• Journal of Microbiology
• /
• v.35 no.2
• /
• pp.152-157
• /
• 1997

Infection of human fibroblast (HF) cells with human cytomegalovirus (HCMV) result in changes in the intracellular level of second messengers. Since nitric oxide (NO) production has been known to be related with other second messengers, it is probable that HCMV infection of HF cells may involve NO. To test this possibility, the amount of NO was measured following ogenous addition of NO generators such as sodium nitroprusside (SNP) or S-nitroso-N-a-cetylpenicillamine (SNAP) immediately after HCMV infection, however, inhibited virus multiplication. Furthermore, immunoblot experiment using monoclonal antibody to HCMV major immediate early (MIE) proteins or CAT assay using pCMVIE/CAT (plasmid containing CAT gene driven by HCMV MIE promoter) revealed that SNP or SNAP blocked the MIE gene expression. SNP was more effective than SNAP in hibiting HCMV multiplication or MIE gene expression. SNP produced more NO than SNAP in inhibiting HCMV multiplication or MIE gene expression. SNP produced more NO than SNAP. Although the mechanism for the inhibition of HCMV multiplication and MIE gene expression by NO is still elusive some correlation with NO-mediated inhibition of HCMV-induced increase in cytosolic free Ca$\^$2+/ concentration ([Ca$\^$2+/]) was observed. The increase of [Ca$\^$2+/] following HCMV infection was inhibited by SNP, and less effectively by SNAP. Raising [Ca$\^$2+/ with bromo-A23187 partially reversed the SNP block of MIE gene expression. Thus, there appear to e some relationships among NO. [Ca$\^$2+/], and HCMV MIE gene expression.

• Bulletin of the Korean Chemical Society
• /
• v.26 no.9
• /
• pp.1385-1389
• /
• 2005

Human hepatitis B virus (HBV) is a pathogen related to the development of liver diseases including chronic hepatitis, cirrhosis, and hepatocellular carcinoma (HCC). However, the efficient methods to suppress HBV replication have not been developed yet. Therefore, we have used RNA interference (RNAi) as a potential tool for the suppression of HBV replication. Here, we designed a 21 nt small intefering dsRNA (siRNA) against hepatitis B virus X (HBx) RNA with 3' overhanging ends derived from T7 promoter. It has been reported that HBV X protein plays an important role in HBV gene expression and viral replication. The suppression of HBx gene expression by the 21 nt siRNA was investigated by Northern blot analysis and chloramphenicol acetyl transferase (CAT) assay. The level of HBx mRNA was decreased by siRNA in a dose-dependent manner. We also found that the 21 nt siRNA inhibited the HBV replication in hepatocellular carcinoma cell.

• BMB Reports
• /
• v.51 no.6
• /
• pp.290-295
• /
• 2018

Y-box binding protein 1 (YB-1) is a member of the cold-shock domain (CSD) protein superfamily. It participates in a wide variety of cellular events, including transcription, RNA splicing, translation, DNA repair, drug resistance, and stress responses. We investigated putative functions of YB-1 in HIV-1 replication. Functional studies using overexpression or knockdown of YB-1 in conjunction with transfection of proviral DNA showed that YB-1 enhances virus production. We found YB-1 regulates HIV-1 production by stimulating viral transcription using HIV-1 LTR sequence U3RU5 with Luciferase assay. We also identified a specific region from amino acids 1 to 324 of YB-1 as necessary for the participation of the protein in the production of virions.

• Fisheries and Aquatic Sciences
• /
• v.18 no.1
• /
• pp.65-71
• /
• 2015

The functional utility of a transgenic marine medaka Oryzias dancena strain carrying the red fluorescent protein (RFP) gene driven by an endogenous choriogenin H (chgH) promoter was evaluated for its ability to detect waterborne $17{\alpha}$-ethinylestradiol (EE2), a synthetic estrogen derivative. The chgH:rfp transgenic marine medaka larvae showed an age-dependent tendency in the efficiency of EE2-mediated transgene expression, in which transgenic larvae older than 6 days post-hatching displayed a more effective response in their transgene expression to EE2 than did younger hatchlings. During experimental exposures to high concentrations of EE2 (200 to 1,000 ng/L), the transgenic responses in the hatchlings were broadly dose- and duration-dependent. With exposures using lower doses of EE2 (25, 50 and 100 ng/L), EE2-induced transgenic RFP was also observed in the transgenic larvae, although the lower doses required exposure of longer duration. Under the EE2 exposure and microscope assay conditions used in our study, transgenic marine medaka larvae exhibited a similar degree of EE2-mediated RFP phenotype expression at various salinity levels (0, 15 and 30 ppt).

• BMB Reports
• /
• v.48 no.11
• /
• pp.597-598
• /
• 2015

Mitochondrial respiratory defect is a key bioenergetics feature of hepatocellular carcinoma (HCC) cells. However, their involvement and roles in HCC development and progression remain unclear. Recently, we identified 10 common mitochondrial defect (CMD) signature genes that may be induced by retrograde signaling-mediated transcriptional reprogramming in response to HCC mitochondrial defects. HCC patients with enriched expression of these genes had poor prognostic outcomes, such as shorter periods of overall survival and recurrence-free survival. Nuclear protein 1 (NUPR1), a key transcription regulator, was up-regulated by Ca⁺⁺-mediated retrograde signaling. NUPR1-centric network analysis and a biochemical promoter-binding assay demonstrated that granulin (GRN) is a key downstream effector of NUPR1 for the regulation of HCC cell invasiveness; association analysis of the NUPR1-GRN pathway supported this conclusion. Mitochondrial respiratory defects and retrograde signaling thus play pivotal roles in HCC progression, highlighting the potential of the NUPR1-GRN axis as a novel diagnostic marker and therapeutic target for HCC.

• Natural Product Sciences
• /
• v.4 no.2
• /
• pp.62-67
• /
• 1998

As a part of search for new biologically active constituents from aloe, we have isolated a glycopeptide, called G1G1M1DI2, from the gel(G1) of Aloe vera. Chemical and spectroscopic evidence indicated that G1G1M1DI2 is a glycopeptide. The molecular weight of G1G1M1DI2 was about 5,500 daltons, and the carbohydrate and protein contents were 20.9% and 32.6%, respectively. Periodate oxidation and enzymic degradation gave peptide moiety and carbohydrate moiety, respectively. Carbohydrate moiety is composed of fucose, galactose, glucose and mannose in a molar ratio of 0.5:2.4;48.8:48.3. Peptide moiety is composed of fifteen amino acids, and glutamic acid and glycine were the major componants. The glycopeptide, G1G1M1DI2, stimulated thymidine uptake of SCC 13 cells about 6.5 times the control. This result suggests that this glycopeptide has a skin cell proliferating activity.

• Korean Journal of Plant Tissue Culture
• /
• v.21 no.1
• /
• pp.55-58
• /
• 1994

Agrobacterium tumefaciens LABA4404 harboring plant binary vector, pBI121, was used for genetic transformation of lettuce (Lactuca sativa t.). Cotyledon segments were infected with A. tumefaciens LBA4404 by cocultivation method and regenerated. Regenerated letture was subject to molecular analyses for integration into plant nuclear genome and expression of ${\beta}$-glucumnidase (GUS) gene. Southern and Northern blot analyses demonstrated that GUS gene was integrated into plant nuclear genome and expressed into its mRNA. The expression of GUS gene into its protein was confirmed by specetrophotometric assay of GUS activity.

• Microbiology and Biotechnology Letters
• /
• v.24 no.1
• /
• pp.72-76
• /
• 1996

To investigate the expression efficiency of new transfer vector of Bombyx mori nuclear polyhedrosis virus (BmNPV), Escherichia coli lacZ gene was inserted into new transfer vector pBmKSK1, under the control of polyhedrin promoter and expressed in BmN-4 cells and larvae of silkworm, Bombyx mori. The recombinant virus containing lacZ gene was isolated from BmN-4 cells coinfected with transfer vectro pBmKSK1-LacZ and wild type BmNPV genome, and analysed by Southern blotting. The expression of ${\beta}$-galactosidase was characterized by SDS-PAGE, Western blotting and ${\beta}$-galactosidase activity assay. The results showed that the level of expression in silkworm larvae was higher than that of BmN-4 cells.

• BMB Reports
• /
• v.39 no.6
• /
• pp.788-793
• /
• 2006

Bacterial DNA containing immunostimulatory CpG motifs can stimulate antigen-presenting cells to express co-stimulatory molecules and to produce various cytokines in vivo and in vitro. In this study, we fragmented macromolecular E.coli genomic DNA with DNase I, and analyzed the ability of the resulting DNA fragments to induce the NF-${\kappa}B$ activation and humoral immune response. Furthermore, using computational analysis and luciferase assay for synthetic ODNs based on the sequence of the immunostimulatory DNA fragments (DF-ODNs), an active component of DF-ODNs sequences was investigated. Experimental results demonstrated that DF-ODN is optimal for the NF-${\kappa}B$-responsive promoter activation in the mouse macrophage cell line and the humoral immune response in vivo. In agreement with the activity of the DF-ODNs processed by DNase I, a synthetic ODN based on the DF-ODN sequences is potent at inducing IL-12 mRNA expression in primary dendritic cells. These results suggest that the discovery and characterization of a highly active natural CpG-ODN may be achieved by the analyses of bacterial DNA fragments generated by a nuclease activity.

• 한국생물공학회:학술대회논문집
• /
• 2002.04a
• /
• pp.261-264
• /
• 2002

CHO (Chinese hamster ovary) cells were transfected with plasmids containing both cis-acting HRE (hypoxia response element) and CMV-promoter that controls tissue-type plasminogen activator (t-PA). CHO cells with HRE produced 16.2 fold higher t-PA concentration than CHO cells without HRE. It was noted that hypoxia strongly induced CHO cell apoptosis. which resulted in decrease of cell viability and protein production. In this study. by introducing Bcl-2, anti-apoptotic gene, we tried to recover cell viability and increase the protein production. When batch culture of both control cells without transfection of Bcl-2 and cells transfected with Bcl-2 were performed in the absence of CoCl ι hypoxia mimic condition. the cells with Bcl-2 were effected specific cell growth rates, maximum cell density. Immunoblotting assay showed Bcl-2 was recombinant with HRE dependent t- P A expression cassette, and their expression level was depended on hypoxia. By introducing Bcl-2, both cell viability and maximum cell density could be increased.