TRIM25 inhibits HBV replication by promoting HBx degradation and the RIG-I-mediated pgRNA recognition.

BACKGROUND: The hepatitis B virus (HBV) vaccine has been efficiently used for decades. However, hepatocellular carcinoma caused by HBV is still prevalent globally. We previously reported that interferon (IFN)-induced tripartite motif-containing 25 (TRIM25) inhibited HBV replication by increasing the IFN expression, and this study aimed to further clarify the anti-HBV mechanism of TRIM25. METHODS: The TRIM25-mediated degradation of hepatitis B virus X (HBx) protein was determined by detecting the expression of HBx in TRIM25-overexpressed or knocked-out HepG2 or HepG2-NTCP cells via Western blotting. Co-immunoprecipitation was performed to confirm the interaction between TRIM25 and HBx, and colocalization of TRIM25 and HBx was identified via immunofluorescence; HBV e-antigen and HBV surface antigen were qualified by using an enzyme-linked immunosorbent assay (ELISA) kit from Kehua Biotech. TRIM25 mRNA, pregenomic RNA (pgRNA), and HBV DNA were detected by quantitative real-time polymerase chain reaction. The retinoic acid-inducible gene I (RIG-I) and pgRNA interaction was verified by RNA-binding protein immunoprecipitation assay. RESULTS: We found that TRIM25 promoted HBx degradation, and confirmed that TRIM25 could enhance the K90-site ubiquitination of HBx as well as promote HBx degradation by the proteasome pathway. Interestingly, apart from the Really Interesting New Gene (RING) domain, the SPRY domain of TRIM25 was also indispensable for HBx degradation. In addition, we found that the expression of TRIM25 increased the recognition of HBV pgRNA by interacting with RIG-I, which further increased the IFN production, and SPRY, but not the RING domain is critical in this process. CONCLUSIONS: The study found that TRIM25 interacted with HBx and promoted HBx-K90-site ubiquitination, which led to HBx degradation. On the other hand, TRIM25 may function as an adaptor, which enhanced the recognition of pgRNA by RIG-I, thereby further promoting IFN production. Our study can contribute to a better understanding of host-virus interaction.

Role of hepatitis B virus non-structural protein HBx on HBV replication, interferon signaling, and hepatocarcinogenesis.

Hepatitis B, a global health concern caused by the hepatitis B virus (HBV), infects nearly 2 billion individuals worldwide, as reported by the World Health Organization (WHO). HBV, a hepatotropic DNA virus, predominantly targets and replicates within hepatocytes. Those carrying the virus are at increased risk of liver cirrhosis and hepatocellular carcinoma, resulting in nearly 900,000 fatalities annually. The HBV X protein (HBx), encoded by the virus's open reading frame x, plays a key role in its virulence. This protein is integral to viral replication, immune modulation, and liver cancer progression. Despite its significance, the precise molecular mechanisms underlying HBx remain elusive. This review investigates the HBx protein's roles in HBV replication, interferon signaling regulation, and hepatocellular carcinoma progression. By understanding the complex interactions between the virus and its host mediated by HBx, we aim to establish a solid foundation for future research and the development of HBx-targeted therapeutics.

When does hepatitis B virus meet long-stranded noncoding RNAs?

Hepatitis B virus (HBV) infection in humans and its associated diseases are long-standing problems. HBV can produce a large number of non-self-molecules during its life cycle, which acts as targets for innate immune recognition and initiation. Among these, interferon and its large number of downstream interferon-stimulated gene molecules are important early antiviral factors. However, the development of an effective antiviral immune response is not simple and depends not only on the delicate regulation of the immune response but also on the various mechanisms of virus-related immune escape and immune tolerance. Therefore, despite there being a relatively well-established consensus on the major pathways of the antiviral response and their component molecules, the complete clearance of HBV remains a challenge in both basic and clinical research. Long-noncoding RNAs (lncRNAs) are generally >200 bp in length and perform different functions in the RNA strand encoding the protein. As an important part of the IFN-inducible genes, interferon-stimulated lncRNAs are involved in the regulation of several HBV infection-related pathways. This review traces the basic elements of such pathways and characterizes the various recent targets of lncRNAs, which not only complement the regulatory mechanisms of pathways related to chronic HBV infection, fibrosis, and cancer promotion but also present with new potential therapeutic targets for controlling HBV infection and the malignant transformation of hepatocytes.

Type-III interferon stimulated gene TRIM31 mutation in an HBV patient blocks its ability in promoting HBx degradation.

TRIM5γ, together with TRIM31, has been shown to promote HBx ubiquitination and degradation. This study aimed to explore whether a patient with HCC (hepatic cell carcinoma) having a small nucleotide inserted into the TRIM31 gene, which made a shorter transcript stop at 768 bp, would result in blocking the activity of TRIM31 in promoting HBx degradation. Besides, this study aimed to determine the binding region of the TRIM31-TRIM5γ-HBx complex. HBV (Hepatitis B virus) infection was reported to induce type-III IFN but not type-I or type-II IFNs, here TRIM31 was found to be a type III rather than a type I stimulated gene, which was indispensable in inhibiting the hepatitis B virus replication by the interferon families. Thus, this study further identified the critical role of TRIM31 in the host-hepatitis B virus interaction.

Type-I-IFN-Stimulated Gene TRIM5γ Inhibits HBV Replication by Promoting HBx Degradation.

To understand the molecular mechanisms that mediate the anti-hepatitis B virus (HBV) effect of interferon (IFN) therapy, we conduct high-throughput bimolecular fluorescence complementation screening to identify potential physical interactions between the HBx protein and 145 IFN-stimulated genes (ISGs). Seven HBx-interacting ISGs have consistent and significant inhibitory effects on HBV replication, among which TRIM5γ suppresses HBV replication by promoting K48-linked ubiquitination and degradation of the HBx protein on the K95 ubiquitin site. The B-Box domain of TRIM5γ under overexpression conditions is sufficient to trigger HBx degradation and is responsible both for interacting with HBx and recruiting TRIM31, which is an ubiquitin ligase that triggers HBx ubiquitination. High expression levels of TRIM5γ in IFN-α-treated HBV patients might indicate a better therapeutic effect. Thus, our studies identify a crucial role for TRIM5γ and TRIM31 in promoting HBx degradation, which may facilitate the development of therapeutic agents for the treatment of patients with IFN-resistant HBV infection.

Type III interferon-induced CBFß inhibits HBV replication by hijacking HBx.

Hepatitis B virus (HBV) and its associated chronic infection remain serious health threats worldwide. However, there is still no impactful approach for clinical treatment of hepatitis B patients. Therefore, developing a better understanding of the interactions between HBV and its host is particularly important. HBV infection has been reported to induce type-III but not type-I or type-II interferon (IFN). In this study, we identified CBFß, an HIV enhancer, as an HBV restriction factor that is specifically induced by type-III IFN in the early stages of HBV infection. Type-III IFN-induced IL-10 played an important role in the production of CBFß. Interestingly, the interaction between CBFß- and HBV-encoded regulatory protein X (HBx) enhanced the stability of CBFß, but notably blocked HBx-mediated promotion of HBV replication. CBFß expression was lower in HBV patients than in healthy persons, and the addition of serum from HBV patients inhibited CBFß expression in HepG2 cells. On the contrary, HBV via HBsAg inhibited type-III IFN-induced CBFß expression and decreased the anti-HBV activity of type-III IFN, suggesting that HBV inhibits antiviral interferon-stimulated gene (ISG) expression and induces IFN resistance. Collectively, our results demonstrate that type-III IFN-triggered and IL-10-induced CBFß are crucial factors for inhibiting HBV replication, and the HBx-CBFß-HBsAg axis reveals a new molecular mechanism of interaction between HBV and its hosts.

Identification of TRIM14 as a Type I IFN-Stimulated Gene Controlling Hepatitis B Virus Replication by Targeting HBx.

Hepatitis B virus (HBV) remains a major cause of hepatic disease that threatens human health worldwide. Type I IFN (IFN-I) therapy is an important therapeutic option for HBV patients. The antiviral effect of IFN is mainly mediated via upregulation of the expressions of the downstream IFN-stimulated genes. However, the mechanisms by which IFN induces ISG production and inhibits HBV replication are yet to be clarified. TRIM14 was recently reported as a key molecule in the IFN-signaling pathway that regulates IFN production in response to viral infection. In this study, we sought to understand the mechanisms by which IFN restricts HBV replication. We confirmed that TRIM14 is an ISG in the hepatic cells, and that the pattern-recognition receptor ligands polyI:C and polydAdT induce TRIM14 dependent on IFN-I production. In addition, IFN-I-activated STAT1 (but not STAT3) directly bound to the TRIM14 promoter and mediated the induction of TRIM14. Interestingly, TRIM14 played an important role in IFN-I-mediated inhibition of HBV, and the TRIM14 SPRY domain interacted with the C-terminal of HBx, which might block the role of HBx in facilitating HBV replication by inhibiting the formation of the Smc-HBx-DDB1 complex. Thus, our study clearly demonstrates that TRIM14 is a STAT1-dependent ISG, and that the IFN-I-TRIM14-HBx axis shows an alternative way to understand the mechanism by which IFN-I inhibits virus replication.

Type I IFN augments IL-27-dependent TRIM25 expression to inhibit HBV replication.

Hepatitis B virus (HBV) can cause chronic hepatitis B, which may lead to cirrhosis and liver cancer. Type I interferon (IFN) is an approved drug for the treatment of chronic hepatitis B. However, the fundamental mechanisms of antiviral action by type I IFN and the downstream signaling pathway are unclear. TRIM25 is an IFN-stimulated gene (ISG) that has an important role in RIG-I ubiquitination and activation. Whether TRIM25 is induced in liver cells by type I IFN to mediate anti-HBV function remains unclear. Here we report that interleukin-27 (IL-27) has a critical role in IFN-induced TRIM25 upregulation. TRIM25 induction requires both STAT1 and STAT3. In TRIM25 knockout HepG2 cells, type I IFN production was consistently attenuated and HBV replication was increased, whereas overexpression of TRIM25 in HepG2 cells resulted in elevated IFN production and reduced HBV replication. More interestingly, we found that TRIM25 expression was downregulated in HBV patients and the addition of serum samples from HBV patients could inhibit TRIM25 expression in HepG2 cells, suggesting that HBV might have involved a mechanism to inhibit antiviral ISG expression and induce IFN resistance. Collectively, our results demonstrate that type I IFN -induced TRIM25 is an important factor in inhibiting HBV replication, and the IFN-IL-27-TRIM25 axis may represent a new target for treating HBV infection.

HBsAg blocks TYPE I IFN induced up-regulation of A3G through inhibition of STAT3.

Interferon (IFN) is a regularly utilized therapeutic for the treatment of chronic hepatitis B and appears to induce superior HBeAg seroconversion comparing nucleos/tide analogs. However, the mechanisms underlying IFN inhibition of HBV replication, as well as poor responses to IFN are unclear. Apobec3G has been reported to be involved in regulating HBV replication. In this study, we investigated Apobec3G expression and regulatory pathways during HBV infection. We show that over-expression of A3G leads to inhibition of HBV replication. We also show that IFN induces a significant increase in A3G protein expression, which is associated with STAT3 activation. We further show that A3G expression in HBV patients is lower compared to non-infected controls, possibly by HBsAg which inhibits IFN induced A3G up-regulation in a dose dependent manner. This process is likely mediated through inhibition of STAT3-Ser727 phosphorylation. The results presented in this study indicate that STAT3 plays an important role in IFN-induced A3G production, and HBsAg may correlated with poor response to IFN treatment.