Inhibition of Hepatitis B Virus Replication by MyD88 Involves Accelerated Degradation of Pregenomic RNA and Nuclear Retention of Pre-S/S RNAs

Li, Jianhua; Lin, Shujing; Chen, Qiying; Lü, Peng; Zhai, Jianwei; Liu, Yinghui; Yuan, Zhenghong

doi:10.1128/jvi.00236-10

Cited by 69 publications

(60 citation statements)

References 57 publications

(76 reference statements)

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“…To rule out the possibility that the reduction in viral RNAs can be partially attributed to accelerated decay of viral RNAs caused by DDX3, we examined whether DDX3 promotes HBV RNA decay, as previously described (39,40). Our data showed that regardless of DDX3 expression, the half-life of pgRNA was determined to be approximately 5 h (Fig.…”

Section: Fig 4 Ddx3 Inhibits Hbv Promoter Activities (A) a Schematicmentioning

confidence: 89%

DDX3 DEAD-Box RNA Helicase Is a Host Factor That Restricts Hepatitis B Virus Replication at the Transcriptional Level

Lee

Windisch

et al. 2014

J Virol

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DDX3 is a member of the DEAD-box RNA helicase family, involved in mRNA metabolism, including transcription, splicing, and translation. We previously identified DDX3 as a hepatitis B virus (HBV) polymerase (Pol) binding protein, and by using a transient transfection, we found that DDX3 inhibits HBV replication at the posttranscriptional level, perhaps following encapsidation. To determine the exact mechanism of the inhibition, we here employed a diverse HBV experimental system. Inconsistently, we found that DDX3-mediated inhibition occurs at the level of transcription. By using tetracycline-inducible HBV-producing cells, we observed that lentivirus-mediated DDX3 expression led to a reduced level of HBV RNAs. Importantly, knockdown of DDX3 by short hairpin RNA resulted in augmentation of HBV RNAs in two distinct HBV replication systems: (i) tetracyclineinducible HBV-producing cells and (ii) constitutive HBV-producing HepG2.2.15 cells. Moreover, DDX3 knockdown in HBVsusceptible HepG2-NTCP cells, where covalently closed circular DNA (cccDNA) serves as the template for viral transcription, resulted in increased HBV RNAs, validating that transcription regulation by DDX3 occurs on a physiological template. Overall, our results demonstrate that DDX3 represents an intrinsic host antiviral factor that restricts HBV transcription. IMPORTANCE Upon entry into host cells, viruses encounter host factors that restrict viral infection. During evolution, viruses have acquired the ability to subvert cellular factors that adversely affect their replication. Such host factors include TRIM5␣ and APOBEC3G, which were discovered in retroviruses. The discovery of host restriction factors provided deeper insight into the innate immune response and viral pathogenesis, leading to better understanding of host-virus interactions. In contrast to the case with retroviruses, little is known about host factors that restrict hepatitis B virus (HBV), a virus distantly related to retroviruses. DDX3DEAD box RNA helicase is best characterized as an RNA helicase involved in RNA metabolism, such as RNA processing and translation. Here, we show that DDX3 inhibits HBV infection at the level of viral transcription. C hronic hepatitis B virus (HBV) infection represents a major public health burden, affecting more than 300 million individuals worldwide, and carries a high risk for developing cirrhosis and hepatocellular carcinoma (HCC) (1). HBV virions contain a small, partially double-stranded circular DNA genome of 3.2 kb in length. Although it is a DNA virus, HBV replicates its DNA genome via reverse transcription. Upon infection, the virion DNA is converted into covalently closed circular DNA (cccDNA), which then serves as the template for viral transcription (2). Among the viral transcripts, only pregenomic RNA (pgRNA), 3.5 kb in length, is selectively packaged into nucleocapsids along with HBV polymerase (Pol). Inside the nucleocapsid, the pgRNA is reverse transcribed by HBV Pol to yield relaxed circular (RC) DNA. These mature RC DNA-containin...

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Section: Fig 4 Ddx3 Inhibits Hbv Promoter Activities (A) a Schematicmentioning

confidence: 89%

DDX3 DEAD-Box RNA Helicase Is a Host Factor That Restricts Hepatitis B Virus Replication at the Transcriptional Level

Lee

Windisch

et al. 2014

J Virol

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“…ISG modulation results in an antiviral response in target cells aimed at limiting both viral replication and spreading. IFN-α has been reported to inhibit HBV replication through a variety of mechanisms, including a block of RNA-containing core particle formation, an accelerated decay of replication-competent core particles, and degradation of the pgRNA (7)(8)(9).…”

Section: Introductionmentioning

confidence: 99%

IFN-α inhibits HBV transcription and replication in cell culture and in humanized mice by targeting the epigenetic regulation of the nuclear cccDNA minichromosome

Belloni¹,

Allweiss²,

Guerrieri³

et al. 2012

J. Clin. Invest.

495

422

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HBV infection remains a leading cause of death worldwide. IFN-α inhibits viral replication in vitro and IntroductionHepatitis B Virus (HBV) infection remains a major health problem worldwide despite the availability of a highly effective preventive vaccine. HBV is a noncytopathic hepatotropic DNA virus that belongs to the family Hepadnaviridae, whose members share a distinctive strategy for replication. HBV replication occurs in the cytoplasm within viral capsids (core particles), where a genomesized RNA replicative intermediate, termed the pregenome (pgRNA), is converted by the virally encoded RNA-dependent and DNA-dependent reverse transcriptase/polymerase into a specific open circular (OC) duplex DNA (1). Transcription in the nucleus of the pgRNA from the covalently closed circular DNA (cccDNA) is the critical step for genome amplification and ultimately determines the rate of HBV replication (2). The cccDNA, which also serves as the template for the transcription of all viral messenger RNAs, is organized into a minichromosome in the nuclei of infected hepatocytes by histone and nonhistone proteins, and its function is regulated, similarly to cellular chromatin, by the activity of nuclear transcription factors, transcriptional coactivators and corepressors, and chromatin-modifying enzymes (2-4).Current antiviral therapies involve the use of nucleoside analogs and pegylated IFN-α (5). IFN-α, a type I IFN, engages the IFN-α/β receptor complex to activate the intracellular Jak/Stat signaling pathway, which modulates the transcription of a diverse set of target genes, referred to as IFN-stimulated genes (ISGs) (6). ISG

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“…Huh7 cells were grown to 30 to 40% confluence and then transiently transfected with the indicated small interfering RNA (siRNA) duplexes (Invitrogen or Santa Cruz Biotechnology) at a final concentration of 40 nM using Lipofectamine 2000 (Invitrogen) according to the manufacturer's instructions and as described previously (47). At 24 h posttransfection, the cells were transfected with 20 nM the same siRNA duplexes and the indicated plasmids.…”

mentioning

confidence: 99%

“…Intracellular viral RNA and DNA analyses were performed as described previously (47). For viral RNA analysis, total cellular RNA was extracted with TRIzol reagents.…”

mentioning

confidence: 99%

Subversion of Cellular Autophagy Machinery by Hepatitis B Virus for Viral Envelopment

Liu

Wang

et al. 2011

J Virol

Self Cite

237

296

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Autophagy is a conserved eukaryotic mechanism that mediates the removal of long-lived cytoplasmic macromolecules and damaged organelles via a lysosomal degradative pathway. Recently, a multitude of studies have reported that viral infections may have complex interconnections with the autophagic process. The findings reported here demonstrate that hepatitis B virus (HBV) can enhance the autophagic process in hepatoma cells without promoting protein degradation by the lysosome. Mutation analysis showed that HBV small surface protein (SHBs) was required for HBV to induce autophagy. The overexpression of SHBs was sufficient to induce autophagy. Furthermore, SHBs could trigger unfolded protein responses (UPR), and the blockage of UPR signaling pathways abrogated the SHB-induced lipidation of LC3-I. Meanwhile, the role of the autophagosome in HBV replication was examined. The inhibition of autophagosome formation by the autophagy inhibitor 3-methyladenine (3-MA) or small interfering RNA duplexes targeting the genes critical for autophagosome formation (Beclin1 and ATG5 genes) markedly inhibited HBV production, and the induction of autophagy by rapamycin or starvation greatly contributed to HBV production. Furthermore, evidence was provided to suggest that the autophagy machinery was required for HBV envelopment but not for the efficiency of HBV release. Finally, SHBs partially colocalized and interacted with autophagy protein LC3. Taken together, these results suggest that the host's autophagy machinery is activated during HBV infection to enhance HBV replication.Hepatitis B virus (HBV) is a noncytopathic, enveloped DNA virus that belongs to the family Hepadnaviridae (57, 70). It is one of the most successful human pathogens, with an estimated 2 billion people infected worldwide, of whom 400 million have chronic HBV infection (54). Chronic HBV infection is correlated with a strongly increased risk for the development of liver cirrhosis and hepatocellular carcinoma (HCC) (49, 54).Effective preventive vaccines against HBV have been available for nearly three decades; however, their effectiveness in preventing blood-borne transmission from an infected mother to her newborn is about 90% (77), and therapeutic vaccines for the treatment of established hepatitis B infection are not available (65, 67). Currently, two types of antiviral therapies are approved: pegylated alpha interferon and nucleoside/nucleotide analogues (60). However, these antivirals cannot completely eradicate the virus, and their efficacy in preventing liver cirrhosis and HCC is limited (21, 64). Thus, the details of the host-virus relationship during HBV infection urgently need to be further clarified to facilitate the development of efficient therapeutic strategies for the control of HBV infection.Autophagy, an evolutionarily conserved intracellular process, involves the formation of a double membrane structure, called the autophagosome, which engulfs long-lived cytoplasmic macromolecules and damaged organelles and delivers them to lysosomes for degrada...

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Inhibition of Hepatitis B Virus Replication by MyD88 Involves Accelerated Degradation of Pregenomic RNA and Nuclear Retention of Pre-S/S RNAs

Cited by 69 publications

References 57 publications

DDX3 DEAD-Box RNA Helicase Is a Host Factor That Restricts Hepatitis B Virus Replication at the Transcriptional Level

DDX3 DEAD-Box RNA Helicase Is a Host Factor That Restricts Hepatitis B Virus Replication at the Transcriptional Level

IFN-α inhibits HBV transcription and replication in cell culture and in humanized mice by targeting the epigenetic regulation of the nuclear cccDNA minichromosome

Subversion of Cellular Autophagy Machinery by Hepatitis B Virus for Viral Envelopment

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