Hepatitis B virus HBx protein localized to the nucleus restores HBx-deficient virus replication in HepG2 cells and in vivo in hydrodynamically-injected mice

Keasler, Victor V.; Hodgson, Amanda; Madden, Charles R.; Slagle, Betty L.

doi:10.1016/j.virol.2009.05.001

Cited by 59 publications

(68 citation statements)

References 56 publications

(97 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Although our observations imply that HBx exerts its role in potentiating HBV replication by acting directly on the cccDNA, we cannot exclude that Ca ϩϩ mobilization and src activation (13,15) might also play a role. Importantly, it has been recently reported that HBx protein localized to the nucleus restores HBx-deficient virus replication in HepG2 cells and in vivo in hydrodynamically injected mice (39), thus supporting the importance of HBx interaction with the cccDNA in the nucleus. Altogether our results indicate that HBx influences the epigenetic control of nuclear HBV cccDNA function by modulating the recruitment of chromatinmodifying enzymes onto the viral minichromosome.…”

Section: Discussionmentioning

confidence: 82%

Nuclear HBx binds the HBV minichromosome and modifies the epigenetic regulation of cccDNA function

Belloni

Pollicino

Nicola

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

424

482

View full text Add to dashboard Cite

HBV cccDNA, the template for transcription of all viral mRNAs, accumulates in the nucleus of infected cells as a stable episome organized into minichromosomes by histones and non-histone viral and cellular proteins. Using a cccDNA-specific chromatin immunoprecipitation (ChIP)-based quantitative assay, we have previously shown that transcription of the HBV minichromosome is regulated by epigenetic changes of cccDNA-bound histones and that modulation of the acetylation status of cccDNA-bound H3/H4 histones impacts on HBV replication. We now show that the cellular histone acetyltransferases CBP, p300, and PCAF/GCN5, and the histone deacetylases HDAC1 and hSirt1 are all recruited in vivo onto the cccDNA. We also found that the HBx regulatory protein produced in HBV replicating cells is recruited onto the cccDNA minichromosome, and the kinetics of HBx recruitment on the cccDNA parallels the HBV replication. As expected, an HBV mutant that does not express HBx is impaired in its replication, and exogenously expressed HBx transcomplements the replication defects. p300 recruitment is severely impaired, and cccDNA-bound histones are rapidly hypoacetylated in cells replicating the HBx mutant, whereas the recruitment of the histone deacetylases hSirt1 and HDAC1 is increased and occurs at earlier times. Finally, HBx mutant cccDNA transcribes significantly less pgRNA. Altogether our results further support the existence of a complex network of epigenetic events that influence cccDNA function and HBV replication and identify an epigenetic mechanism (i.e., to prevent cccDNA deacetylation) by which HBx controls HBV replication.histone acetylation ͉ HATs ͉ HDACs H epatitis B virus (HBV) infection is a major health problem, with Ϸ400 million people chronically infected worldwide who are at high risk of developing liver cirrhosis and hepatocellular carcinoma (HCC) (1). The epidemiological evidence linking HBV infection to HCC is very strong, and despite the mechanisms underlying HBV-associated carcinogenesis remain to be fully defined, a growing number of studies support a direct role of HBV in the process (2-5). The HBV-encoded regulatory protein hepatitis B virus X protein (HBx) is thought to contribute to HBV oncogenicity (5, 6). HBx transforms SV40-immortalized murine hepatocytes, induces cell cycle progression within the regenerating liver, causes liver cancer in some transgenic mice, and acts as a cofactor to accelerate cancer development in other mouse models (6-11). HBx is a 154-amino acid protein with an N-terminal negative regulatory domain and C-terminal transactivation or coactivation domain that has been detected both in the cytoplasm and in the nuclei of infected hepatocytes (6,12,13). Studies in transfected cells have shown that HBx expression affects several cellular functions such as cytoplasmic calcium regulation, cell signaling, transcription, cell proliferation, DNA repair, and apoptosis (11, 13-16). To perform its multiple functions, HBx interacts with many cellular partners including the tumor suppressor p53,...

show abstract

Section: Discussionmentioning

confidence: 82%

Nuclear HBx binds the HBV minichromosome and modifies the epigenetic regulation of cccDNA function

Belloni

Pollicino

Nicola

et al. 2009

Proc. Natl. Acad. Sci. U.S.A.

424

482

View full text Add to dashboard Cite

show abstract

“…Results have shown that the absence of HBx can induce a 65% reduction in HBV replication (6), and trans-complementation of HBx for the HBx-deficient plasmid can restore replication to wild-type levels (6)(7)(8). Another advanced study performed with the hydrodynamic injection of an HBx-deficient plasmid into mice showed the same results in vivo (6,9,10). These results demonstrated that HBx has an important role in modulating HBV replication.…”

Section: Introductionmentioning

confidence: 71%

Hepatitis B virus X protein modulates remodelling of minichromosomes related to hepatitis B virus replication in HepG2 cells

Luo

Chen

Gong

et al. 2012

International Journal of Molecular Medicine

View full text Add to dashboard Cite

Abstract. Hepatitis B virus (HBV) covalently closed circular DNA (cccDNA) is organised into minichromosomes by histone and non-histone proteins. Remodelling of minichromosomes is crucial for the regulation of HBV replication, which is dependent on the presence of the hepatitis B virus X protein (HBx). However, the mechanisms of HBx-dependent HBV replication remain obscure. The objective of this study was to investigate the mechanism of HBx-dependent HBV replication through the pathway of chromatin remodelling. The role of HBx was investigated by transfecting human HepG2 cells with the linear full-length HBV genome (wild-type) or HBx-deficient mutant HBV DNA (HBx mutant). Our results showed that although the formation of cccDNA was not affected by HBx, HBV replication, transcription and antigen secretion were all significantly reduced, resulting from the absence of HBx. The acetylation, mono-methylation and phosphorylation of cccDNA-bound histone H3 were associated with HBV replication. In addition, the levels of cccDNA-bound methylated, phosphorylated and acetylated histone H3 decreased sharply in HBx mutant HBV DNA. HBx modulated not only the status of acetylation but also the methylation and phosphorylation of histone H3 bound to the cccDNA during HBV replication in HepG2 cells. These findings suggest that HBx plays an important role in modulating the remodelling of minichromosomes related to HBV replication and it may regulate viral replication through the pathway of chromatin remodelling.

show abstract

“…Human embryonic kidney 293T and liver HepG2 cells were maintained in Eagle's minimum essential medium (EMEM) with 10% fetal bovine serum (FBS), as previously described (23). Plasmid DNA encoding IPS-1 containing hemagglutinin (HA) and FLAG epitopes was obtained from Zhijian Chen (28).…”

Section: Methodsmentioning

confidence: 99%

“…Plasmid DNA encoding IPS-1 containing hemagglutinin (HA) and FLAG epitopes was obtained from Zhijian Chen (28). Plasmid DNA encoding HBx (subtype adw2) was prepared by cloning into plasmid vector pSI (Promega) as previously described (23). pSI-X plasmids with an in-frame nuclear localization signal (pSI-NLS-X) or a nuclear export signal (pSI-NES-X) were previously described (23).…”

Section: Methodsmentioning

confidence: 99%

Hepatitis B Virus Regulatory HBx Protein Binds to Adaptor Protein IPS-1 and Inhibits the Activation of Beta Interferon

Kumar

Jung

Hodgson

et al. 2011

J Virol

Self Cite

123

119

View full text Add to dashboard Cite

Hepatitis B virus (HBV) is a small (3.2-kb) DNA virus that causes acute and chronic inflammation of the liver, and the latter is a risk factor for the development of hepatocellular carcinoma (HCC) (39). Worldwide, an estimated 350 million people have chronic HBV and are at risk for severe liver disease (39). New insight into the virus-host interactions underlying chronic virus replication was provided with the demonstration that HBV infection fails to activate the innate immune response in chimpanzees (52). This observation was recently confirmed in acutely infected humans (10) and primary human hepatocytes exposed to HBV (17). Several studies have clearly demonstrated that HBV replication is controlled by an activated adaptive immune response (4,14,36,53,54), suggesting that HBV has evolved a strategy to dampen activation of the innate immune response.The sole HBV regulatory protein, the 17-kDa HBx protein, plays an essential role in virus replication in HepG2 cells (3) and in hydrodynamically injected mice (22). Given the abundance of properties attributed to HBx (reviewed in reference 2), it is likely that HBx has more than one function during the virus life cycle. These functions may be mediated, in part, through HBx interactions with cellular proteins. Indeed, screening protocols such as the yeast two-hybrid assay have been used to identify over 30 HBx-interacting proteins (reviewed in reference 13). The biologic importance of these virus-host protein interactions is difficult to assess due to a paucity of virus replication assays. In the related woodchuck virus replication model, it was demonstrated that the HBx interaction with cellular DDB1 is critical for virus replication (42), and this was confirmed in plasmid-transfected HepG2 cells (27). However, the role of other HBx binding partners in virus replication remains unknown. Cells respond to virus infection through the recognition of viral pathogen-associated molecular patterns (PAMPs). Several cytoplasmic host pattern recognition receptors (PRRs) are responsible for this, including the Toll-like receptors (TLRs) and retinoic acid-inducible gene I (RIG-I) and other RIG-Ilike receptors (RLRs), such as MDA-5 and LGP2 (43; also reviewed in reference 55). Following recognition of the viral DNA or RNA, the PRRs undergo conformational changes that activate downstream pathways, ultimately leading to the induction of type I interferon (IFN) and proinflammatory cytokines. A key adaptor protein in this process is the beta interferon promoter stimulator 1 (IPS-1) protein (21), also known as mitochondrial antiviral signaling protein (MAVS) (40), VISA (58), and Cardiff (35; also reviewed in reference 20). Upon its activation, IPS-1 recruits kinases that phosphorylate latent transcription factors required for the production of 21,25). Interestingly, IPS-1 is targeted for interaction by several viral proteins, effectively inactivating the innate antiviral immune response (43).The goal of the present study was to identify HBx-interacting proteins in the liver of the H...

show abstract

Hepatitis B virus HBx protein localized to the nucleus restores HBx-deficient virus replication in HepG2 cells and in vivo in hydrodynamically-injected mice

Cited by 59 publications

References 56 publications

Nuclear HBx binds the HBV minichromosome and modifies the epigenetic regulation of cccDNA function

Nuclear HBx binds the HBV minichromosome and modifies the epigenetic regulation of cccDNA function

Hepatitis B virus X protein modulates remodelling of minichromosomes related to hepatitis B virus replication in HepG2 cells

Hepatitis B Virus Regulatory HBx Protein Binds to Adaptor Protein IPS-1 and Inhibits the Activation of Beta Interferon

Contact Info

Product

Resources

About