PRMT5 restricts HBV replication through a two-part mechanism including epigenetic suppression of cccDNA transcription and interference with pregenomic RNA encapsidation; these findings improve the understanding of epigenetic regulation of HBV transcription and host-HBV interaction, thus providing new insights into targeted therapeutic intervention. (Hepatology 2017;66:398-415).
Background and Aims
Interferon (IFN)‐α, composed of numerous subtypes, plays a crucial role in immune defense. As the most studied subtype, IFN‐α2 has been used for treating chronic hepatitis B virus (HBV) infection, with advantages of finite treatment duration and sustained virologic response, but its efficacy remains relatively low. This study aimed to screen for IFN‐α subtypes with the highest anti‐HBV potency and to characterize mechanisms of IFN‐α–mediated HBV restriction.
Approach and Results
Using cell culture–based HBV infection systems and a human‐liver chimeric mouse model, IFN‐α subtype–mediated antiviral response and signaling activation were comprehensively analyzed. IFN‐α14 was identified as the most effective subtype in suppression of HBV covalently closed circular DNA transcription and HBV e antigen/HBV surface antigen production, with median inhibitory concentration values approximately 100‐fold lower than those of the conventional IFN‐α2. IFN‐α14 alone elicited IFN‐α and IFN‐γ signaling crosstalk in a manner similar to the combined use of IFN‐α2 and IFN‐γ, inducing multiple potent antiviral effectors, which synergistically restricted HBV replication. Guanylate binding protein 5, one of the most differentially expressed genes between IFN‐α14–treated and IFN‐α2–treated liver cells, was identified as an HBV restriction factor. A strong IFN‐α–IFN‐α receptor subunit 1 interaction determines the anti‐HBV activity of IFN‐α. The in vivo anti‐HBV activity of IFN‐α14 and treatment‐related transcriptional patterns were further confirmed, and few adverse effects were observed.
Conclusions
A concerted IFN‐α and IFN‐γ response in liver, which could be efficiently elicited by IFN‐α subtype 14, is associated with potent HBV suppression. These data deepen the understanding of the divergent activities of IFN‐α subtypes and the mechanism underlying the synergism between IFN‐α and IFN‐γ signaling, with implications for improved IFN therapy and HBV curative strategies.
The antibacterial peptide CM4 (ABP-CM4), isolated from Chinese Bombys mori, is a 35-residue cationic, amphipathic alpha-helical peptide that exhibits a broad range of antimicrobial activity. To explore a new approach for the expression of ABP-CM4 in E. coli, the gene ABP-CM4, obtained by recursive PCR (rPCR), was cloned into the vector pET32a to construct a fusion expression plasmid. The fusion protein Trx-CM4 was expressed in soluble form, purified by Ni(2+)-chelating chromatography, and cleaved by formic acid to release recombinant CM4. Purification of rCM4 was achieved by affinity chromatography and reverse-phase HPLC. The purified of recombinant peptide showed antimicrobial activities against E. coli K(12)D(31), Penicillium chrysogenum, Aspergillus niger and Gibberella saubinetii. According to the antimicrobial peptide database (http://aps.unmc.edu/AP/main.html), 116 peptides contain a Met residue, but only 5 peptides contain the AspPro site, indicating a broader application of formic acid than CNBr in cleaving fusion protein. The successful application to the expression of the ABP-CM4 indicates that the system is a low-cost, efficient way of producting milligram quantities of ABP-CM4 that is biologically active.
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