• Journal of Life Science
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• v.26 no.7
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• pp.835-846
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• 2016

Chemo-resistance is the biggest issue of effective cancer therapy. ABCG2 is highly correlated with multi-drug resistance, and represent a typical phenotype of multiple cancer stem-like cells. Accumulating evidence recently reported that oncolytic viruses represent a new strategy for multiple aggressive cancers and drug resistant cancers including cancer stem cell-like cells and ABCG2 expressing cells. In this study, we generated an evolutionally engineered vaccinia virus, SLJ-496, for drug-resistant cancer therapy. We first showed that SLJ-496 treatment enhanced tumor affinity using cytopathic effect assay, plaque assay, as well as cell viability assay. Next, we clearly demonstrated that in vitro SLJ-496 treatment represents significant cytotoxic effect in multiple cancers including colorectal cancer cells (HT-29, HCT-116, HCT-8), gastric cancer cells (AGS, NCI-N87, MKN-28), Hepatocellular carcinoma cells (SNU-449, SNU-423, SNU-475, HepG2), as well as mesothelioma cell (NCI-H226, NCI-H28, MSTO-221h). Highly ABCG2 expressing HT-29 cells represent cancer stem like phenotype including stem cell marker expression, and self-renewal bioactivities. Interestingly, we demonstrated that in vitro treatment of SLJ-496 showed significant cytotoxicity effect, as well as viral replication capacity in ABCG2 overexpressing cell. In addition, we also demonstrated the cytotoxic effect of SLJ-496 in Adriamycin-resistant cell lines, SNU-620 and ADR-300. Taken together, these findings provide us a pivotal clue that cancer therapy using SLJ-496 vaccinia virus might be new therapeutic strategy to overcome ABCG2 expressing cancer stem-like cell and multiple chemo-resistance cancer cells.

• Journal of Life Science
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• v.28 no.9
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• pp.1007-1015
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• 2018

The E6-associated protein (E6AP) is known to induce the ubiquitination and proteasomal degradation of HCV core protein and thereby directly impair capsid assembly, resulting in a decline in HCV replication. To counteract this anti-viral host defense system, HCV core protein has evolved a strategy to inhibit E6AP expression via DNA methylation. In the present study, we further explored the mechanism by which HCV core protein inhibits E6AP expression. HCV core protein upregulated both the protein levels and enzyme activities of DNA methyltransferase 1 (DNMT1), DNMT3a, and DNMT3b to inhibit E6AP expression via promoter hypermethylation in HepG2 cells but not in Hep3B cells, which do not express p53. Interestingly, p53 overexpression alone in Hep3B cells was sufficient to activate DNMTs in the absence of HCV core protein and thereby inhibit E6AP expression via promoter hypermethylation. In addition, upregulation of p53 was absolutely required for the HCV core protein to inhibit E6AP expression via promoter hypermethylation, as evidenced by both p53 knockdown and ectopic expression experiments. Accordingly, levels of the ubiquitinated forms of HCV core protein were lower in HepG2 cells than in Hep3B cells. Based on these observations, we conclude that HCV core protein evades ubiquitin-dependent proteasomal degradation in a p53-dependent manner.

• Korean Journal of Veterinary Research
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• v.34 no.3
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• pp.517-527
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• 1994

An attempt was made to isolate a causative viral agents from the fecal specimens of the diseased dogs with the gastroenteritis symptoms. Two coronavirus-like agents were isolated by serial dilution end point method and plaque assay. The isolates were characterized in terms of cytopathology, antigenicity, replication, physicochemical and morphological properties. The results obtained through the experiment were as follows; 1. Among 7 fecal specimens collected from the dogs with enteric disease, 2(28.6%)coronavirus-like agents showing typical cytopathic effects of canine coronavirus were isolated, and designated as CCV D1 and CCV D2, respectively. 2. By the cross-neutralization test and indirect immunofluoresence antibody test, the isolates were antigenically indentified as the standard CCV. The viruses were replicated only in the cytoplasm of A-72 cells. 3. The isolates showed no haemagglutinating activity against the erythrocytes from 11 kinds of animals. 4. The electron microscopic observation for the isolates showed spherical and pleomorphic features, covered with club-shaped projections on the surface. The size of particles was ranged from 70 to 150nm. 5. In one-step growth curve for the isolates in A-72 cells, maximum titers of intracellular vius was $10^{4.6}$ $TCID_{50}/0.1ml$ at 46 hrs postinoculation(pi) of CCV Dl and $10^{4.4}$ $TCID_{50}/0.1ml$ at 34 hrs pi of CCV D2. The maximum titers of extracellular virus was $10^{5.5}$ $TCID_{50}/0.1ml$ at 58 hrs pi of CCV D1 and $10^{5.8}$ $TCID_{50}/0.1ml$ at 46 hrs pi of CCV D2. 6. In physicochemical property test, the isolates were very sensitive to choroform and were found to be RNA virus. The viruses was stable at pH 3.0 for 1 hr and at $22{^{\circ}C}$ for 5 hrs. However, infectivity titers reduced remarkably by treatment with $56{^{\circ}C}$ for 10min.

• Biomedical Science Letters
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• v.9 no.4
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• pp.177-181
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• 2003

The baculovirus/Spodoptera frugiperda (Sf) cell system has become popular for the production of large amounts of the human erythrocyte glucose transporter, GLUT1, heterologously. However, it was not possible to show that the expressed transporter in insect cells could actually transport glucose. The possible reason for this was that the activity of the endogenous insect glucose transporter was extremely high and so rendered transport activity resulting from the expression of exogenous transporter very difficult to detect. Sf21-AE cells are commonly employed as the host permissive cell line to support the baculovirus AcNPV replication and protein synthesis. The cells grow well on TC-100 medium that contains 0.1 % D-glucose as the major carbon source, strongly suggesting the presence of endogenous glucose transporters. However, unlike the human glucose transporter, very little is known about properties of the endogenous sugar transporter(s) in insect cells. Thus, the uptake of 2-deoxy-D-glucose (2dGlc) by Sf21-AE cells and the inhibition of 2dGlc transport in the insect cells by fructose and cytochalasin B were investigated in the present work. The binding assay of cytochalasin B was also performed, which could be used as a functional assay for the endogenous glucose transporter(s) in the insect cells. Sf21-AE cells were infected with the recombinant virus AcNPV-GT or no virus, at a multiplicity of infection (MOI) of 5. Infected cells were resuspended in PBS plus and minus 300 mM fructose, and plus and minus 20 $\mu$M cytochalasin B for use in transport assays. Uptake was measured at 28$^{\circ}C$ for 1 min, with final concentration of 1 mM deoxy-D-glucose, 2-[1,2-$^3$H]- or glucose, L-[l,$^3$H]-, used at a specific radioactivity of 4 Ci/mol. The results obtained demonstrated that the sugar uptake in uninfected cells was stereospecific, and was strongly inhibited by fructose but only poorly inhibitable by cytochalasin B. It is therefore suggested that the Sf21-AE glucose transporter has very low affinity for cytochalasin B, a potent inhibitor of human erythrocyte glucose transporter.

• Korean Journal of Microbiology
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• v.47 no.4
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• pp.342-347
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• 2011

T cells play a key role in viral infection. However, in patients with chronic hepatitis C virus (HCV) infection, HCV-specific T cells are dysfunctional and impaired in the liver, which is the primary site for HCV replication. There are multiple potential mechanisms for HCV-specific T cell dysfunction including induction of immune inhibitory pathways (program death-1; PD-1, cytotoxic t lymphocyte associated antigen-4; CTLA-4) and immune tolerance induced specific for the liver. However, the interaction between hepatocytes and HCV-specific CD8 T cells has not clearly established. In this study, we confirmed huh (human hepatoma) 7.5 cells expressing HLA (human leukocyte antigen) A2 presented antigen to activate HCV-specific CD8 T cells in HLA A2-restricted manner and expression of PD-L (program death ligand) 1 on huh7.5 cells reduced HCV-specific CD8 T cell activation, suggesting an immune modulatory activity. Loss of HCV-specific tetramer responses following antigenic stimulation correlated with increased caspase-3 activity. In addition, PD-L1 on huh7.5 cells rescued HCV-specific CD8 T cells from apoptosis. Our results suggest that the interaction between PD-L1 and PD-1 can recover the function of HCV-specific CD8 T cells in the liver, which could be applied in therapy of HCV chronic infection.

• Korean Journal of Plant Tissue Culture
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• v.25 no.4
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• pp.245-250
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• 1998

Tobacco mosaic virus(TMV) coat protein cDNA was transformed to Nicotiana tabacum cv. NC82 and the transgenic tobacco plants resistant to TMV infection were isolated in the next generation. The expression of TMV coat protein cDNA and genetic stability of the fifth generation of TMV resistant transgenic tobacco plants at the higher temperature were investigated. The TMV coat protein cDNA was amplified by genomic PCR in all the TMV resistant transgenic tobacco plants. The TMV coat protein expressed in the transgenic tobacco plants was detected at very low level by immunoblot hybridization. Even in tansgenic plants that showed the viral symptom only on very late sucker growth (delay type plants), the coat protein expression in the suckers was much less than that of susceptible tobacco infected with TMV. The TMV coat protein expressed in the transgenic tobacco plants was below 0.01% of total protein. Transcription and expression of the coat protein cDNA in delay type plants were observbed at high temperature (38$^{\circ}C$), and TMV replication was suppressed at both 28$^{\circ}C$ and 38$^{\circ}C$. This indicates that unlike the resistance conferred by 'N' gene. TMV resistance of transgenic tobacco plant won't break down at high temperature.

• Journal of Life Science
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• v.32 no.9
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• pp.734-741
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• 2022

Currently, around 40 million people worldwide are living with human immunodeficiency virus (HIV) infection making HIV a critical global health risk. Present therapies for HIV infection consist of drug cocktails that target different steps of the HIV life cycle to prevent infection, replication, and release of the virus. Due to its mutating nature, drug resistance coupled with side-effects of long-term drug use, novel strategies, and pharmaceuticals to treat and manage HIV infection are constant needs and continuously being studied. Plants allocate a major repertoire of chemical diversity and are therefore regarded as an important source of new bioactive agents that can be utilized against HIV. Since the early 1990s, upon recommendations of the World Health Organization, numerous studies reported phytochemicals from different structural classes such as flavonoids, coumarins, tannins and terpenes with strong inhibitory effects against HIV infection. The present review gathered and presented recent research (2021-present) on plant extracts and phytochemicals that exhibit anti-HIV properties with the aim of providing insights into future studies where ethnomedical and underutilized plant sources may yield important natural products against HIV. Considering the relation and importance of HIV treatment with current viral infection risks such as SARS-CoV-2, screening plants for anti-HIV agents is an important step towards the discovery of novel antivirals.

• BMB Reports
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• v.53 no.5
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• pp.248-253
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• 2020

Gene expression in HIV-1 is regulated by the promoters in 5' long-terminal repeat (LTR) element, which contain multiple DNA regulatory elements that serve as binding sites for cellular transcription factors. YY1 could repress HIV-1 gene expression and latent infection. Here, however, we observed that virus production can be increased by YY1 over-expression and decreased under YY1 depleted condition by siRNA treatment. To identify functional domain(s) of YY1 activation, we constructed a number of YY1 truncated mutants. Our data show that full-length YY1 enhances the viral transcription both through U3 and U3RU5 promoters. Moreover, the C-terminal region (296-414 residues) of YY1 is responsible for the transcriptional upregulation, which could be enhanced further in the presence of the viral Tat protein. The central domain of YY1 (155-295 residues) does not affect LTR activity but has a negative effect on HIV-1 gene expression. Taken together, our study shows that YY1 could act as a transcriptional activator in HIV-1 replication, at least in the early stages of infection.

• Toxicological Research
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• v.17
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• pp.157-166
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• 2001

Almost all currently available retroviral vectors based on murine leukemia virus (MLV) contain one or more viral coding sequences. Because these sequences are also present in the packaging genome, it has been suggested that homologous recombination may occur between the same nucleotide sequence in the packaging genome and the vector, resulting in the production of replication competent retrovirus (RCR). Up until now, it has been difficult to completely remove viral coding sequences since some were thought to be involved in the optimum function of the retroviral vector. For example, the gag coding sequence present in almost all available retroviral vectors has been believed to be necessary for efficient viral packaging, while the pol coding sequence present in the highly efficient vector MFG has been thought to be involved in achieving the high levels of gene expression. However, we have now developed a series of retroviral vectors that are absent of any retroviral coding sequences but produce even higher levels of gene expression without compromising viral titer. In these vectors, the intron and exon sequences from heterologous cellular or viral genes are present. When compared to the well known MLV-based vectors, some of these newly developed vectors have been shown to produce significantly higher levels of gene expression for a longer period. In an experimental system that can maximize the production of RCR, our newly constructed vectors produced an absence of RCR. These vectors should prove to be safer than other currently available retroviral vectors containing one or more viral coding sequences.

• BMB Reports
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• v.38 no.4
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• pp.381-385
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• 2005

Severe acute respiratory syndrome (SARS) is an emerging infectious disease associated with a novel coronavirus (CoV) that was identified and molecularly characterized in 2003. Previous studies on various coronaviruses indicate that protein-protein interactions amongst various coronavirus proteins are critical for viral assembly and morphogenesis. It is necessary to elucidate the molecular mechanism of SARS-CoV replication and rationalize the anti-SARS therapeutic intervention. In this study, we employed an in vitro GST pull-down assay to investigate the interaction between the membrane (M) and the nucleocapsid (N) proteins. Our results show that the interaction between the M and N proteins does take place in vitro. Moreover, we provide an evidence that 12 amino acids domain (194-205) in the M protein is responsible for binding to N protein. Our work will help shed light on the molecular mechanism of the virus assembly and provide valuable information pertaining to rationalization of future anti-viral strategies.