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

Ko, Chunkyu; Lee, Soo Young; Windisch, Marc P.; Ryu, Wang Shick

doi:10.1128/jvi.02035-14

Cited by 76 publications

(83 citation statements)

References 47 publications

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“…Given these crucial roles in RNA biology, several RNA viruses interact with DDX3X, often with important consequences for viral replication. This includes hepatitis C virus (HCV), norovirus, West Nile virus, and Japanese encephalitis virus (3)(4)(5)(6)(7). This is also the case for HIV and hepatitis B virus (HBV), a peculiar DNA virus that relies on an RNA template and reversed transcription to replicate its genome (8,9).…”

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confidence: 99%

The ATP-Dependent RNA Helicase DDX3X Modulates Herpes Simplex Virus 1 Gene Expression

Khadivjam

Stegen

Hogue-Racine

et al. 2017

J Virol

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The human protein DDX3X is a DEAD box ATP-dependent RNA helicase that regulates transcription, mRNA maturation, and mRNA export and translation. DDX3X concomitantly modulates the replication of several RNA viruses and promotes innate immunity. We previously showed that herpes simplex virus 1 (HSV-1), a human DNA virus, incorporates DDX3X into its mature particles and that DDX3X is required for optimal HSV-1 infectivity. Here, we show that viral gene expression, replication, and propagation depend on optimal DDX3X protein levels. Surprisingly, DDX3X from incoming viral particles was not required for the early stages of the HSV-1 infection, but, rather, the protein controlled the assembly of new viral particles. This was independent of the previously reported ability of DDX3X to stimulate interferon type I production. Instead, both the lack and overexpression of DDX3X disturbed viral gene transcription and thus subsequent genome replication. This suggests that in addition to its effect on RNA viruses, DDX3X impacts DNA viruses such as HSV-1 by an interferon-independent pathway. IMPORTANCE Viruses interact with a variety of cellular proteins to complete their life cycle. Among them is DDX3X, an RNA helicase that participates in most aspects of RNA biology, including transcription, splicing, nuclear export, and translation. Several RNA viruses and a limited number of DNA viruses are known to manipulate DDX3X for their own benefit. In contrast, DDX3X is also known to promote interferon production to limit viral propagation. Here, we show that DDX3X, which we previously identified in mature HSV-1 virions, stimulates HSV-1 gene expression and, consequently, virion assembly by a process that is independent of its ability to promote the interferon pathway. KEYWORDS DDX3X, helicase, herpes, host-pathogen interaction, DNA virus, RNA virus, interferon, herpes simplex virus, host-pathogen interactions, transcriptional regulation, translational control T he human DDX3 protein is a member of a large family of DEAD box ATP-dependent RNA helicases. In humans, it is encoded by the X (DDX3X) and Y (DDX3Y) chromosomes, albeit the latter is restricted to testes (1). It participates in different stages of cellular gene expression, such as transcription, mRNA maturation, and mRNA export and translation (2). Given these crucial roles in RNA biology, several RNA viruses interact with DDX3X, often with important consequences for viral replication. This includes hepatitis C virus (HCV), norovirus, West Nile virus, and Japanese encephalitis virus (3-7). This is also the case for HIV and hepatitis B virus (HBV), a peculiar DNA virus that relies on an RNA template and reversed transcription to replicate its genome (8,9). Furthermore, DDX3X also contributes to innate immunity against these viruses. For instance, DDX3X stimulates interferon (IFN) type I production by binding IKK (IB kinase epsilon) and TBK1 (tank-binding kinase 1), leading to IRF3 phosphorylation and activation (10,

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confidence: 99%

The ATP-Dependent RNA Helicase DDX3X Modulates Herpes Simplex Virus 1 Gene Expression

Khadivjam

Stegen

Hogue-Racine

et al. 2017

J Virol

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“…HepG2 and HEK293 cells were maintained in Dulbecco's modified Eagle's medium (DMEM) containing 10% fetal bovine serum (FBS). HepG2-NTCP cells were maintained in DMEM containing 10% FBS and 2 mM L-glutamine (21). All cells were maintained in a humidified incubator at 37°C with 5% CO 2 .…”

Section: Methodsmentioning

confidence: 99%

“…Lentiviruses expressing green fluorescent protein (GFP) or DDB1-targeting shRNA were obtained by cotransfection of HEK293T cells with the psPAX2, pMD2.G, and pLKO.1-shGFP or pLKO.1-shDDB1 constructs, as previously described (21). Briefly, aliquots of virus supplemented with 4 g/ml Polybrene (Sigma) were used to transduce HepG2 and HepG2-NTCP cells.…”

Section: Methodsmentioning

confidence: 99%

“…The lentivirus plasmid vector pLKO.1-puro containing short hairpin RNA (shRNA) targeting DDB1 was purchased from Sigma-Aldrich. The Luc reporter constructs, including pHBV-Luc, pHBV-S1-Luc, and pHBV-S2-Luc, were previously described (20,21). pHBV-X/EnhI-Luc was constructed by the insertion of fragments containing the X promoter/enhancer I region.…”

Section: Methodsmentioning

confidence: 99%

“…All cells were maintained in a humidified incubator at 37°C with 5% CO 2 . Transient transfection was performed by using polyethylenimine (PEI) (linear, 25 kDa) or Lipofectamine 2000 (Invitrogen), as previously described (21).…”

Section: Methodsmentioning

confidence: 99%

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DDB1 Stimulates Viral Transcription of Hepatitis B Virus via HBx-Independent Mechanisms

Kim

Lee

Son

et al. 2016

J Virol

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HBx, a small regulatory protein of hepatitis B virus (HBV), augments viral DNA replication by stimulating viral transcription. Among numerous reported HBx-binding proteins, DDB1 has drawn attention, because DDB1 acts as a substrate receptor of the Cul4-DDB1 ubiquitin E3 ligase. Previous work reported that the DDB1-HBx interaction is indispensable for HBx-stimulated viral DNA replication, suggesting that the Cul4-DDB1 ubiquitin E3 ligase might target cellular restriction factors for ubiquitination and proteasomal degradation. To gain further insight into the DDB1-HBx interaction, we generated HBx mutants deficient for DDB1 binding (i.e., R96A, L98A, and G99A) and examined whether they support HBx-stimulated viral DNA replication. In contrast to data from previous reports, our results showed that the HBx mutants deficient for DDB1 binding supported viral DNA replication to nearly wild-type levels, revealing that the DDB1-HBx interaction is largely dispensable for HBx-stimulated viral DNA replication. Instead, we found that DDB1 directly stimulates viral transcription regardless of HBx expression. Through an HBV infection study, importantly, we demonstrated that DDB1 stimulates viral transcription from covalently closed circular DNA, a physiological template for viral transcription. Overall, we concluded that DDB1 stimulates viral transcription via a mechanism that does not involve an interaction with HBx. IMPORTANCEDDB1 constitutes a cullin-based ubiquitin E3 ligase, where DDB1 serves as an adaptor linking the cullin scaffold to the substrate receptor. Previous findings that the DDB1-binding ability of HBx is essential for HBx-stimulated viral DNA replication led to the hypothesis that HBx could downregulate host restriction factors that limit HBV replication through the cullin ubiquitin E3 ligase that requires the DDB1-HBx interaction. Consistent with this hypothesis, recent work identified Smc5/6 as a host restriction factor that is regulated by the viral cullin ubiquitin E3 ligase. In contrast, here we found that the DDB1-HBx interaction is largely dispensable for HBx-stimulated viral DNA replication. Instead, our results clearly showed that DDB1, regardless of HBx expression, enhances viral transcription. Overall, besides its role in the viral cullin ubiquitin E3 ligase, DDB1 itself stimulates viral transcription via HBx-independent mechanisms. H epatitis B virus (HBV) infection represents a major global public health concern, with over 300 million chronically infected patients worldwide. Chronic HBV infection carries a great risk of developing into severe liver diseases, including cirrhosis and liver cancer, which result in a million deaths annually (1). HBV carries a small (ca. 3.2-kb), relaxed circular (RC), partially double-stranded DNA. Following entry into the host cell, the viral RC DNA traffics to the nucleus, where it is converted into a covalently closed circular DNA (cccDNA), the template for viral transcription (2). Four viral RNAs are transcribed from their respective promoters and serve a...

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Global microRNA expression profiling in the liver biopsies of hepatitis B virus–infected patients suggests specific microRNA signatures for viral persistence and hepatocellular injury

Singh¹,

Rooge²,

Varshney³

et al. 2018

Hepatology

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

Cited by 76 publications

References 47 publications

The ATP-Dependent RNA Helicase DDX3X Modulates Herpes Simplex Virus 1 Gene Expression

The ATP-Dependent RNA Helicase DDX3X Modulates Herpes Simplex Virus 1 Gene Expression

DDB1 Stimulates Viral Transcription of Hepatitis B Virus via HBx-Independent Mechanisms

Global microRNA expression profiling in the liver biopsies of hepatitis B virus–infected patients suggests specific microRNA signatures for viral persistence and hepatocellular injury

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