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Biomedical subjects

Akio Nomoto

Publications and source records attributed to Akio Nomoto.

26 records · Page 2Linked to original sources
PubMed2002

The core protein of hepatitis C virus is imported into the nucleus by transport receptor Kap123p but inhibits Kap121p-dependent nuclear import of yeast AP1-like transcription factor in yeast cells.

The core protein of hepatitis C virus (HCV) is a major component of the viral nucleocapsid. The HCV core protein includes nuclear localization signal-like sequences and has various effects on cellular metabolism, playing roles, for example, in the regulation of transcription, apoptosis, and transformation. To examine the possibility of an effect of the core protein on nucleocytoplasmic transport, we used the yeast Saccharomyces cerevisiae as a model system. The core protein (p23) is processed to p21 and is localized in both the cytoplasm and nucleus in yeast cells, similar to that observed in mammalian cells in several cases. The nuclear import of the core protein requires the activity of small GTPase Ran/Gsp1p and is mediated by Kap123p in yeast cells. When the core protein was expressed in yeast cells, the import of the yeast AP1-like transcription factor Yap1p into the nucleus was inhibited. Experiments in vitro involving Kap121p, also known as Pse1p, a receptor for the nuclear import of Yap1p, indicated that the amount of Yap1p bound to Kap121p was reduced in the presence of core protein. These results suggest that the HCV core protein affects cellular metabolism by disturbing transport of proteins to the nucleus.

Active Transport, Cell Nucleus↗
PubMed2002

The tumor suppressor protein TSLC1 is involved in cell-cell adhesion.

TSLC1 is a tumor suppressor gene encoding a member of the immunoglobulin (Ig) superfamily. The significant homology of its extracellular domain with those of other Ig superfamily cell adhesion molecules (IgCAMs) has raised the possibility that TSLC1 participates in cell-cell interactions. In this study, the physiological properties of TSLC1 were investigated in Madin-Darby canine kidney (MDCK) cells expressing TSLC1 tagged with green fluorescent protein (GFP) as well as in the cells that express endogenous TSLC1. Biochemical analysis has revealed that TSLC1 is an N-linked glycoprotein with a molecular mass of 75 kDa and that it forms homodimers through cis interaction within the plane of the cell membranes. Confocal laser scanning microcopy of the cells expressing TSLC1 showed the localization patterns characteristic to adhesion molecules. At the beginning of cell attachment, TSLC1 accumulated in interdigitated structures at cell-cell boundaries, but, when cells reached a confluence, TSLC1 was distributed all along the cell membranes. In polarized cells, TSLC1 was recruited to the lateral membrane, implying trans interaction of TSLC1 between neighboring cells. In support of this notion, MDCK cells expressing TSLC1-GFP showed a significant level of cell aggregation in the absence or presence of Ca(2+) and Mg(2+). Taken together, these results indicate that TSLC1 mediates intracellular adhesion through homophilic interactions in a Ca(2+)/Mg(2+)-independent manner.

Calcium↗
PubMed2002

Isolation of vaccine-derived type 1 polioviruses displaying similar properties to virulent wild strain Mahoney from sewage in Japan.

Type 1, 2, and 3 vaccine-derived polioviruses were isolated from a sewage disposal plant located downstream of the Oyabe River in Toyama Prefecture, Japan, between October 1993 and September 1995. Neurovirulence was analyzed in 13 type 1 vaccine-derived strains, using mutant analysis by polymerase chain reaction and restriction enzyme cleavage (MAPREC). Nine strains (69%) were estimated to have marked neurovirulence. Some of the neutralizing antigenic sites, temperature sensitivity, and plaque-forming ability of two virulent vaccine-derived poliovirus strains were similar to Mahoney strain. The neutralizing activity of human sera obtained after oral poliomyelitis vaccine (OPV) administration against one of the virulent vaccine-derived polioviruses was examined. Although all human sera showed sufficient neutralizing activity for the prevention of poliomyelitis by vaccine-derived poliovirus strains, a lower titer than that against Sabin type 1 strain was observed. Vaccination against virulent vaccine-derived poliovirus will be effective. However, the environmental presence of viruses that have properties similar to those Mahoney strain is a threat. The introduction of inactivated poliovirus vaccine (IPV), and well-maintained herd immunity, together with reinforced environmental surveillance is important for the final phase of the polio eradication program by the World Health Organization (WHO).

Animals↗
PubMed2002

Expression of brain-derived neurotrophic factor in the central nervous system of mice using a poliovirus-based vector.

Brain-derived neurotrophic factor (BDNF) is a promising candidate for the gene therapy of neurological disease. To deliver BDNF to neurons of the central nervous system (CNS), a nucleotide sequence encoding the mature peptide of BDNF was inserted into the genome of poliovirus, a neurotropic virus that is known to replicate mainly in motor neurons of the spinal cord of the CNS. Thus, the recombinant poliovirus constructed was replication-competent. The expression of BDNF in cultured cells infected with the recombinant poliovirus was evident when the cells were analyzed using an immunofluorescence assay and Western blotting. When the recombinant viruses were injected intramuscularly into transgenic mice that carry the human poliovirus receptor gene, the antigens of poliovirus and BDNF were detected in the motor neurons of the spinal cord at 3 days postinfection, and had disappeared by 7 days postinfection. This study suggests that poliovirus can be used as a virus vector for the delivery of neurotrophic factors to the motor neurons of the central nervous system and may provide a new approach for the treatment of motor neuron diseases.

Animals↗
PubMed2002

Isolation and analysis of retroviral integration targets by solo long terminal repeat inverse PCR.

Upon retroviral infection, the genomic RNA is reverse transcribed to make proviral DNA, which is then integrated into the host chromosome. Although the viral elements required for successful integration have been extensively characterized, little is known about the host DNA structure constituting preferred targets for proviral integration. In order to elucidate the mechanism for the target selection, comparison of host DNA sequences at proviral integration sites may be useful. To achieve simultaneous analysis of the upstream and downstream host DNA sequences flanking each proviral integration site, a Moloney murine leukemia virus-based retroviral vector was designed so that its integrated provirus could be removed by Cre-loxP homologous recombination, leaving a solo long terminal repeat (LTR). Taking advantage of the solo LTR, inverse PCR was carried out to amplify both the upstream and downstream cellular flanking DNA. The method called solo LTR inverse PCR, or SLIP, proved useful for simultaneously cloning the upstream and downstream flanking sequences of individual proviral integration sites from the polyclonal population of cells harboring provirus at different chromosomal sites. By the SLIP method, nucleotide sequences corresponding to 38 independent proviral integration targets were determined and, interestingly, atypical virus-host DNA junction structures were found in more than 20% of the cases. Characterization of retroviral integration sites using the SLIP method may provide useful insights into the mechanism for proviral integration and its target selection.

Animals↗
PubMed2001

Suppression of apoptotic and necrotic cell death by poliovirus.

To determine an antiapoptotic activity of poliovirus type 1 (PV-1), we examined the effect of PV-1 infection on apoptosis that was induced in HEp-2 cells by the treatment with 1 M sorbitol. The virus did not induce apoptosis in the infected cells and could suppress both the fragmentation of chromosomal DNA and morphological cell and cell nuclei changes in the sorbitol-treated cells, indicating that PV-1 induces an antiapoptotic state. Comparison of the kinetics showed that this ability of the virus appeared in the infected cells at the time of progeny virus formation (maturation step of virus multiplication). Simultaneously with this antiapoptotic activity, PV-1 infection also suppressed non-apoptotic cell death induced by sodium chloride. Electron microscopic observation revealed that the cells killed by the sodium chloride treatment had undergone liquefactive necrosis, indicating that PV-1 can inhibit both apoptosis and necrosis. In addition, PV-1 can grow in the apoptotic cells, although the virus yield was reduced to a quarter of the yield in normal cells.

Apoptosis↗
PubMed1999

Lower concentration of La protein required for internal ribosome entry on hepatitis C virus RNA than on poliovirus RNA.

Translation initiation of poliovirus and hepatitis C virus (HCV) RNA occurs by entry of ribosomes to the internal RNA sequence, called the internal ribosomal entry site (IRES). Both IRES bind to the La protein and are thought to require the protein for their translation initiation activity, although they are greatly different in both the primary and predicted secondary structures. To compare the La protein requirement for these IRES, we took advantage of I-RNA from the yeast Saccharomyces cerevisiae, which has been reported to bind to La protein and block poliovirus IRES-mediated translation initiation. In a cell-free translation system prepared from HeLa cells, yeast I-RNA inhibited translation initiation on poliovirus RNA as expected, but did not significantly inhibit translation initiation on HCV RNA. However, the translation initiation directed by either IRES was apparently inhibited by I-RNA in rabbit reticulocyte lysates, in which La protein is limiting. I-RNA-mediated inhibition of HCV IRES-dependent translation in rabbit reticulocyte lysates was reversed by exogenous addition of purified recombinant La protein of smaller amounts than necessary to reverse poliovirus IRES-dependent translation. These results suggest that HCV IRES requires lower concentrations of La protein for its function than does poliovirus IRES. Immunofluorescence studies showed that HCV infection appeared not to affect the subcellular localization of La protein, which exists mainly in the nucleus, although La protein redistributed to the cytoplasm after poliovirus infection. The data are compatible with the low requirement of La protein for HCV IRES activity.

Animals↗