HBV enveloped particle secretion is positively regulated by GRP78 through direct interaction with preS1

Hepatitis B virus (HBV) infection is a common cause of liver diseases worldwide. Existing drugs do not effectively eliminate HBV from infected hepatocytes; thus, novel curative therapies are needed. Enveloped-particle secretion is a key but poorly studied aspect of the viral life cycle. Here, we report that GRP78 positively regulates HBV enveloped-particle secretion. GRP78 is the specific target of preS1 binding; HBV can upregulate GRP78 in liver cell lines and sera from chronic hepatitis B patients. GRP78 promoted intact HBV-particle secretion in liver cell lines and an HBV transgenic-mouse model. Some peptides screened from preS1 via phage display could inhibit viral-particle secretion by interacting with GRP78 via hydrogen bonds and hydrophobic interactions, thereby disturbing the interaction with HBV particles. These results provide insight into enveloped-particle secretion in the HBV life cycle. GRP78 might be a potential target for HBV-infection treatment via restricting GRP78–preS1 interactions to block viral-particle secretion.

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Hepatitis B Virus Replication Induces Methylation of both Host and Viral DNA

Journal of Virology ◽

10.1128/jvi.02280-09 ◽

2010 ◽

Vol 84 (9) ◽

pp. 4321-4329 ◽

Cited By ~ 78

Author(s):

Perumal Vivekanandan ◽

Hubert Darius-J Daniel ◽

Rajesh Kannangai ◽

Francisco Martinez-Murillo ◽

Michael Torbenson

Keyword(s):

Gene Expression ◽

Hepatitis B Virus ◽

Hepatitis B ◽

Viral Replication ◽

Cell Lines ◽

Natural Infection ◽

Cpg Islands ◽

Hbv Infection ◽

Viral Dna ◽

B Virus

ABSTRACT Control of viral replication is a major therapeutic goal to reduce morbidity and mortality from chronic hepatitis B virus (HBV) infection. Recently, methylation has been identified as a novel host defense mechanism, and methylation of viral DNA leads to downregulation of HBV gene expression. To better understand the mechanisms of HBV methylation, cell lines were exposed to HBV using a model system that mimics natural infection and the expression of host DNA methyltransferase genes (DNMTs) was measured. DNMT1, DNMT2, and DNMT3 were all significantly upregulated in response to HBV. DNMT3 was further studied because of its known role in the de novo methylation of DNA. Cotransfection experiments with full-length HBV and DNMT3 led to the downregulation of viral protein and pregenomic RNA production. To investigate whether the upregulation of DNMTs could also have an effect on the methylation of host DNA, cell lines were exposed to HBV in two independent model systems, one that mimics natural infection and a second model with temporary transfection. Host DNA methylation was measured by DNA microarray analysis. Increased methylation of host CpG islands was detected in both experimental systems. Two CpG islands, corresponding to genes SUFU and TIRAP, were selected, and the downregulation of these genes in hepatocellular carcinomas was confirmed. In conclusion, hepatocytes respond to HBV infection by upregulating DNMTs. The DNMTs methylate viral DNA, leading to decreased viral gene expression and decreased viral replication. However, virus-induced overexpression of DNMTs also leads to methylation of host CpG islands.

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Increased ERp57 Expression in HBV-Related Hepatocellular Carcinoma: Possible Correlation and Prognosis

BioMed Research International ◽

10.1155/2017/1252647 ◽

2017 ◽

Vol 2017 ◽

pp. 1-8 ◽

Cited By ~ 2

Author(s):

Miao Liu ◽

Lingyao Du ◽

Zhiliang He ◽

Libo Yan ◽

Ying Shi ◽

...

Keyword(s):

Hepatocellular Carcinoma ◽

Hepatitis B ◽

Cell Line ◽

Cell Lines ◽

Liver Cell ◽

Normal Liver ◽

Hbv Infection ◽

Altered Expression ◽

Liver Cell Line ◽

Liver Tissues

Aim.ERp57 is involved in virus induced endoplasmic reticulum stress (ERS) and plays an important role in tumorigenesis. This study aimed to find whether HBV infection altered ERp57 expression and whether ERp57 regulation was involved in hepatitis B virus-related hepatocellular carcinoma (HBV-HCC) genesis.Materials and Methods.HBV-HCC tissues, chronic hepatitis B (CHB) liver tissues, and normal liver tissues were acquired. ERp57 expressions in these tissues were detected through immunohistochemistry (IHC). And ERp57 expression in liver cell line L02, HBV replicative liver cell line L02-pHBV4.1, and HCC cell lines were detected through western blot for verification. Then medical data on patients providing HCC tissues were collected and analyzed along with ERp57 expression.Results.Higher ERp57 expression was found in HCC and CHB tissues (p<0.001). And HCC cell lines and L02-pHBV4.1 presented higher ERp57 expression as well. In patients, ERp57 expression showed significant differences between death and survival groups (p=0.037). And cumulative survival in patients with higher ERp57 (score⩾8.75) is significantly lower (p=0.009).Conclusion.Our study found increased expression of ERp57 in HBV-HCC. Such altered expression could be related to HBV infection and high ERp57 expression may lead to poor prognosis of HBV-HCC patients.

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Induction of apoptosis after switch-on of the hepatitis B virus X gene mediated by the Cre/loxP recombination system

Journal of General Virology ◽

10.1099/0022-1317-80-12-3257 ◽

1999 ◽

Vol 80 (12) ◽

pp. 3257-3265 ◽

Cited By ~ 50

Author(s):

Yoshizumi Shintani ◽

Hiroshi Yotsuyanagi ◽

Kyoji Moriya ◽

Hajime Fujie ◽

Takeya Tsutsumi ◽

...

Keyword(s):

Cell Death ◽

Hepatitis B Virus ◽

Hepatitis B ◽

Cell Lines ◽

Liver Cell ◽

Cre Recombinase ◽

Wild Type ◽

Cell Clones ◽

B Virus ◽

X Gene

The HBx protein of hepatitis B virus is a multifunctional protein that is implicated in the pathogenesis of hepatocellular carcinoma by regulating gene transcription, causing cell proliferation and, as shown recently, inducing cell death. However, analysis of the effects of HBx in stable cultured cell clones has been hampered because only cell lines that adapted to the effects of HBx were selected during the establishment of cell clones. Here, we describe a system in which transcription of the X gene of hepatitis B virus is switched on by the use of the site-specific Cre recombinase. Two human liver cell lines, HLF and HepG2, were used, the former with a mutant p53 allele and the latter with wild-type p53. The stable cell clones isolated, which carried the X gene in a transcriptionally silent state, were infected with recombinant adenovirus carrying Cre recombinase. Ninety-six hours after adenovirus infection, cell clones that expressed HBx had undergone TUNEL-positive cell death with characteristics of apoptosis. Apoptosis was induced despite concomitant inactivation of the p53 protein as a result of its cytoplasmic translocation by HBx. In contrast, neither the X gene-carrying cells infected with wild-type adenovirus nor various control cells infected with Cre-expressing adenovirus exhibited apoptosis. These results indicate that the expression of HBx protein leads to liver cell apoptosis independently of the p53 pathway. The significance of HBx-induced apoptosis in natural infection is unclear, but it may contribute to the development of hepatitis and serve to spread progeny virus to neighbouring cells while evading the host immune responses.

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Study on CD4/CD8 in liver portal area(Po-L3/T8) and CD8/CD5 in parenchyma(Pa-T8/T1) of liver cell injury on hepatitis B virus(HBV) infection.

Kanzo ◽

10.2957/kanzo.31.1143 ◽

1990 ◽

Vol 31 (10) ◽

pp. 1143-1151

Author(s):

Hitoshi SATA ◽

Mikio ZENIYA ◽

Masami SAKAGUCHI ◽

Sanae OKUYAMA ◽

Tomonobu KAWABE ◽

...

Keyword(s):

Hepatitis B Virus ◽

Hepatitis B ◽

Liver Cell ◽

Cell Injury ◽

Hbv Infection ◽

Portal Area ◽

B Virus ◽

Liver Cell Injury

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Small Interfering RNA Screening for the Small GTPase Rab Proteins Identifies Rab5B as a Major Regulator of Hepatitis B Virus Production

Journal of Virology ◽

10.1128/jvi.00621-19 ◽

2019 ◽

Vol 93 (15) ◽

Cited By ~ 3

Author(s):

Jun Inoue ◽

Masashi Ninomiya ◽

Teruyuki Umetsu ◽

Takuya Nakamura ◽

Takayuki Kogure ◽

...

Keyword(s):

Life Cycle ◽

Hepatitis B Virus ◽

Hepatitis B ◽

Hbv Dna ◽

Vesicular Trafficking ◽

Hbv Infection ◽

Rab Proteins ◽

Early Endosomes ◽

B Virus ◽

Interfering Rna

ABSTRACTViruses are considered to use vesicular trafficking in infected cells, but the details of assembly/release pathways of hepatitis B virus (HBV) are still unknown. To identify key regulators of HBV production, we performed short interfering RNA (siRNA) screening for Rab proteins, which are considered to act as molecular switches in vesicular trafficking using HepG2.2.15 cells. Among 62 Rab proteins, the suppression of Rab5B most significantly increased HBV DNA in the culture supernatant. Surprisingly, 5 days after the transfection of Rab5B siRNA, HBV DNA in the supernatant was increased more than 30-fold, reflecting the increase of infectious HBV particles. Northern blotting showed that transcription of 2.4/2.1-kb mRNA coding envelope proteins containing large hepatitis B surface protein (LHBs) was increased. Analysis of hepatocyte nuclear factors (HNFs) showed that transcription of HNF4α, which is known to enhance 2.4-kb mRNA transcription, was regulated by Rab5B. Also, it was revealed that LHBs had accumulated in the endoplasmic reticulum (ER) after Rab5B depletion but not in the multivesicular body (MVB), which is thought to be an organelle utilized for HBV envelope formation. Therefore, it was considered that Rab5B is required for the transport of LHBs from the ER to MVB. Immunofluorescent microscopy showed that HBs proteins, including LHBs, colocalized with HBc in the ER of Rab5B-depleted cells, suggesting that HBV envelopment occurs not only in the MVB but also in the ER. In conclusion, Rab5B is a key regulator of HBV production and could be a target of antiviral therapy.IMPORTANCEHBV infection is a worldwide health problem, but the mechanisms of how HBV utilizes cellular machinery for its life cycle are poorly understood. In particular, it has been unclear how the viral components and virions are transported among the organelles. The HBV budding site has been reported to be the ER or MVB, but it has not been clearly determined. In this study, siRNA-based screening of Rab proteins using HBV-expressing cells showed that Rab5B, one of the Rab5 isoforms, has important roles in late steps of the HBV life cycle. Although Rab5 is known to work on early endosomes, this study showed that Rab5B plays a role in the transport of LHBs between the ER and MVB. Furthermore, it affects the transcription of LHBs. This is the first report on the mechanisms of HBV envelope protein transport among the organelles, and the results provide important insights into the therapeutic control of HBV infection.

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Identification and characterization of lncRNA AP000253 in occult hepatitis B virus infection

Virology Journal ◽

10.1186/s12985-021-01596-y ◽

2021 ◽

Vol 18 (1) ◽

Author(s):

Qingqin Hao ◽

Zheng Wang ◽

Qinghui Wang ◽

Bo Chen ◽

Huizhong Qian ◽

...

Keyword(s):

Hepatitis B Virus ◽

Plasma Level ◽

Hepatitis B ◽

Cell Lines ◽

Hepatoma Cell ◽

Hbv Infection ◽

Occult Hbv ◽

Potential Biomarker ◽

B Virus ◽

Liver Tissues

Abstract Background Recent studies suggest that lncRNAs may play significant roles in the development of hepatitis B virus (HBV) infection. However, as a special stage of HBV infection, the lncRNA expression in occult HBV infection (OBI) remains unclear. Methods The plasma level of 15 HBV infection-related lncRNAs was initially detected using qRT-PCR in 10 OBI and 10 healthy controls (HCs) in discovery phase. Significantly dysregulated lncRNAs were subsequently validated in another 64 OBI, 20 HCs, 31 chronic hepatitis B (CHB) and 20 asymptomatic HBsAg carriers (ASC). Moreover, the AP000253 expression in liver tissues and its potential biological functions in HBV infection were further investigate with public transcriptomic data and HBV-expressing cell lines. Results Among candidate lncRNAs, the plasma level of AP000253 decreased significantly in OBI, ASC and CHB patients compared to HCs, while no difference was found among OBI, ASC and CHB patients. In liver tissues, similar AP000253 expression was also observed from the GSE83148 dataset, while that in HBV-expressing hepatoma cells was opposite. ROC curve analysis indicated that plasma AP000253 yielded an AUC of 0.73 with 60% sensitivity and 75% specificity when differentiating OBI from HCs, but it could not specifically separate the stage of chronic HBV infection. Furthermore, functional experiments suggested that AP000253 could promote HBV transcription and replication in hepatoma cell lines. Conclusions AP000253 might be involved in HBV replication, and be served as a potential biomarker for HBV infection. In the setting of blood donations, plasma AP000253 would be more useful to moderately distinguish OBI in HBsAg-negative donors. However, the AP000253 expression in liver tissues and associated molecular mechanism of HBV infection deserve further study in future.

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Robust Human and Murine Hepatocyte Culture Models of Hepatitis B Virus Infection and Replication

Journal of Virology ◽

10.1128/jvi.01255-18 ◽

2018 ◽

Vol 92 (23) ◽

Cited By ~ 2

Author(s):

Luhua Qiao ◽

Jianhua Sui ◽

Guangxiang Luo

Keyword(s):

Cell Culture ◽

Hepatitis B Virus ◽

Hepatitis B ◽

Cell Lines ◽

Hbv Infection ◽

Hepatitis B Virus Infection ◽

B Virus ◽

Culture Models ◽

Cell Culture Models ◽

Murine Hepatocyte

ABSTRACTHepatitis B virus (HBV) is a major cause of chronic liver diseases, including hepatitis, cirrhosis, and hepatocellular carcinoma. HBV research has been hampered by the lack of robust cell culture and small animal models of HBV infection. The discovery of sodium taurocholate cotransporting polypeptide (NTCP) as an HBV receptor has been a landmark advance in HBV research in recent years. Ectopic expression of NTCP in nonpermissive HepG2, Huh7, and AML12 cell lines confers HBV susceptibility. However, HBV replication in these human and murine hepatocyte cell lines appeared suboptimal. In the present study, we constructed stable NTCP-expressing HepG2 and AML12 cell lines and found that HBV permissiveness is correlated with NTCP expression. More significantly, we developed robust HBV cell culture models by treating the HBV-infected cells with dimethyl sulfoxide (DMSO) and hydrocortisone, which significantly promoted HBV replication and production. Mechanistic studies suggested that hydrocortisone significantly enhanced the transcription and expression of PGC1α and HNF4α, which are known to promote HBV transcription and replication. These new human and murine hepatocyte culture systems of HBV infection and replication will accelerate the determination of molecular aspects underlying HBV infection, replication, and morphogenesis in human and murine hepatocytes. We anticipate that our HBV cell culture models will also facilitate the discovery and development of antiviral drugs towards the ultimate eradication of chronic hepatitis B virus infection.IMPORTANCEHBV research has been greatly hampered by the lack of robust cell culture and small animal models of HBV infection and propagation. The discovery of NTCP as an HBV receptor has greatly impacted the field of HBV research. Although HBV infection of NTCP-expressing human and murine hepatocyte cell lines has been demonstrated, its replication in cell culture appeared inefficient. To further improve cell culture systems of HBV infection and replication, we constructed NTCP-expressing HepG2 and AML12 cell lines that are highly permissive to HBV infection. More significantly, we found that DMSO and hydrocortisone markedly enhanced HBV transcription and replication in human and murine hepatocytes when added to the cell culture medium. These new cell culture models of HBV infection and replication will facilitate HBV research and antiviral drug discovery towards the ultimate elimination of chronic hepatitis B virus infection.

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Large liver cell change/dysplasia in hepatitis B virus-related liver cirrhosis

The Korean Journal of Hepatology ◽

10.3350/kjhep.2009.15.3.375 ◽

2009 ◽

Vol 15 (3) ◽

pp. 375 ◽

Cited By ~ 1

Author(s):

Haeryoung Kim ◽

Young Nyun Park

Keyword(s):

Liver Cirrhosis ◽

Hepatitis B Virus ◽

Hepatitis B ◽

Liver Cell ◽

B Virus

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How Far we are towards Eradication of HBV Infection

Journal of Gastrointestinal and Liver Diseases ◽

10.15403/jgld.2014.1121.244.hbv ◽

2015 ◽

Vol 24 (4) ◽

pp. 473-479 ◽

Cited By ~ 4

Author(s):

Mihai Voiculescu

Keyword(s):

Immune Response ◽

Hepatitis B Virus ◽

T Cell ◽

Hepatitis B ◽

T Cell Receptors ◽

Hepatitis B Surface Antigen ◽

Hbv Infection ◽

Major Health Problem ◽

Cell Receptors ◽

B Virus

Hepatitis B virus (HBV) infection is a major health problem with an important biological and a significant socio-economic impact all over the world. There is a high pressure to come up with a new and more efficient strategy against HBV infection, especially after the recent success of HCV treatment. Preventing HBV infection through vaccine is currently the most efficient way to decrease HBV-related cirrhosis and liver cancer incidence, as well as the best way to suppress the HBV reservoir. The vaccine is safe and efficient in 80-95% of cases. One of its most important roles is to reduce materno-fetal transmission, by giving the first dose of vaccine in the first 24 hours after birth. Transmission of HBV infection early in life is still frequent, especially in countries with high endemicity.Successful HBV clearance by the host is immune-mediated, with a complex combined innate and adaptive cellular and humoral immune response. Different factors, such as the quantity and the sequence of HBV epitope during processing by dendritic cells and presenting by different HLA molecules or the polymorphism of T cell receptors (TOL) are part of a complex network which influences the final response. A new potential therapeutic strategy is to restore T-cell antiviral function and to improve innate and adaptive immune response by immunotherapeutic manipulation.It appears that HBV eradication is far from being completed in the next decades, and a new strategy against HBV infection must be considered. Abbreviations: ALT: alanine aminotransferase; APC: antigen presenting cells; cccDNA: covalently closed circular DNA; HBIG: hepatitis B immunoglobulin; HbsAg: hepatitis B surface antigen; HBV: hepatitis B virus; HCC: hepatocellular carcinoma; CTL: cytotoxic T lymphocyte; IFN: interferon; NUC: nucleos(t)ide analogues; pg RNA: pre genomic RNA; TLR: toll-like receptors; TOL: T cell receptors.

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