Publications by authors named "Lily Hui-Ching Wang"

32 Publications

A novel platform for discovery of differentially expressed microRNAs in patients with repeated implantation failure.

Fertil Steril 2021 Apr 3. Epub 2021 Apr 3.

Institute of Molecular and Cellular Biology, National Tsing Hua University, Hsinchu, Taiwan; Department of Medical Science, National Tsing Hua University, Hsinchu, Taiwan. Electronic address:

Objective: To identify predictor microRNAs (miRNAs) from patients with repeated implantation failure (RIF).

Design: Systemic analysis of miRNA profiles from the endometrium of patients undergoing in vitro fertilization (IVF).

Setting: University research institute, private IVF center, and molecular testing laboratory.

Patient(s): Twenty five infertile patients in the discovery cohort and 11 patients in the validation cohort.

Interventions(s): None.

Main Outcome Measure(s): A signature set of miRNA associated with the risk of RIF.

Result(s): We designed a reproductive disease-related PanelChip to access endometrium miRNA profiles in patients undergoing IVF. Three major miRNA signatures, including hsa-miR-20b-5p, hsa-miR-155-5p, and hsa-miR-718, were identified using infinite combination signature search algorithm analysis from 25 patients in the discovery cohort undergoing IVF. These miRNAs were used as biomarkers in the validation cohort of 11 patients. Finally, the 3-miRNA signature was capable of predicting patients with RIF with an accuracy >90%.

Conclusion(s): Our findings indicated that specific endometrial miRNAs can be applied as diagnostic biomarkers to predict RIF. Such information will definitely help to increase the success rate of implantation practice.
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http://dx.doi.org/10.1016/j.fertnstert.2021.01.055DOI Listing
April 2021

Aerobic glycolysis supports hepatitis B virus protein synthesis through interaction between viral surface antigen and pyruvate kinase isoform M2.

PLoS Pathog 2021 Mar 15;17(3):e1008866. Epub 2021 Mar 15.

Institute of Molecular and Cellular Biology, National Tsing Hua University, Hsinchu, Taiwan.

As an intracellular pathogen, the reproduction of the hepatitis B virus (HBV) depends on the occupancy of host metabolism machinery. Here we test a hypothesis if HBV may govern intracellular biosynthesis to achieve a productive reproduction. To test this hypothesis, we set up an affinity purification screen for host factors that interact with large viral surface antigens (LHBS). This identified pyruvate kinase isoform M2 (PKM2), a key regulator of glucose metabolism, as a binding partner of viral surface antigens. We showed that the expression of viral LHBS affected oligomerization of PKM2 in hepatocytes, thereby increasing glucose consumption and lactate production, a phenomenon known as aerobic glycolysis. Reduction of PKM2 activity was also validated in several different models, including HBV-infected HepG2-NTCP-C4 cells, adenovirus mediated HBV gene transduction and transfection with a plasmid containing complete HBV genome on HuH-7 cells. We found the recovery of PKM2 activity in hepatocytes by chemical activators, TEPP-46 or DASA-58, reduced expressions of viral surface and core antigens. In addition, reduction of glycolysis by culturing in low-glucose condition or treatment with 2-deoxyglucose also decreased expressions of viral surface antigen, without affecting general host proteins. Finally, TEPP-46 largely suppressed proliferation of LHBS-positive cells on 3-dimensional agarose plates, but showed no effect on the traditional 2-dimensional cell culture. Taken together, these results indicate that HBV-induced metabolic switch may support its own translation in hepatocytes. In addition, aerobic glycolysis is likely essential for LHBS-mediated oncogenesis. Accordingly, restriction of glucose metabolism may be considered as a novel strategy to restrain viral protein synthesis and subsequent oncogenesis during chronic HBV infection.
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http://dx.doi.org/10.1371/journal.ppat.1008866DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8009439PMC
March 2021

Identifying primate ACE2 variants that confer resistance to SARS-CoV-2.

Mol Biol Evol 2021 Mar 1. Epub 2021 Mar 1.

Biodiversity Research Center, Academia Sinica, Nankang, Taipei 11529, Taiwan.

SARS-CoV-2 infects humans through the binding of viral S-protein (spike protein) to human ACE2 (angiotensin I converting enzyme 2). The structure of the ACE2-S-protein complex has been deciphered and we focused on the 27 ACE2 residues that bind to S-protein. From human sequence databases, we identified 9 ACE2 variants at ACE2-S-protein binding sites. We used both experimental assays and protein structure analysis to evaluate the effect of each variant on the binding affinity of ACE2 to S-protein. We found one variant causing complete binding disruption, two and three variants, respectively, strongly and mildly reducing the binding affinity, and two variants strongly enhancing the binding affinity. We then collected the ACE2 gene sequences from 57 non-human primates. Among the six apes and 20 Old World monkeys (OWMs) studied we found no new variants. In contrast, all 11 New World monkeys (NWMs) studied share four variants each causing a strong reduction in binding affinity, the Philippine tarsier also possesses three such variants, and 18 of the 19 prosimian species studied share one variant causing a strong reduction in binding affinity. Moreover, one OWM and three prosimian variants increased binding affinity by > 50%. Based on these findings we proposed that the common ancestor of primates was strongly resistant to and that of NWMs was completely resistant to SARS-CoV-2 and so is the Philippine tarsier, whereas apes and OWMs, like most humans, are susceptible. This study increases our understanding of the differences in susceptibility to SARS-CoV-2 infection among primates.
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http://dx.doi.org/10.1093/molbev/msab060DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7989403PMC
March 2021

Kinetic Characterization and Inhibitor Screening for the Proteases Leading to Identification of Drugs against SARS-CoV-2.

Antimicrob Agents Chemother 2021 03 18;65(4). Epub 2021 Mar 18.

Institute of Biological Chemistry, Academia Sinica, Taipei, Taiwan

Coronavirus (CoV) disease 2019 (COVID-19), caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has claimed many lives worldwide and is still spreading since December 2019. The 3C-like protease (3CL) and papain-like protease (PL) are essential for maturation of viral polyproteins in SARS-CoV-2 life cycle and thus regarded as key drug targets for the disease. In this study, 3CL and PL assay platforms were established, and their substrate specificities were characterized. The assays were used to screen collections of 1,068 and 2,701 FDA-approved drugs. After excluding the externally used drugs which are too toxic, we totally identified 12 drugs as 3CL inhibitors and 36 drugs as PL inhibitors active at 10 μM. Among these inhibitors, six drugs were found to suppress SARS-CoV-2 with the half-maximal effective concentration (EC) below or close to 10 μM. This study enhances our understanding on the proteases and provides FDA-approved drugs for prevention and/or treatment of COVID-19.
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http://dx.doi.org/10.1128/AAC.02577-20DOI Listing
March 2021

Development and Evaluation of Vero Cell-Derived Master Donor Viruses for Influenza Pandemic Preparedness.

Vaccines (Basel) 2020 Oct 25;8(4). Epub 2020 Oct 25.

National Institution of Infectious Diseases and Vaccinology, National Health Research Institutes (NHRI), Zhunan, Miaoli 35053, Taiwan.

The embryonated egg-based platform currently produces the majority of seasonal influenza vaccines by employing a well-developed master donor virus (MDV, A/PR/8/34 (PR8)) to generate high-growth reassortants (HGRs) for A/H1N1 and A/H3N2 subtypes. Although the egg-based platform can supply enough seasonal influenza vaccines, it cannot meet surging demands during influenza pandemics. Therefore, multi-purpose platforms are desirable for pandemic preparedness. The Vero cell-based production platform is widely used for human vaccines and could be a potential multi-purpose platform for pandemic influenza vaccines. However, many wild-type and egg-derived influenza viruses cannot grow efficiently in Vero cells. Therefore, it is critical to develop Vero cell-derived high-growth MDVs for pandemic preparedness. In this study, we evaluated two in-house MDVs (Vero-15 and VB5) and two external MDVs (PR8 and PR8-HY) to generate Vero cell-derived HGRs for five avian influenza viruses (AIVs) with pandemic potentials (H5N1 clade 2.3.4, H5N1 clade 2.3.2.1, American-lineage H5N2, H7N9 first wave and H7N9 fifth wave). Overall, no single MDV could generate HGRs for all five AIVs, but this goal could be achieved by employing two in-house MDVs (vB5 and Vero-15). In immunization studies, mice received two doses of Vero cell-derived inactivated H5N1 and H7N9 whole virus antigens adjuvanted with alum and developed robust antibody responses.
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http://dx.doi.org/10.3390/vaccines8040626DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7712210PMC
October 2020

Investigating Core Signaling Pathways of Hepatitis B Virus Pathogenesis for Biomarkers Identification and Drug Discovery via Systems Biology and Deep Learning Method.

Biomedicines 2020 Aug 31;8(9). Epub 2020 Aug 31.

Laboratory of Automatic Control, Signal Processing and Systems Biology, Department of Electrical Engineering, National Tsing Hua University, Hsinchu 30013, Taiwan.

Hepatitis B Virus (HBV) infection is a major cause of morbidity and mortality worldwide. However, poor understanding of its pathogenesis often gives rise to intractable immune escape and prognosis recurrence. Thus, a valid systematic approach based on big data mining and genome-wide RNA-seq data is imperative to further investigate the pathogenetic mechanism and identify biomarkers for drug design. In this study, systems biology method was applied to trim false positives from the host/pathogen genetic and epigenetic interaction network (HPI-GEN) under HBV infection by two-side RNA-seq data. Then, via the principal network projection (PNP) approach and the annotation of KEGG (Kyoto Encyclopedia of Genes and Genomes) pathways, significant biomarkers related to cellular dysfunctions were identified from the core cross-talk signaling pathways as drug targets. Further, based on the pre-trained deep learning-based drug-target interaction (DTI) model and the validated pharmacological properties from databases, i.e., drug regulation ability, toxicity, and sensitivity, a combination of promising multi-target drugs was designed as a multiple-molecule drug to create more possibility for the treatment of HBV infection. Therefore, with the proposed systems medicine discovery and repositioning procedure, we not only shed light on the etiologic mechanism during HBV infection but also efficiently provided a potential drug combination for therapeutic treatment of Hepatitis B.
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http://dx.doi.org/10.3390/biomedicines8090320DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7555687PMC
August 2020

Heparan sulfate targeting strategy for enhancing liposomal drug accumulation and facilitating deep distribution in tumors.

Drug Deliv 2020 Dec;27(1):542-555

Institute of Molecular and Cellular Biology, National Tsing Hua University, Hsinchu, Taiwan.

Nanoparticles (NPs), such as liposomes, effectively evade the severe toxicity of unexpected accumulation and passively shuttle drugs into tumor tissues by enhanced permeability and retention. In the case of non-small cell lung cancer and pancreatic ductal adenocarcinoma, cancer-associated fibroblasts promote the aggregation of a gel-like extracellular matrix that forms a physical barrier in the desmoplastic stroma of the tumor. These stroma are composed of protein networks and glycosaminoglycans (GAGs) that greatly compromise tumor-penetrating performance, leading to insufficient extravasation and tissue penetration of NPs. Moreover, the presence of heparan sulfate (HS) and related proteoglycans on the cell surface and tumor extracellular matrix may serve as molecular targets for NP-mediated drug delivery. Here, a GAG-binding peptide (GBP) with high affinity for HS and high cell-penetrating activity was used to develop an HS-targeting delivery system. Specifically, liposomal doxorubicin (L-DOX) was modified by post-insertion with the GBP. We show that the uptake of L-DOX in A549 lung adenocarcinoma cells increased by GBP modification. Cellular uptake of GBP-modified L-DOX (L-DOX-GBP) was diminished in the presence of extracellular HS but not in the presence of other GAGs, indicating that the interaction with HS is critical for the cell surface binding of L-DOX-GBP. The cytotoxicity of doxorubicin positively correlated with the molecular composition of GBP. Moreover, GBP modification improved the distribution and anticancer efficiency of L-DOX, with enhanced desmoplastic targeting and extensive distribution. Taken together, GBP modification may greatly improve the tissue distribution and delivery efficiency of NPs against HS-abundant desmoplastic stroma-associated neoplasm.
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http://dx.doi.org/10.1080/10717544.2020.1745326DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7170378PMC
December 2020

Cell Penetrating Peptide as a High Safety Anti-Inflammation Ingredient for Cosmetic Applications.

Biomolecules 2020 01 7;10(1). Epub 2020 Jan 7.

Institute of Molecular and Cellular Biology, National Tsing Hua University, Hsinchu 30013, Taiwan.

Cosmeceutical peptides have become an important topic in recent decades in both academic and industrial fields. Many natural or synthetic peptides with different biological functions including anti-ageing, anti-oxidation, anti-infection and anti-pigmentation have been developed and commercialized. Current cosmeceutical peptides have already satisfied most market demand, remaining: "cargos carrying skin penetrating peptide with high safety" still an un-met need. To this aim, a cell-penetrating peptide, CPP, which efficiently transported cargos into epithelial cells was exanimated. CPP was evaluated with cell model and 3D skin model following OECD guidelines without using animal models. As a highly stable peptide, CPP neither irritated nor sensitized skin, also did not disrupt skin barrier. In addition, such high safety peptide had anti-inflammation activity without allergic effect. Moreover, cargo carrying activity of CPP was assayed using HaCaT cell model and rapid CPP penetration was observed within 30 min. Finally, CPP possessed transepidermal activity in water in oil formulation without disruption of skin barrier. All evidences indicated that CPP was an ideal choice for skin penetrating and its anti-inflammatory activity could improve skin condition, which made CPP suitable and attractive for novel cosmeceutical product development.
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http://dx.doi.org/10.3390/biom10010101DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7023394PMC
January 2020

Comparing progression molecular mechanisms between lung adenocarcinoma and lung squamous cell carcinoma based on genetic and epigenetic networks: big data mining and genome-wide systems identification.

Oncotarget 2019 Jun 4;10(38):3760-3806. Epub 2019 Jun 4.

Laboratory of Automatic Control, Signaling Processing, and Systems Biology, Department of Electrical Engineering, National Tsing Hua University, Hsinchu 30013, Taiwan.

Non-small-cell lung cancer (NSCLC) is the predominant type of lung cancer in the world. Lung adenocarcinoma (LADC) and lung squamous cell carcinoma (LSCC) are subtypes of NSCLC. We usually regard them as different disease due to their unique molecular characteristics, distinct cells of origin and dissimilar clinical response. However, the differences of genetic and epigenetic progression mechanism between LADC and LSCC are complicated to analyze. Therefore, we applied systems biology approaches and big databases mining to construct genetic and epigenetic networks (GENs) with next-generation sequencing data of LADC and LSCC. In order to obtain the real GENs, system identification and system order detection are conducted on gene regulatory networks (GRNs) and protein-protein interaction networks (PPINs) for each stage of LADC and LSCC. The core GENs were extracted via principal network projection (PNP). Based on the ranking of projection values, we got the core pathways in respect of KEGG pathway. Compared with the core pathways, we found significant differences between microenvironments, dysregulations of miRNAs, epigenetic modifications on certain signaling transduction proteins and target genes in each stage of LADC and LSCC. Finally, we proposed six genetic and epigenetic multiple-molecule drugs to target essential biomarkers in each progression stage of LADC and LSCC, respectively.
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http://dx.doi.org/10.18632/oncotarget.26940DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6557199PMC
June 2019

Investigation mechanisms between normal, developing and regenerating livers for regenerative liver drug design.

Regen Med 2019 05 17;14(5):359-387. Epub 2019 Jun 17.

Lab of Automatic Control, Signal Processing, and Systems Biology, Department of Electrical Engineering, National Tsing Hua University, Hsinchu 30013, Taiwan.

A systematic multimolecule drug design procedure is proposed for promoting hepatogenesis and liver regeneration. Genome-wide microarray data including three hepatic conditions are obtained from the GEO database (GSE15238). System modeling and big data mining methods are used to construct real genome-wide genetic-and-epigenetic networks (GWGENs). Then, we extracted the core GWGENs by applying principal network projection on real GWGENs of normal, developing and regenerating livers, respectively. After that, we investigated the significant signal pathways and epigenetic modifications in the core GWGENs to identify potential biomarkers as drug targets. A multimolecule drug consisting of sulmazole, clofibrate, colchicine, furazolidone, nadolol, eticlopride and felbinac is proposed to target on novel biomarkers for promoting hepatogenesis and liver regeneration.
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http://dx.doi.org/10.2217/rme-2018-0058DOI Listing
May 2019

Intrahepatic hepatitis B virus large surface antigen induces hepatocyte hyperploidy via failure of cytokinesis.

J Pathol 2018 08 4;245(4):502-513. Epub 2018 Jul 4.

Institute of Molecular and Cellular Biology, National Tsing Hua University, Hsinchu, Taiwan.

Hepatitis B virus (HBV) is an aetiological factor for liver cirrhosis and hepatocellular carcinoma (HCC). Despite current antiviral therapies that successfully reduce the viral load in patients with chronic hepatitis B, persistent hepatitis B surface antigen (HBsAg) remains a risk factor for HCC. To explore whether intrahepatic viral antigens contribute directly to hepatocarcinogenesis, we monitored the mitotic progression of HBV-positive cells. Cytokinesis failure was increased in HBV-positive HepG2.2.15 and 1.3ES2 cells, as well as in HuH-7 cells transfected with a wild-type or X-deficient HBV construct, but not in cells transfected with an HBsAg-deficient construct. We show that expression of viral large surface antigen (LHBS) was sufficient to induce cytokinesis failure of immortalized hepatocytes. Premitotic defects with DNA damage and G /M checkpoint attenuation preceded cytokinesis in LHBS-positive cells, and ultimately resulted in hyperploidy. Inhibition of polo-like kinase-1 (Plk1) not only restored the G /M checkpoint in these cells, but also suppressed LHBS-mediated in vivo tumourigenesis. Finally, a positive correlation between intrahepatic LHBS expression and hepatocyte hyperploidy was detected in >70% of patients with chronic hepatitis B. We conclude that HBV LHBS provokes hyperploidy by inducing DNA damage and upregulation of Plk1; the former results in atypical chromatin structures, and the latter attenuates the function of the G /M DNA damage checkpoint. Our data uncover a mechanism by which genomic integrity of hepatocytes is disrupted by viral LHBS. These findings highlight the role of intrahepatic surface antigen as an oncogenic risk factor in the development of HCC. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
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http://dx.doi.org/10.1002/path.5102DOI Listing
August 2018

Heparin-Promoted Cellular Uptake of the Cell-Penetrating Glycosaminoglycan Binding Peptide, GBP, Depends on a Single Tryptophan.

ACS Chem Biol 2017 02 21;12(2):398-406. Epub 2016 Dec 21.

Institute of Bioinformatics and Structural Biology, National Tsing Hua University , Hsinchu 30013, Taiwan.

A 10-residue, glycosaminoglycan-binding peptide, GBP, derived from human eosinophil cationic protein has been recently designated as a potent cell-penetrating peptide. A model system containing peptide, glycan, and lipid was monitored by nuclear magnetic resonance (NMR) spectroscopy to determine the cell-penetrating mechanism. Heparin octasaccharide with dodecylphosphocholine (DPC) lipid micelle was titrated into the GBP solution. Our data revealed substantial roles for the charged residues Arg5 and Lys7 in recognizing heparin, whereas Arg3 had less effect. The aromatic residue Trp4 acted as an irreplaceable moiety for membrane insertion, as the replacement of Trp4 with Arg4 abolished cell penetration, although it significantly improved the heparin-binding ability. GBP bound either heparin or lipid in the presence or absence of the other ligand indicating that the peptide has two alternative binding sites: Trp4 is responsible for lipid insertion, and Arg5 and Lys7 are for GAG binding. We developed a molecular model showing that the two effects synergistically promote the penetration. The loss of either effect would abolish the penetration. GBP has been proven to enter cells through macropinocytosis. The GBP treatment inhibited A549 lung cancer cell migration and invasion, implying that the cellular microenvironment would be modulated by GBP internalization. The intracellular penetration of GBP leading to inhibition of epithelial cell migration and invasion depends on the presence of the tryptophan residue in its sequence compared with similar derivative peptides. Therefore, GBP shows substantial potential as a novel delivery therapeutic through rapid and effective internalization and interference with cell mobility.
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http://dx.doi.org/10.1021/acschembio.6b00864DOI Listing
February 2017

Hepatitis B virus PreS2-mutant large surface antigen activates store-operated calcium entry and promotes chromosome instability.

Oncotarget 2016 Apr;7(17):23346-60

Institute of Molecular and Cellular Biology, National Tsing Hua University, Hsinchu 300, Taiwan.

Hepatitis B virus (HBV) is a driver of hepatocellular carcinoma, and two viral products, X and large surface antigen (LHBS), are viral oncoproteins. During chronic viral infection, immune-escape mutants on the preS2 region of LHBS (preS2-LHBS) are gain-of-function mutations that are linked to preneoplastic ground glass hepatocytes (GGHs) and early disease onset of hepatocellular carcinoma. Here, we show that preS2-LHBS provoked calcium release from the endoplasmic reticulum (ER) and triggered stored-operated calcium entry (SOCE). The activation of SOCE increased ER and plasma membrane (PM) connections, which was linked by ER- resident stromal interaction molecule-1 (STIM1) protein and PM-resident calcium release- activated calcium modulator 1 (Orai1). Persistent activation of SOCE induced centrosome overduplication, aberrant multipolar division, chromosome aneuploidy, anchorage-independent growth, and xenograft tumorigenesis in hepatocytes expressing preS2- LHBS. Chemical inhibitions of SOCE machinery and silencing of STIM1 significantly reduced centrosome numbers, multipolar division, and xenograft tumorigenesis induced by preS2-LHBS. These results provide the first mechanistic link between calcium homeostasis and chromosome instability in hepatocytes carrying preS2-LHBS. Therefore, persistent activation of SOCE represents a novel pathological mechanism in HBV-mediated hepatocarcinogenesis.
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http://dx.doi.org/10.18632/oncotarget.8109DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5029631PMC
April 2016

PICH promotes sister chromatid disjunction and co-operates with topoisomerase II in mitosis.

Nat Commun 2015 Dec 8;6:8962. Epub 2015 Dec 8.

Center for Chromosome Stability and Center for Healthy Aging, Department of Cellular and Molecular Medicine, University of Copenhagen, DK-2200 Copenhagen, Denmark.

PICH is a SNF2 family DNA translocase that binds to ultra-fine DNA bridges (UFBs) in mitosis. Numerous roles for PICH have been proposed from protein depletion experiments, but a consensus has failed to emerge. Here, we report that deletion of PICH in avian cells causes chromosome structural abnormalities, and hypersensitivity to an inhibitor of Topoisomerase II (Topo II), ICRF-193. ICRF-193-treated PICH(-/-) cells undergo sister chromatid non-disjunction in anaphase, and frequently abort cytokinesis. PICH co-localizes with Topo IIα on UFBs and at the ribosomal DNA locus, and the timely resolution of both structures depends on the ATPase activity of PICH. Purified PICH protein strongly stimulates the catalytic activity of Topo II in vitro. Consistent with this, a human PICH(-/-) cell line exhibits chromosome instability and chromosome condensation and decatenation defects similar to those of ICRF-193-treated cells. We propose that PICH and Topo II cooperate to prevent chromosome missegregation events in mitosis.
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http://dx.doi.org/10.1038/ncomms9962DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4686863PMC
December 2015

The photothermal effect of silica-carbon hollow sphere-concanavalin A on liver cancer cells.

J Mater Chem B 2015 Mar 16;3(12):2447-2454. Epub 2015 Feb 16.

Department of Biomedical Engineering, National Cheng Kung University, Tainan 70101, Taiwan.

Hepatocellular carcinoma (HCC) is one of the most common cancers and causes of death by cancer. Concanavalin A (ConA) lectin can specifically bind to the glycoprotein receptors of HCC, which are produced by the aberrant overexpression of liver cancer cells. ConA was used in the current study to conjugate on silica-carbon hollow spheres (SCHSs) and applied in the thermal ablation therapy of liver cancer cell lines under near-infrared (NIR) laser irradiation. We found that the amount of ConA-SCHS complex binding to hepatoma cells was significantly higher than that seen with normal hepatocytes, based on flow cytometric analysis and confocal imaging. Hepatoma cells incubated with ConA-SCHSs were thus more easily killed by the subsequent irradiation with a NIR laser. The results show that the ConA-SCHS complex may enhance the interaction with highly expressed ConA receptors on hepatoma cells, and thus serve as an effective photothermal therapy agent for liver cancer treatment.
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http://dx.doi.org/10.1039/c5tb00056dDOI Listing
March 2015

Hepatitis B virus pre-S2 mutant large surface protein inhibits DNA double-strand break repair and leads to genome instability in hepatocarcinogenesis.

J Pathol 2015 Jul 22;236(3):337-47. Epub 2015 Apr 22.

Department of Medical Laboratory Science and Biotechnology, College of Medicine, National Cheng Kung University, Tainan, Taiwan.

Although hepatitis B virus (HBV) has been established to cause hepatocellular carcinoma (HCC), the exact mechanism remains to be clarified. Type II ground glass hepatocytes (GGHs) harbouring the HBV pre-S2 mutant large surface protein (LHBS) have been recognized as a morphologically distinct hallmark of HCC in the advanced stages of chronic HBV infection. Considering its preneoplastic nature, we hypothesized that type II GGH may exhibit high genomic instability, which is important for the carcinogenic process in chronic HBV carriers. In this study we found that pre-S2 mutant LHBS directly interacted with importin α1, the key factor that recognizes cargos undergoing nuclear transportation mediated by the importin α/β-associated nuclear pore complex (NPC). By interacting with importin α1, which inhibits its function as an NPC factor, pre-S2 mutant LHBS blocked nuclear transport of an essential DNA repair and recombination factor, Nijmegen breakage syndrome 1 (NBS1), upon DNA damage, thereby delaying the formation of nuclear foci at the sites of DNA double-strand breaks (DSBs). Pre-S2 mutant LHBS was also found to block NBS1-mediated homologous recombination repair and induce multi-nucleation of cells. In addition, pre-S2 mutant LHBS transgenic mice showed genomic instability, indicated by increased global gene copy number variations (CNVs), which were significantly higher than those in hepatitis B virus X mice, indicating that pre-S2 mutant LHBS is the major viral oncoprotein inducing genomic instability in HBV-infected hepatocytes. Consistently, the human type II GGHs in HCC patients exhibited increased DNA DSBs representing significant genomic instability. In conclusion, type II GGHs harbouring HBV pre-S2 mutant oncoprotein represent a high-risk marker for the loss of genome integrity in chronic HBV carriers and explain the complex chromosome changes in HCCs. Mouse array CGH raw data: GEO Accession No. GSE61378 (http://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE61378).
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http://dx.doi.org/10.1002/path.4531DOI Listing
July 2015

Erratum to: PGRMC1 contributes to doxorubicin-induced chemoresistance in MES-SA uterine sarcoma.

Cell Mol Life Sci 2015 Jun;72(12):2411-3

Institute of Bioinformatics and Structural Biology, National Tsing Hua University, No. 101, Kuang-Fu Rd. Sec. 2, Hsin-chu, 30013, Taiwan.

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http://dx.doi.org/10.1007/s00018-015-1852-zDOI Listing
June 2015

Sgo1 is a potential therapeutic target for hepatocellular carcinoma.

Oncotarget 2015 Feb;6(4):2023-33

Institute of Molecular and Cellular Biology, National Tsing Hua University, Hsinchu, Taiwan.

Shugoshin-like protein 1 (Sgo1) is an essential protein in mitosis; it protects sister chromatid cohesion and thereby ensures the fidelity of chromosome separation. We found that the expression of Sgo1 mRNA was relatively low in normal tissues, but was upregulated in 82% of hepatocellular carcinoma (HCC), and correlated with elevated alpha-fetoprotein and early disease onset of HCC. The depletion of Sgo1 reduced cell viability of hepatoma cell lines including HuH7, HepG2, Hep3B, and HepaRG. Using time-lapse microscopy, we showed that hepatoma cells were delayed and ultimately die in mitosis in the absence of Sgo1. In contrast, cell viability and mitotic progression of immortalized cells were not significantly affected. Notably, mitotic cell death induced upon Sgo1 depletion was suppressed upon inhibitions of cyclin-dependent kinase-1 and Aurora kinase-B, or the depletion of mitotic arrest deficient-2. Thus, mitotic cell death induced upon Sgo1 depletion in hepatoma cells is mediated by persistent activation of the spindle assembly checkpoint. Together, these results highlight the essential role of Sgo1 in the maintenance of a proper mitotic progression in hepatoma cells and suggest that Sgo1 is a promising oncotarget for HCC.
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http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4385833PMC
http://dx.doi.org/10.18632/oncotarget.2764DOI Listing
February 2015

PGRMC1 contributes to doxorubicin-induced chemoresistance in MES-SA uterine sarcoma.

Cell Mol Life Sci 2015 Jun 18;72(12):2395-409. Epub 2015 Jan 18.

Institute of Bioinformatics and Structural Biology, National Tsing Hua University, No. 101, Kuang-Fu Rd. Sec. 2, Hsin-chu, 30013, Taiwan.

Chemotherapy is one of the major categories of medical oncology and a primary tumor treatment; however, the effectiveness of chemotherapy is restricted by drug resistance. Overcoming resistance to chemotherapy and investigating molecular targeted therapies are challenges currently faced during resistance management. Progesterone receptor membrane component 1 (PGRMC1) is an adapter protein mediating cholesterol synthesis, steroid signaling, and cytochrome p450 activation. Attention has recently focused on the role of PGRMC1 in cell survival, anti-apoptosis, and damage response. In the present study, we used knockdown and overexpression approaches in the following set of uterine sarcoma models to further evaluate the role of PGRMC1 in drug resistance: the doxorubicin-sensitive MES-SA cells and the doxorubicin-resistant MES-SA/DxR-2 µM and MES-SA/DxR-8 µM cells (with different levels of doxorubicin resistance). PGRMC1 repressed doxorubicin-induced cytotoxicity and exhibited an anti-apoptotic effect; it also promoted cell proliferation and cell cycle progression to the S phase. Of note, PGRMC1 overexpression led to the epithelial-mesenchymal transition (EMT) of the sensitive MES-SA cells, thus facilitating their migration and invasion. The combination of PGRMC1 knockdown and the P-glycoprotein inhibitor verapamil significantly decreased the viability of P-glycoprotein-overexpressing MES-SA/DxR-8 μM cells after doxorubicin treatment. Taken together, our results show that PGRMC1 contributed to chemoresistance through cell proliferation, anti-apoptosis, and EMT induction, leading to the suggestion that PGRMC1 may serve as a therapeutic target in combination with an inhibitor in different drug resistance pathways and indicating the usefulness of predictive resistance biomarkers in uterine sarcoma.
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http://dx.doi.org/10.1007/s00018-014-1831-9DOI Listing
June 2015

SH2B1 and IRSp53 proteins promote the formation of dendrites and dendritic branches.

J Biol Chem 2015 Mar 13;290(10):6010-21. Epub 2015 Jan 13.

From the Institute of Molecular Medicine, Department of Medical Science, National Tsing Hua University, Hsinchu, Taiwan 30013, China

SH2B1 is an adaptor protein known to enhance neurite outgrowth. In this study, we provide evidence suggesting that the SH2B1 level is increased during in vitro culture of hippocampal neurons, and the β isoform (SH2B1β) is the predominant isoform. The fact that formation of filopodia is prerequisite for neurite initiation suggests that SH2B1 may regulate filopodium formation and thus neurite initiation. To investigate whether SH2B1 may regulate filopodium formation, the effect of SH2B1 and a membrane and actin regulator, IRSp53 (insulin receptor tyrosine kinase substrate p53), is investigated. Overexpressing both SH2B1β and IRSp53 significantly enhances filopodium formation, neurite outgrowth, and branching. Both in vivo and in vitro data show that SH2B1 interacts with IRSp53 in hippocampal neurons. This interaction depends on the N-terminal proline-rich domains of SH2B1. In addition, SH2B1 and IRSp53 co-localize at the plasma membrane, and their levels increase in the Triton X-100-insoluble fraction of developing neurons. These findings suggest that SH2B1-IRSp53 complexes promote the formation of filopodia, neurite initiation, and neuronal branching.
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http://dx.doi.org/10.1074/jbc.M114.603795DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4358244PMC
March 2015

The emerging role of hepatitis B virus pre-S2 deletion mutant proteins in HBV tumorigenesis.

J Biomed Sci 2014 Oct 15;21:98. Epub 2014 Oct 15.

National Institute of Infectious Diseases and Vaccinology, National Health Research Institutes, Tainan, Taiwan.

Chronic hepatitis B virus (HBV) infection can cause hepatocellular carcinoma (HCC). Several hypotheses have been proposed to explain the mechanisms of HBV tumorigenesis, including inflammation and liver regeneration associated with cytotoxic immune injuries and transcriptional activators of mutant HBV gene products. The mutant viral oncoprotein-driven tumorigenesis is prevailed at the advanced stage or anti-HBe-positive phase of chronic HBV infection. Besides HBx, the pre-S2 (deletion) mutant protein represents a newly recognized oncoprotein that is accumulated in the endoplasmic reticulum (ER) and manifests as type II ground glass hepatocytes (GGH). The retention of pre-S2 mutant protein in ER can induce ER stress and initiate an ER stress-dependent VEGF/Akt/mTOR and NFκB/COX-2 signal pathway. Additionally, the pre-S2 mutant large surface protein can induce an ER stress-independent pathway to transactivate JAB-1/p27/RB/cyclin A,D pathway, leading to growth advantage of type II GGH. The pre-S2 mutant protein-induced ER stress can also cause DNA damage, centrosome overduplication, and genomic instability. In 5-10% of type II GGHs, there is co-expression of pre-S2 mutant protein and HBx antigen which exhibited enhanced oncogenic effects in transgenic mice. The mTOR signal cascade is consistently activated throughout the course of pre-S2 mutant transgenic livers and in human HCC tissues, leading to metabolic disorders and HCC tumorigenesis. Clinically, the presence of pre-S2 deletion mutants in sera frequently develop resistance to nucleoside analogues anti-virals and predict HCC development. The pre-S2 deletion mutants and type II GGHs therefore represent novel biomarkers of HBV-related HCCs. A versatile DNA array chip has been developed to detect pre-S2 mutants in serum. Overall, the presence of pre-S2 mutants in serum has implications for anti-viral treatment and can predict HCC development. Targeting at pre-S2 mutant protein-induced, ER stress-dependent, mTOR signal cascade and metabolic disorders may offer potential strategy for chemoprevention or therapy in high risk chronic HBV carriers.
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http://dx.doi.org/10.1186/s12929-014-0098-7DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4200140PMC
October 2014

Cholesterol glucosylation by Helicobacter pylori delays internalization and arrests phagosome maturation in macrophages.

J Microbiol Immunol Infect 2016 Oct 26;49(5):636-645. Epub 2014 Jul 26.

Institute of Molecular and Cellular Biology and Department of Life Science, National Tsing Hua University, Hsinchu, Taiwan; Biomedical Science and Engineering Center, National Tsing Hua University, Hsinchu, Taiwan. Electronic address:

Background/purpose: Helicobacter pylori colonizes the human stomach and contributes to chronic inflammation of the gastric mucosa. H. pylori persistence occurs because of insufficient eradication by phagocytic cells. A key factor of H. pylori, cholesterol-α-glucosyltransferase encoded by capJ that extracts host cholesterol and converts it to cholesteryl glucosides, is important to evade host immunity. Here, we examined whether phagocytic trafficking in macrophages was perturbed by capJ-carrying H. pylori.

Methods: J774A.1 cells were infected with H. pylori at a multiplicity of infection of 50. Live-cell imaging and confocal microscopic analysis were applied to monitor the phagocytic trafficking events. The viability of H. pylori inside macrophages was determined by using gentamicin colony-forming unit assay. The phagocytic routes were characterized by using trafficking-intervention compounds.

Results: Wild type (WT) H. pylori exhibited more delayed entry into macrophages and also arrested phagosome maturation more than did capJ knockout mutant. Pretreatment of genistein and LY294002 prior to H. pylori infection reduced the internalization of WT but not capJ-knockout H. pylori in macrophages.

Conclusion: Cholesterol glucosylation by H. pylori interferes with phagosome trafficking via a lipid-raft and PI3K-dependent manner, which retards engulfment of bacteria for prolonged intracellular survival of H. pylori.
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http://dx.doi.org/10.1016/j.jmii.2014.05.011DOI Listing
October 2016

Ground-glass hepatocytes co-expressing hepatitis B virus X protein and surface antigens exhibit enhanced oncogenic effects and tumorigenesis.

Hum Pathol 2014 Jun 28;45(6):1294-301. Epub 2014 Feb 28.

National Institute of Infectious Diseases and Vaccinology, National Health Research Institutes, Tainan 70456, Taiwan; Department of Pathology, National Cheng Kung University Hospital, Tainan 70403, Taiwan; Department of Surgery, National Cheng Kung University Hospital, Tainan 70403, Taiwan. Electronic address:

Hepatitis B virus (HBV) X protein (HBx) and pre-S2 deletion mutant large surface antigens are oncoproteins that induce hepatocellular carcinoma (HCC). The interaction of these two oncoproteins in hepatocytes and its significance in tumorigenesis remain to be elucidated. In this study, we observed the co-expression of HBx with surface antigens in ground-glass hepatocytes in 5 of 20 hepatitis B surface antigen-positive livers. In vitro, hepatocytes co-expressing HBx and a pre-S2 mutant showed enhanced expression of vascular endothelial growth factor-A, phosphorylated Akt 1/2/3, phosphorylated extracellular signal-regulated kinase 1/2, and phosphorylated mammalian target of rapamycin signals. Transgenic mice harboring both HBx and pre-S2 mutant construct plasmids developed HCCs at an average of 15.1 months, earlier than animals carrying either HBx (16.9 months) or pre-S2 mutant (24.5 months) alone. The oncogenic signals of vascular endothelial growth factor-A, phosphorylated Akt 1/2/3, phosphorylated extracellular signal-regulated kinase 1/2, and phosphorylated mammalian target of rapamycin were sequentially and differentially activated at different stages in tumorigenesis. Phosphorylated mTOR was consistently activated in transgenic and human HCCs. We conclude that ground-glass hepatocytes co-expressing HBx and surface antigens exhibit enhanced oncogenic effects and tumorigenesis in chronic HBV infections. The mTOR signal cascade may be the key regulator in HBV tumorigenesis and may be useful targets in the design of HCC therapy.
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http://dx.doi.org/10.1016/j.humpath.2013.10.039DOI Listing
June 2014

Pathogenesis of virus-associated human cancers: Epstein-Barr virus and hepatitis B virus as two examples.

J Formos Med Assoc 2014 Sep 3;113(9):581-90. Epub 2013 Oct 3.

Department of Pathology, College of Medicine, National Cheng Kung University and Hospital, Tainan, Taiwan; National Institute of Infectious Diseases and Vaccinology, National Health Research Institute, Tainan, Taiwan. Electronic address:

Virus-associated human cancers may exhibit two characteristic histopathologic features: (1) the inflammation-rich background as observed in Epstein-Barr virus-associated Hodgkin lymphoma (HL) and nasopharyngeal carcinoma (NPC); and (2) the characteristic nuclear morphology such as the Reed-Sternberg cells in HL. Besides, the hepatocytes of chronic hepatitis B virus (HBV) infection frequently exhibit characteristic ground glass hepatocytes, a phenomenon associated with endoplasmic reticulum stress response induced by the overloaded or malfolded HBV surface antigens. In this review, we explore specifically the pathogenesis of Epstein-Barr virus-associated HL and NPC, and HBV-associated hepatocellular carcinoma based on the observed histopathologic features. We propose that the retention of viral proteins induces inflammation, endoplasmic reticulum stress, and genomic instability in HL, NPC, and hepatocellular carcinoma, whereby the viral oncoproteins may play additional transactivational roles to induce host genes for transformation, invasion, and metastasis. Therapeutic implications based on the pathogenesis of virus-associated cancers are discussed.
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http://dx.doi.org/10.1016/j.jfma.2013.09.001DOI Listing
September 2014

Microtubule depolymerization activates the Epstein-Barr virus lytic cycle through protein kinase C pathways in nasopharyngeal carcinoma cells.

J Gen Virol 2013 Dec 23;94(Pt 12):2750-2758. Epub 2013 Sep 23.

Department of Medical Science, National Tsing Hua University, Hsinchu 300, Taiwan.

Elevated levels of antibodies against Epstein-Barr virus (EBV) and the presence of viral DNA in plasma are reliable biomarkers for the diagnosis of nasopharyngeal carcinoma (NPC) in high-prevalence areas, such as South-East Asia. The presence of these viral markers in the circulation suggests that a minimal level of virus reactivation may have occurred in an infected individual, although the underlying mechanism of reactivation remains to be elucidated. Here, we showed that treatment with nocodazole, which provokes the depolymerization of microtubules, induces the expression of two EBV lytic cycle proteins, Zta and EA-D, in EBV-positive NPC cells. This effect was independent of mitotic arrest, as viral reactivation was not abolished in cells synchronized at interphase. Notably, the induction of Zta by nocodazole was mediated by transcriptional upregulation via protein kinase C (PKC). Pre-treatment with inhibitors for PKC or its downstream signalling partners p38 mitogen-activated protein kinase (MAPK) and c-Jun N-terminal kinase (JNK) abolished the nocodazole-mediated induction of Zta and EA-D. Interestingly, the effect of nocodazole, as well as colchicine and vinblastine, on lytic gene expression occurred only in NPC epithelial cells but not in cells derived from lymphocytes. These results establish a novel role of microtubule integrity in controlling the EBV life cycle through PKC and its downstream pathways, which represents a tissue-specific mechanism for controlling the life-cycle switch of EBV.
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http://dx.doi.org/10.1099/vir.0.058040-0DOI Listing
December 2013

Hepatic expression of HCV RNA-dependent RNA polymerase triggers innate immune signaling and cytokine production.

Mol Cell 2012 Oct 6;48(2):313-21. Epub 2012 Sep 6.

Laboratory of Gene Regulation and Signal Transduction, Departments of Pharmacology and Pathology, School of Medicine, University of California, San Diego, La Jolla, CA 92093-0723, USA.

Innate immunity controls pathogen replication and spread. Yet, certain pathogens, such as Hepatitis C Virus (HCV), escape immune elimination and establish persistent infections that promote chronic inflammation and related diseases. Whereas HCV regulatory proteins that attenuate antiviral responses are known, those that promote inflammation and liver injury remain to be identified. Here, we show that transient expression of HCV RNA-dependent RNA polymerase (RdRp), NS5B, in mouse liver and human hepatocytes results in production of small RNA species that activate innate immune signaling via TBK1-IRF3 and NF-κB and induce cytokine production, including type I interferons (IFN) and IL-6. NS5B-expression also results in liver damage.
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http://dx.doi.org/10.1016/j.molcel.2012.07.032DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3483424PMC
October 2012

The involvement of MCT-1 oncoprotein in inducing mitotic catastrophe and nuclear abnormalities.

Cell Cycle 2012 Mar 1;11(5):934-52. Epub 2012 Mar 1.

Institute of Molecular and Genomic Medicine, National Health Research Institutes, Miaoli, Taiwan.

Centrosome amplification and chromosome abnormality are frequently identified in neoplasia and tumorigenesis. However, the mechanisms underlying these defects remain unclear. We here identify that MCT-1 is a centrosomal oncoprotein involved in mitosis. Knockdown of MCT-1 protein results in intercellular bridging, chromosome mis-congregation, cytokinesis delay, and mitotic death. Introduction of MCT-1 oncogene into the p53 deficient cells (MCT-1-p53), the mitotic checkpoint kinases and proteins are deregulated synergistically. These biochemical alterations are accompanied with increased frequencies of cytokinesis failure, multi-nucleation, and centrosome amplification in subsequent cell cycle. As a result, the incidences of polyploidy and aneuploidy are progressively induced by prolonged cell cultivation or further promoted by sustained spindle damage on MCT-1-p53 background. These data show that the oncoprotein perturbs centrosome structure and mitotic progression, which provide the molecular aspect of chromsomal abnormality in vitro and the information for understanding the stepwise progression of tumors under oncogenic stress.
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http://dx.doi.org/10.4161/cc.11.5.19452DOI Listing
March 2012

Induction of Bcl-2 expression by hepatitis B virus pre-S2 mutant large surface protein resistance to 5-fluorouracil treatment in Huh-7 cells.

PLoS One 2011 22;6(12):e28977. Epub 2011 Dec 22.

Department of Biotechnology, Chia Nan University of Pharmacy and Science, Tainan, Taiwan.

Background: Hepatocellular carcinoma (HCC) is one of the most common malignancies worldwide with poor prognosis due to resistance to conventional chemotherapy and limited efficacy of radiotherapy. Our previous studies have indicated that expression of Hepatitis B virus pre-S2 large mutant surface antigen (HBV pre-S2Δ) is associated with a significant risk of developing HCC. However, the relationship between HBV pre-S2Δ protein and the resistance of chemotherapeutic drug treatment is still unclear.

Methodology/principal Findings: Here, we show that the expression of HBV pre-S2Δ mutant surface protein in Huh-7 cell significantly promoted cell growth and colony formation. Furthermore, HBV pre-S2Δ protein increased both mRNA (2.7±0.5-fold vs. vehicle, p=0.05) and protein (3.2±0.3-fold vs. vehicle, p=0.01) levels of Bcl-2 in Huh-7 cells. HBV pre-S2Δ protein also enhances Bcl-2 family, Bcl-xL and Mcl-1, expression in Huh-7 cells. Meanwhile, induction of NF-κB p65, ERK, and Akt phosphorylation, and GRP78 expression, an unfolded protein response chaperone, were observed in HBV pre-S2Δ and HBV pre-S-expressing cells. Induction of Bcl-2 expression by HBV pre-S2Δ protein resulted in resistance to 5-fluorouracil treatment in colony formation, caspase-3 assay, and cell apoptosis, and can enhance cell death by co-incubation with Bcl-2 inhibitor. Similarly, transgenic mice showed higher expression of Bcl-2 in liver tissue expressing HBV pre-S2Δ large surface protein in vivo.

Conclusion/significance: Our result demonstrates that HBV pre-S2Δ increased Bcl-2 expression which plays an important role in resistance to 5-fluorouracil-caused cell death. Therefore, these data provide an important chemotherapeutic strategy in HBV pre-S2Δ-associated tumor.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0028977PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3245229PMC
May 2012

Aberrant cyclin A expression and centrosome overduplication induced by hepatitis B virus pre-S2 mutants and its implication in hepatocarcinogenesis.

Carcinogenesis 2012 Feb 9;33(2):466-72. Epub 2011 Dec 9.

Institute of Molecular and Cellular Biology, National Tsing Hua University, Hsinchu 300, Taiwan.

Ground glass hepatocytes harboring hepatitis B virus (HBV) pre-S2 mutants have been recognized as pre-neoplastic lesions of hepatocellular carcinoma (HCC). The pre-S2 mutants accumulated in endoplasmic reticulum (ER) can induce ER stress, upregulate cyclin A and promote hepatocyte proliferation. Notably, cyclin A was aberrantly detected in the cytoplasm, instead of nucleus, of pre-S2 mutant-transgenic mice livers, thereby raising the potential role of cytoplasmic cyclin A in HBV hepatocarcinogenesis. In this study, we confirmed that cyclin A was detected in the cytoplasm in the majority of HBV-related HCC tissues. In vitro, the pre-S2 mutant-initiated ER stress could induce cytoplasmic cyclin A mediated via cleavage by the calcium-dependent protease μ-calpain, resulting in an N-terminal truncated product which was preferentially located in the cytoplasm. The aberrant cyclin A expression subsequently induced centrosome overduplication, and this effect was abolished by calpain-specific inhibitors or RNA interference targeting to cyclin A. Overall, our data indicate that HBV pre-S2 mutant may elicit aberrant cyclin A expression and centrosome overduplication through ER stress induction and thereby represent a potential mechanism for the chromosome instability in HBV hepatocarcinogenesis.
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http://dx.doi.org/10.1093/carcin/bgr296DOI Listing
February 2012

Centromere DNA decatenation depends on cohesin removal and is required for mammalian cell division.

J Cell Sci 2010 Mar 9;123(Pt 5):806-13. Epub 2010 Feb 9.

Department of Cell Biology, Max-Planck Institute of Biochemistry, D-82152 Martinsried, Germany.

Sister chromatid cohesion is mediated by DNA catenation and proteinaceous cohesin complexes. The recent visualization of PICH (Plk1-interacting checkpoint helicase)-coated DNA threads in anaphase cells raises new questions as to the role of DNA catenation and its regulation in time and space. In the present study we show that persistent DNA catenation induced by inhibition of Topoisomerase-IIalpha can contribute to sister chromatid cohesion in the absence of cohesin complexes and that resolution of catenation is essential for abscission. Furthermore, we use an in vitro chromatid separation assay to investigate the temporal and functional relationship between cohesin removal and Topoisomerase-IIalpha-mediated decatenation. Our data suggest that centromere decatenation can occur only after separase activation and cohesin removal, providing a plausible explanation for the persistence of centromere threads after anaphase onset.
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http://dx.doi.org/10.1242/jcs.058255DOI Listing
March 2010