-activator protein, Tat, which is located predominantly in the cell nucleolus. To study the role of the basic region of Tat in nucleolar localization, we constructed fusion genes encoding serially deleted segments of Tat joined to the amino-terminal end of the Escherichia coli beta-galactosidase molecule. We show that the basic region of Tat was sufficient for nuclear localization but not for nucleolar localization. Addition of three amino acids (59, 60, and 61) of the Tat sequence at the C-terminal end of the basic region was necessary for the chimeric beta-galactosidase to localize in the nucleus as well as in the nucleolus. We demonstrate that a short amino acid sequence (G-48 RKKRRQRRRA HQ N-61), when fused to the amino terminus of beta-galactosidase, can act as a nucleolar localization signal.

Amino Acid Sequence↗
PubMed1990

Effect of resin use in the post-embedding procedure on immunoelectron microscopy of membranous antigens, with special reference to sensitivity.

To investigate quantitatively the effect of resins on the sensitivity of immunoelectron microscopy of membranous antigen, ultra-thin sections of bovine epithelial tissue embedded in five different kinds of resins [JB-4 (JB4), LR Gold (LRG), Lowicryl K4M (K4M), Quetol 812 (Q812), and Spurr's (Spurr) resin] were labeled specifically with anti-desmosomal glycoprotein I(DGI) antibody followed by protein A-gold (PAG) conjugates. When we compared the labeling intensity expressed as the number of PAG particles per 500-nm length of the desmosomal region along the membrane, three hydrophilic resins (JB4, LRG, and K4M) showed much greater levels of labeling intensity than did epoxy resins (Q812 and Spurr), which had a negative value. The three hydrophilic resins showed only minor differences in their levels of labeling intensity. The intensity obtained with JB4, which was the highest of the three, was further increased by pretreatment of the ultra-thin sections with methyl methacrylate monomer (MM) for 5 min. On the basis of these results, wide applicability of this new technique for membranous antigens, which have been difficult to detect positively by any previously employed techniques, is suggested.

Animals↗
PubMed1989

Rat cytotoxic T lymphocytes against human T-lymphotropic virus type 1-infected cells recognize gag gene and env gene encoded antigens.

T cell immune responses in syngeneic WKA/H rats were analyzed by using lymphoid cell lines, TARS-1, TART-1, and TARL-2, infected with human T-lymphotropic virus type 1 (HTLV-1). Spleen cells of rats in which these cell lines had been rejected were sensitized in vitro with the same cell lines, and cells cytotoxic to these HTLV-1+ cell lines, and cells cytotoxic to these HTLV-1+ cell lines were generated. The effector cells were CTL of the CD5+ CD8+ phenotype and showed restriction of MHC class I Ag. Direct tests as well as cold target cell inhibition tests with an array of cell populations showed that these CTL reacted only with syngeneic HTLV-1+ cell lines. When xenogeneic HTLV-1+ cell lines were similarly utilized for in vitro sensitization, rat CTL specific for syngeneic HTLV-1+ cells were generated. They were not, however, reactive with xenogeneic HTLV-1+ cells used for sensitization. Syngeneic rat cells selectively expressing gag, env, or pX gene coded Ag were prepared by infection of recombinant vaccinia viruses. In cold target cell inhibition tests of anti-HTLV-1 CTL with thus prepared cells, cytotoxicity against the syngeneic HTLV-1+ cells line, TARS-1, was inhibited by syngeneic cells expressing gag gene or env gene coded Ag. Inhibition was, however, more consistent and more dominant by cells with gag gene than those with env gene. Syngeneic cells with pX gene and MHC class I incompatible cells with gag, env, or pX gene did not inhibit cytotoxicity.

Animals↗
PubMed1989

Development of heat-stable recombinant rinderpest vaccine.

Recombinant vaccinia virus (RVV) containing the full-length cDNA of rinderpest virus (RV)-haemagglutinin (H) gene was constructed. The H gene was inserted into the attenuated vaccine strain of vaccinia virus (VV), Le 16 m0, with two different promoters, namely cowpox virus A-type inclusion body (ATI) promoter or VV 7.5 kilodalton (P7.5) promoter. These RVVs produced the same sized fully glycosylated RV-H protein in RK 13 cells as that of the authentic RV-H. Their heat stability in the lyophylized state was similar to that of the parental VV. All rabbits immunized with these RVVs produced virus neutralizing (VN) antibody to RV as well as anti RV-H antibody. Four weeks after immunization, these animals were challenged with RV intravenously. None of the RVV-immunized rabbits developed any clinical signs of RV infection except one which was immunized with RVV containing the ATI promoter and developed low VN titer. These results indicate the possibility of developing a heat-stable recombinant vaccine for the eradication of rinderpest in tropical countries without cold storage systems.

Animals↗
PubMed1989

Fiberoptic sigmoidoscopy as the first screening procedure for colorectal neoplasms in an asymptomatic population.

As the first screening for colorectal neoplasms, a total of 2243 examinations by fiberoptic sigmoidoscopy (FS) were performed on 1573 asymptomatic patients. One hundred seventy neoplasms were found in 134 patients (6 percent of total examinations), including nine adenocarcinomas (0.4 percent). A flat, intramucosal cancer and a small, 5-mm cancer detected by FS could not be demonstrated by barium enema examination. On further investigation, additional adenomas were identified in the proximal colon in 22 percent of patients in whom neoplasms had first been detected by FS and in only 4.4 percent of those in whom FS findings were negative but other criteria were positive. It is concluded that FS is the procedure of choice for the first screening of asymptomatic patients, not only because it enables detection of flat and small lesions within its observation range, but also because it could lead to detection of proximal neoplasms.

Adult↗
PubMed1988

Sequence requirements for nucleolar localization of human T cell leukemia virus type I pX protein, which regulates viral RNA processing.

The posttranscriptional regulator (p27x-III) of human T cell leukemia virus type I (HTLV-I) is located predominantly in the cell nucleolus. A highly basic amino-terminal sequence (NH2-Met-Pro-Lys-Thr-Arg-Arg-Arg-Pro-Arg-Arg-Ser-Gln-Arg-Lys-Arg-Pro-Pro -Thr- Pro) in this protein, when fused to the amino termini of beta-galactosidase and p40x of HTLV-I, acts as an autonomous signal capable of directing the hybrid proteins to the cell nucleolus.

Amino Acid Sequence↗
PubMed1988

Immunogenicity of a recombinant vaccinia virus expressing envelope a glycoprotein of bovine leukaemia virus.

We constructed a recombinant vaccinia virus (RVV) expressing envelope (env) glycoprotein (gp51) of bovine leukaemia virus (BLV): the expression of gp51 was detected by Western blot of the lysates of rabbit kidney cells infected with the RVV. The rabbits inoculated intradermally with the RVV alone failed to induce detectable anti-gp51 antibodies even 10 weeks after immunization. However, when these animals were boosted with inactivated BLV virion in saline, significant levels of anti-gp51 antibodies were induced as shown both in Western blot and immunodiffusion analyses. In these animals, antibodies against gag product (p24) were not detected. On the other hand, the rabbits inoculated with wild-type vaccinia virus and boosted similarly three times with the BLV virion in saline did not induce detectable anti-gp51 antibodies at all. The present experiment revealed that the RVV possessed the capability to endow immunological memory without inducing apparent anti-gp51 antibody responses, meaning that the RVV activated helper T cells far more strongly than B cells. The applicability of the RVV to vaccine development is discussed.

Animals↗
PubMed1988

Cloning and characterization of the gene encoding the major protein of the A-type inclusion body of cowpox virus.

We have mapped and determined the nucleotide sequence of the cowpox virus (CPV) gene coding for the protein of A-type inclusion bodies (ATI). The ATI gene was mapped to the central part of the largest HindIII restriction fragment of the CPV genome. Vaccinia virus (VV) also contained a similar sequence at the equivalent position in its genome, which even under stringent conditions hybridized with the CPV ATI gene. Nucleotide sequence analysis revealed that the upstream region of the coding frame contained a sequence closely resembling the VV late promoter consensus sequence and that the gene encoded a protein of Mr 150,000.

Amino Acid Sequence↗
PubMed1988

Processing of gag precursor polyprotein of human T-cell leukemia virus type I by virus-encoded protease.

The biological activity encoded in the putative protease gene (pro) of human T-cell leukemia virus type I was investigated by using a vaccinia virus expression vector. The 53-kilodalton gag precursor polyprotein was processed into the mature p19, p24, and p15 gag proteins when the gag and protease-coding sequence was expressed under the control of a vaccinia virus promoter, suggesting that the protease may be synthesized through the mechanism of ribosomal frame shifting. The processing defect of a protease mutant could be complemented by cointroduction of a wild-type construct into the cell, demonstrating that the pro gene encodes the biologically active protease molecules which are capable of processing the gag precursor polyprotein in vivo in trans. A study involving the use of a variety of mutants constructed in vitro revealed that the protease consists of a nonessential carboxy-terminal region and a part essential for its activity, including the putative catalytic residue, aspartic acid. Furthermore, a cluster of adenine residues positioned at the overlapping region between the gag and pro genes was shown to be involved in the ribosomal frameshifting event for the synthesis of protease. To mimic the formation of the 76-kilodalton gag-pro precursor polyprotein formed by ribosomal slipping, the coding frames of the gag and pro gene were adjusted. The processing of the gag-pro precursor polyprotein depended on an intact protease gene, implying that a cis-acting function of human T-cell leukemia virus type I protease may be necessary to trigger the initial cleavage event that leads to the release of protease from the precursor protein.

Amino Acid Sequence↗
PubMed1988

Effects and virulences of recombinant vaccinia viruses derived from attenuated strains that express the human T-cell leukemia virus type I envelope gene.

We constructed recombinant vaccinia viruses (RVVs) that expressed human T-cell leukemia virus type I (HTLV-I) envelope glycoproteins by using attenuated vaccinia viruses (VVs) which have much lower neurovirulence than the WR strain that is extensively used as a vector. The RVV produced from the LC16mO strain, one of the attenuated VVs, elicited a high titer of anti-HTLV-I antibody in rabbits and protected them against HTLV-I infection. The env gene was inserted into the VV hemagglutinin gene. The resultant inactivation of the hemagglutinin gene led to the attenuation of VVs, but the extent of their attenuation depended on the VV strain. The propagation of LC16mO and its RVV in rabbit brain was poorer than that of LO-1, a cloned derivative of Lister strain, and its RVV, although LC16mO replicated in other organs better than did LO-1. Taken together, these results suggest that LC16mO is a good candidate as a vector for vaccination of humans.

Animals↗
PubMed1987

On the molecular organization, diversity and functions of desmosomal proteins.

After a brief review of epithelial cell junctions, the authors describe a series of observations and experiments directed toward elucidating the molecular organization and functions of the proteins of desmosomes (maculae adherentes). Their immunofluorescence observations reveal differences between the proteins of junctional complex and other desmosomes of the same cells. Quantitative immuno-localization studies are described using protein A-colloidal gold markers and a series of monospecific, polyclonal antibodies directed against cytokeratins and four desmosomal protein families. It is shown that desmoplakin III (about 81 kDa) is restricted to the desmosomal cytoplasmic plaque, which possesses outer and inner dense zones. Desmoplakins I/II (about 240 kDa and 210 kDa) extend across the same range and beyond into the intermediate filament-rich cytoplasm. Two glycoprotein families, desmoglein I (DGI; about 150 kDa) and desmoglein II (DGII; about 97-118 kDa), extend from the desmosomal midline (or beyond) across the desmoglea (intercellular space), through the plasma membrane and across both layers of the plaque, terminating near its cytoplasmic border. Cytokeratins do not extend into the desmosome. With new procedures utilizing guanidine HCl for preparing and fractionating desmosomal proteins, DGII has been purified to homogeneity. DGII, DGI and a third protein are all shown to bind Ca2+, which is known to promote desmosome assembly.

Animals↗