Hepatitis B Virus HBx Protein Mediates the Degradation of Host Restriction Factors through the Cullin 4 DDB1 E3 Ubiquitin Ligase Complex

The hepatitis B virus (HBV) regulatory HBx protein is required for infection, and its binding to cellular damaged DNA binding protein 1 (DDB1) is critical for this function. DDB1 is an adaptor protein for the cullin 4A Really Interesting New Gene (RING) E3 ubiquitin ligase (CRL4) complex and functions by binding cellular DDB1 cullin associated factor (DCAF) receptor proteins that recruit substrates for ubiquitination and degradation. We compared the proteins found in the CRL4 complex immunoprecipitated from uninfected versus HBV-infected hepatocytes from human liver chimeric mice for insight into mechanisms by which HBV and the cell interact within the CRL4 complex. Consistent with its role as a viral DCAF, HBx was found in the HBV CRL4 complexes. In tissue culture transfection experiments, we showed that HBx expression led to decreased levels of known restriction factor structural maintenance of chromosomes protein 6 (SMC6) and putative restriction factors stromal interaction molecule 1 (STIM1, zinc finger E-box binding homeobox 2 (ZEB2), and proteasome activator subunit 4 (PSME4). Moreover, silencing of these proteins led to increased HBV replication in the HepG2-sodium taurocholate cotransporting polypeptide (NTCP) infection model. We also identified cellular DCAF receptors in CRL4 complexes from humanized mice. Increasing amounts of HBx did not reveal competitive DCAF binding to cullin4 (CUL4)-DDB1 in plasmid-transfected cells. Our results suggest a model in which HBx benefits virus replication by directly or indirectly degrading multiple cellular restriction factors.

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Hepatitis B virus X protein activates E3 ubiquitin ligase Siah-1 to control virus propagation via a negative feedback loop

Journal of General Virology ◽

10.1099/jgv.0.000856 ◽

2017 ◽

Vol 98 (7) ◽

pp. 1774-1784 ◽

Cited By ~ 6

Author(s):

Sujeong Yeom ◽

Soo Shin Kim ◽

Hyerin Jeong ◽

Kyung Lib Jang

Keyword(s):

Hepatitis B Virus ◽

Hepatitis B ◽

Ubiquitin Ligase ◽

Negative Feedback ◽

Feedback Loop ◽

E3 Ubiquitin Ligase ◽

Negative Feedback Loop ◽

Virus Propagation ◽

B Virus ◽

Control Virus

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Hepatitis B Virus X Protein Sensitizes TRAIL-Induced Hepatocyte Apoptosis by Inhibiting the E3 Ubiquitin Ligase A20

PLoS ONE ◽

10.1371/journal.pone.0127329 ◽

2015 ◽

Vol 10 (5) ◽

pp. e0127329 ◽

Cited By ~ 14

Author(s):

Hang Zhang ◽

Changxin Huang ◽

Yan Wang ◽

Zhe Lu ◽

Ningtong Zhuang ◽

...

Keyword(s):

Hepatitis B Virus ◽

Hepatitis B ◽

Ubiquitin Ligase ◽

E3 Ubiquitin Ligase ◽

Hepatocyte Apoptosis ◽

X Protein ◽

B Virus

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O-024: A novel CRISPR/Cas9-based approach to transient activation of intracellular host restriction factors results in strong suppression of hepatitis B virus and degradation of cccDNA

Journal of Viral Hepatitis ◽

10.1111/jvh.20_12922 ◽

2018 ◽

Vol 25 ◽

pp. 16-17 ◽

Cited By ~ 1

Keyword(s):

Hepatitis B Virus ◽

Hepatitis B ◽

Restriction Factors ◽

Strong Suppression ◽

Host Restriction ◽

Host Restriction Factors ◽

Transient Activation ◽

B Virus

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Mitochondria ubiquitin ligase, MARCH5 resolves hepatitis B virus X protein aggregates in the liver pathogenesis

Cell Death and Disease ◽

10.1038/s41419-019-2175-z ◽

2019 ◽

Vol 10 (12) ◽

Cited By ~ 5

Author(s):

Young-Suk Yoo ◽

Yeon-Ji Park ◽

Ho-Soo Lee ◽

Nguyen Thi Kim Oanh ◽

Mi-Young Cho ◽

...

Keyword(s):

Liver Disease ◽

Hepatitis B Virus ◽

Hepatitis B ◽

Ubiquitin Ligase ◽

E3 Ubiquitin Ligase ◽

Protein Aggregates ◽

Agarose Gels ◽

Protein Levels ◽

B Virus ◽

Proteasome Pathway

AbstractInfection of hepatitis B virus (HBV) increase the incidence of chronic liver disease and hepatocellular carcinoma (HCC). The hepatitis B viral x (HBx) protein encoded by the HBV genome contributes to the pathogenesis of HCC and thus, negative regulation of HBx is beneficial for the alleviation of the disease pathogenesis. MARCH5 is a mitochondrial E3 ubiquitin ligase and here, we show that high MARCH5 expression levels are correlated with improved survival in HCC patients. MARCH5 interacts with HBx protein mainly accumulated in mitochondria and targets it for degradation. The N-terminal RING domain of MARCH5 was required for the interaction with HBx, and MARCH5H43W lacking E3 ligase activity failed to reduce HBx protein levels. High expression of HBx results in the formation of protein aggregates in semi-denaturing detergent agarose gels and MARCH5 mediates the elimination of protein aggregates through the proteasome pathway. HBx-induced ROS production, mitophagy, and cyclooxygenase-2 gene expression were suppressed in the presence of high MARCH5 expression. These results suggest MARCH5 as a target for alleviating HBV-mediated liver disease.

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Notch signaling facilitates hepatitis B virus covalently closed circular DNA transcription via cAMP response element-binding protein with E3 ubiquitin ligase-modulation

Scientific Reports ◽

10.1038/s41598-018-38139-5 ◽

2019 ◽

Vol 9 (1) ◽

Cited By ~ 3

Author(s):

Zijing Wang ◽

Kazunori Kawaguchi ◽

Masao Honda ◽

Shinichi Hashimoto ◽

Takayoshi Shirasaki ◽

...

Keyword(s):

Hepatitis B Virus ◽

Hepatitis B ◽

Notch Signaling ◽

Ubiquitin Ligase ◽

E3 Ubiquitin Ligase ◽

Camp Response Element Binding ◽

Camp Response Element ◽

Circular Dna ◽

B Virus ◽

Element Binding Protein

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E3 ubiquitin ligase TRIM21 restricts hepatitis B virus replication by targeting HBx for proteasomal degradation

Antiviral Research ◽

10.1016/j.antiviral.2021.105107 ◽

2021 ◽

Vol 192 ◽

pp. 105107

Author(s):

Yahui Song ◽

Min Li ◽

Yanqi Wang ◽

Hongkai Zhang ◽

Lin Wei ◽

...

Keyword(s):

Hepatitis B Virus ◽

Hepatitis B ◽

Virus Replication ◽

Ubiquitin Ligase ◽

E3 Ubiquitin Ligase ◽

Proteasomal Degradation ◽

B Virus

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P185 PROTEIN KINASE C-INDUCED ENDOCYTOSIS OF SODIUM TAUROCHOLATE COTRANSPORTING POLYPEPTIDE MEDIATES CELLULAR ENTRY OF HEPATITIS B VIRUS IN AN IN VITRO HBV MODEL

Journal of Hepatology ◽

10.1016/s0168-8278(14)60347-0 ◽

2014 ◽

Vol 60 (1) ◽

pp. S127-S128

Author(s):

J.W. Chung ◽

I.Y. Moon ◽

S.W. Yang ◽

E.S. Jang ◽

S.-H. Jeong ◽

...

Keyword(s):

Protein Kinase C ◽

Hepatitis B Virus ◽

Protein Kinase ◽

Hepatitis B ◽

Sodium Taurocholate ◽

Kinase C ◽

B Virus ◽

Cellular Entry ◽

Hbv Model

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NTCP oligomerization occurs downstream of the NTCP-EGFR interaction during hepatitis B virus internalization

Journal of Virology ◽

10.1128/jvi.00938-21 ◽

2021 ◽

Author(s):

Kento Fukano ◽

Mizuki Oshima ◽

Senko Tsukuda ◽

Hideki Aizaki ◽

Mio Ohki ◽

...

Keyword(s):

Epidermal Growth Factor Receptor ◽

Bile Acid ◽

Hepatitis B Virus ◽

Hepatitis B ◽

Viral Entry ◽

Sodium Taurocholate ◽

Growth Factor Receptor ◽

Host Cells ◽

B Virus ◽

Epidermal Growth

Sodium taurocholate cotransporting polypeptide (NTCP) is a receptor that is essential for hepatitis B virus (HBV) entry into the host cell. A number of HBV entry inhibitors targeting NTCP have been reported to date; these inhibitors have facilitated a mechanistic analysis of the viral entry process. However, the mechanism of HBV internalization into host cells after interaction of virus with NTCP remains largely unknown. Recently, we reported that troglitazone, a thiazolidinedione derivative, specifically inhibits both HBV internalization and NTCP oligomerization, resulting in inhibition of HBV infection. Here, using troglitazone as a chemical probe to investigate entry process, the contribution of NTCP oligomerization to HBV internalization was evaluated. Using surface plasmon resonance and transporter kinetics, we found that troglitazone directly interacts with NTCP and non-competitively interferes with NTCP-mediated bile acid uptake, suggesting that troglitazone allosterically binds to NTCP, rather than to the bile acid-binding pocket. Additionally, alanine scanning mutagenesis showed that a mutation at phenylalanine 274 of NTCP (F274A) caused a loss of HBV susceptibility and disrupted both the oligomerization of NTCP and HBV internalization without affecting viral attachment to the cell surface. An inhibitor of the interaction between NTCP and epidermal growth factor receptor (EGFR), another host cofactor essential for HBV internalization, impeded NTCP oligomerization. Meanwhile, co-immunoprecipitation analysis revealed that neither troglitazone nor the F274A mutation in NTCP affect the NTCP-EGFR interaction. These findings suggest that NTCP oligomerization is initiated downstream of the NTCP-EGFR interaction, and then triggers HBV internalization. This study provides significant insight into the HBV entry mechanisms. Importance Hepatitis B virus (HBV) infection is mediated by a specific interaction with sodium taurocholate cotransporting polypeptide (NTCP), a viral entry receptor. Although the virus-receptor interactions are believed to trigger viral internalization into host cells, the exact molecular mechanisms of HBV internalization are not understood. In this study, we revealed the mode of action whereby troglitazone, a specific inhibitor of HBV internalization, impedes NTCP oligomerization, and identified NTCP phenylalanine 274 as a residue essential for this oligomerization. We further analyzed the association between NTCP oligomerization and HBV internalization, a process that is mediated by epidermal growth factor receptor (EGFR), another essential host cofactor for HBV internalization. Our study provides critical information on the mechanism of HBV entry, and suggests that oligomerization of the viral receptor serves as an attractive target for drug discovery.

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