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ABSTRACT: Hepatitis B virus (HBV) X protein (HBx) is a key player in HBV replication as well as HBV-induced hepatocellular carcinoma (HCC). However, the pathogenesis of HBV infection and the mechanisms of host-virus interactions are still elusive. In this study, a combination of affinity purification and mass spectrometry was applied to identify the host factors interacting with HBx in hepatoma cells. Thirteen proteins were identified as HBx binding partners. Among them, we first focused on determining the functional significance of the interaction between HBx and hepatocystin. A physical interaction between HBx and hepatocystin was confirmed by co-immunoprecipitation and Western blotting. Immunocytochemistry demonstrated that HBx and hepatocystin colocalized in the hepatoma cells. Domain mapping of both proteins revealed that the HBx C-terminus (amino acids 110-154) was responsible for binding to the mannose 6-phosphate receptor homology domain (amino acids, 419-525) of hepatocystin. Using translation and proteasome inhibitors, we found that hepatocystin overexpression accelerated HBx degradation via a ubiquitin-independent proteasome pathway. We demonstrated that this effect was mediated by an interaction between both proteins using a HBx deletion mutant. Hepatocystin overexpression significantly inhibited HBV DNA replication and expression of HBs antigen concomitant with HBx degradation. Using the hepatocystin mutant constructs that bind HBx, we also confirmed that hepatocystin inhibited HBx-dependent HBV replication. In conclusion, we demonstrated for the first time that hepatocystin functions as a chaperon-like molecule by accelerating HBx degradation, and thereby inhibits HBV replication. Our results suggest that inducing hepatocystin may provide a novel therapeutic approach to control HBV infection.
Biochimica et Biophysica Acta 05/2013; · 4.66 Impact Factor
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Eun-Sook Park,
Yong Kwang Park,
Chanyoung Shin,
Seung Hwa Park, Sung Hyun Ahn,
Doo Hyun Kim,
Keo-Heun Lim,
So Young Kwon,
Kwang Pyo Kim,
Sung-Il Yang,
Baik L Seong,
Kyun-Hwan Kim
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ABSTRACT: Liver regeneration after liver damage caused by toxins and pathogens is critical for liver homeostasis. Retardation of liver proliferation was reported in hepatitis B virus (HBV) X protein (HBx)-transgenic mice. However, the underlying mechanism of the HBx-mediated disturbance of liver regenerationis unknown.We investigated the molecular mechanismof the inhibition of liver regeneration using liver cell lines and a mouse model. The mouse model of acute HBV infection was established by hydrodynamic injection of viral DNA.Liver regeneration after partial hepatectomy was significantly inhibited in the HBV DNA-treatedmice. Mechanism studies have revealed that the expression of urokinase-type plasminogen activator (uPA), which regulates the activation of hepatocyte growth factor (HGF), was significantly decreased in the liver tissues of HBV or HBx-expressing mice. The down-regulation of uPA was further confirmed using liver cell lines transiently or stably transfected with HBx and the HBV genome. HBx suppresseduPA expression through the epigenetic regulationof the uPA promoter in mice liver tissues and human liver cell lines. Expression of HBx strongly induced hyper-methylation of the uPA promoter byrecruiting DNA methyltransferase (DNMT) 3A2.Conclusions:Taken together,these resultssuggest that infection of HBV impairs liver regeneration throughthe epigenetic dysregulation of liver regeneration signals by HBx. (HEPATOLOGY 2013.).
Hepatology 03/2013; · 11.66 Impact Factor
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ABSTRACT: Sustained activation of NF-κB is one of the causative factors for various liver diseases, including liver inflammation and hepatocellular carcinoma (HCC). It has been known that activating the NF-κB signal by hepatitis B virus X protein (HBx) is implicated in the development of HCC. However, despite numerous studies on HBx-induced NF-κB activation, the detailed mechanisms still remain unsolved. Recently, p22-FLIP, a cleavage product of c-FLIPL, has been reported to induce NF-κB activation through interaction with the IκB kinase (IKK) complex in primary immune cells. Since our previous report on the interaction of HBx with c-FLIPL, we explored whether p22-FLIP is involved in the modulation of HBx function. First, we identified the expression of endogenous p22-FLIP in liver cells. NF-κB reporter assay and electrophoretic mobility shift assay (EMSA) revealed that the expression of p22-FLIP synergistically enhances HBx-induced NF-κB activation. Moreover, we found that HBx physically interacts with p22-FLIP and NEMO and potentially forms a ternary complex. Knock-down of c-FLIP leading to the downregulation of p22-FLIP showed that endogenous p22-FLIP is involved in HBx-induced NF-κB activation, and the formation of a ternary complex is necessary to activate NF-κB signaling. In conclusion, we showed a novel mechanism of HBx-induced NF-κB activation in which ternary complex formation is involved among HBx, p22-FLIP and NEMO. Our findings will extend the understanding of HBx-induced NF-κB activation and provide a new target for intervention in HBV-associated liver diseases and in the development of HCC.
PLoS ONE 01/2013; 8(3):e57331. · 4.09 Impact Factor
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ABSTRACT: We investigated the localized surface plasmon resonances of individual AgAuAg nanorods (NRs) using the dark-field spectro-microscopy technique. We find that the scattering spectra of such hetero-NRs show longitudinal resonance wavelengths that are nearly insensitive to the relative composition of Ag and Au. Instead, the resonance is mostly governed by the overall length of the nanorod. This shows that the plasmons oscillate along the entire length of the NR without the significant perturbation at the Ag-Au interfaces. The results demonstrate that the overall geometry as well as the composition determine the tunability of the hetero-metallic nanostructures, and provide an important design rule for the composition-tunable bimetallic plasmon structures.
Physical Chemistry Chemical Physics 12/2012; · 3.57 Impact Factor
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So Young Kwon,
Yong Kwang Park, Sung Hyun Ahn,
Eun Sook Cho,
Won Hyeok Choe,
Chang Hong Lee,
Byung Kook Kim,
Soon Young Ko,
Hyo Sun Choi,
Eun-Sook Park,
Gu Choul Shin,
Kyun-Hwan Kim
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ABSTRACT: Clevudine (CLV) is a nucleoside analog with potent antiviral activity against chronic hepatitis B virus (HBV) infection. Viral resistance to CLV in patients receiving CLV therapy has not been reported. The aim of this study was to characterize CLV-resistant HBV in patients with viral breakthrough (BT) during long-term CLV therapy. The gene encoding HBV reverse transcriptase (RT) was analyzed from chronic hepatitis B patients with viral BT during CLV therapy. Sera collected from the patients at baseline and at the time of viral BT were studied. To characterize the mutations of HBV isolated from the patients, we subjected the HBV mutants to in vitro drug susceptibility assays. Several conserved mutations were identified in the RT domain during viral BT, with M204I being the most common. In vitro phenotypic analysis showed that the mutation M204I was predominantly associated with CLV resistance, whereas L229V was a compensatory mutation for the impaired replication of the M204I mutant. A quadruple mutant (L129M, V173L, M204I, and H337N) was identified that conferred greater replicative ability and strong resistance to both CLV and lamivudine. All of the CLV-resistant clones were lamivudine resistant. They were susceptible to adefovir, entecavir, and tenofovir, except for one mutant clone. In conclusion, the mutation M204I in HBV RT plays a major role in CLV resistance and leads to viral BT during long-term CLV treatment. Several conserved mutations may have a compensatory role in replication. Drug susceptibility assays reveal that adefovir and tenofovir are the most effective compounds against CLV-resistant mutants. These data may provide additional therapeutic options for CLV-resistant patients.
Journal of Virology 02/2010; 84(9):4494-503. · 5.40 Impact Factor
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ABSTRACT: We carried out the near-field optical imaging of isolated and dimerized gold nanocubes to directly investigate the strong coupling between two adjacent nanoparticles. The high-resolution (approximately 10 nm) local field maps (intensities and phases) of self-assembled nanocube dimers reveal antisymmetric plasmon modes that are starkly different from a simple superposition of two monomeric dipole plasmons, which is fully reproduced by the electrodynamics simulations. The result decisively proves that, for the closely spaced pair of nanoparticles (interparticle distance/particle size approximately 0.04), the strong Coulombic attraction between the charges at the interparticle gap dominates over the intraparticle charge oscillations, resulting in a hybridized dimer plasmon mode that is qualitatively different from those expected from a simple dipole-dipole coupling model.
Nano Letters 08/2009; 9(10):3619-25. · 13.20 Impact Factor
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ChemPhysChem 11/2008; 9(17):2491-4. · 3.41 Impact Factor
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ABSTRACT: Scattering-type apertureless near-field microscopy (ANSOM) provides high-resolution dielectric maps of indium gallium nitride (InGaN) semiconductor nanoparticles at visible (633 nm) wavelengths. A specific size-dependent contrast reversal is observed in the ANSOM images of InGaN nanoparticles grown on a layer of gallium nitride (GaN). Model calculations demonstrate that the observed contrast reversal is the result of the competition between the tip-particle versus tip-substrate dipolar coupling.
Nano Letters 09/2007; 7(8):2258-62. · 13.20 Impact Factor
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ABSTRACT: Scattering-type apertureless near-field microscopy (ANSOM) provides high-resolution dielectric maps of indium gallium nitride (InGaN) semiconductor nanoparticles at visible (633 nm) wavelengths. A specific size-dependent contrast reversal is observed in the ANSOM images of InGaN nanoparticles grown on a layer of gallium nitride (GaN). Model calculations demonstrate that the observed contrast reversal is the result of the competition between the tip−particle versus tip−substrate dipolar coupling.
07/2007;
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ABSTRACT: Capillary RPLC/ESI-MS (cRPLC/ESI-MS) is one of the most powerful analytical tools for current proteomic research. The development of cRPLC techniques coupled online to a mass spectrometer has focused on increasing the separation efficiency, detection sensitivity, and throughput. Recently, the use of high-pressure (over 10,000 psi) LC systems that utilize long, small inner diameter capillary columns has gained much attention for proteomic analyses. In this study, we developed an ultrahigh-pressure dual online SPE/capillary RPLC (DO-SPE/cRPLC) system. This LC system employs two online SPE columns and two capillary columns (75 microm inner diameter x 1 m length) in a single separation system, and has a maximum operating pressure of 10,000 psi. This DO-SPE/cRPLC system is capable of providing high-resolution separation in addition to several other advantageous features, such as high reproducibility in terms of the LC retention time, rapid sample injection, online desalting, online sample enrichment of dilute samples, and increased throughput as a result of essentially removing the column equilibration time between successive experiments. We coupled the DO-SPE/cRPLC system online to a tandem mass spectrometer to allow high-throughput proteomic analyses. In this paper, we demonstrate the efficiency of this DO-SPE/cRPLC/MS/MS system by its use in the analyses of proteomic samples exhibiting different levels of complexity.
Electrophoresis 04/2007; 28(6):1012-21. · 3.30 Impact Factor
