Hepatitis A virus-induced hsa-miR-146a-5p attenuates IFN-β signaling by targeting adaptor protein TRAF6

Abstract

Hepatitis A virus (HAV), a unique hepatotropic human picornavirus, is the causative agent of acute hepatitis A in humans. Some studies have shown that HAV antagonizes the innate immune response by disrupting interferon-beta (IFN-β) signaling by viral proteins. However, whether microRNAs (miRNAs), a class of non-coding RNAs, are involved in the antagonism of IFN-β induction upon HAV infection is still unclear. In this study, we investigated the effects and mechanisms by which HAV-induced miRNAs antagonize IFN-β signaling. A variety of analytical methods, including miRNA microarray, RT-qPCR, dual-luciferase reporter assay, and Western blotting, were performed using HAV-infected cells. The results indicated that HAV infection upregulates the expression of hsa-miR-146a-5p, which in turn partially suppresses the induction of IFN-β synthesis, thereby promoting viral replication. Mechanistically, TRAF6 (TNF receptor-associated factor 6), a key adaptor protein in the RIG-I/MDA5-mediated IFN-I signaling pathway, is targeted and degraded by hsa-miR-146a-5p. As TRAF6 is necessary for IFN-β induction, inhibition of this protein attenuates IFN-β signaling. Taken together, the results from this study indicated that HAV disrupts RIG-I/MDA5-mediated IFN-I signaling partially through the cleavage of the essential adaptor molecule TRAF6 via hsa-miR-146a-5p.

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Abbreviations

HAV:

Hepatitis A virus

miRNA:

microRNA

qRT-PCR:

Real-time reverse transcription PCR

MAVS:

Mitochondrial antiviral signaling protein

TNF:

Tumor necrosis factor

CPE:

Cytopathic effect

IFN-I:

Type I interferon

IFN-α/β:

Interferon α/β

PRRs:

Pattern-recognition receptors

TLR3:

Toll-like receptor 3

RIG-I:

Retinoic acid-inducible gene I

MDA5:

Oncorhynchus mykiss melanoma differentiation associated gene 5

IFNAR1/2:

Interferon-associated receptor 1/2

JAK/STAT:

Janus kinase/signal transducer and activator of tran-ions

ISG:

Interferon-stimulated gene

NEMO:

Nuclear factor-κB essential modulator

TRIM14:

Tripartite motif containing 14

TBK1:

TANK-binding kinase 1

IKKε:

IΚB kinase ε

IRF3:

Interferon regulatory factor 3

RANTES:

C-C motif chemokine ligand 5

KSHV:

Kaposi’s sarcoma associated herpesvirus

USP3:

Ubiquitin-specific peptidase 3

EV71:

Enterovirus 71

VSV:

Vesicular stomatitis virus

IRAK1/2:

Interleukin 1 receptor associated kinase 1/2

HSV:

Herpes simplex virus

IRF1:

Interferon regulatory factor 1

MOI:

Multiplicity of infection

ELISA:

Enzyme-linked immunosorbent assay

TCID50 :

Tissue culture infectious dose

Ct:

Cycle threshold

PBS:

Phosphate-buffered saline

RIN:

RNA integrity number

GAPDH:

Glyceraldehyde-3-phosphate dehydrogenase

RLR:

RIG-I-like receptors

NF-κB:

Nuclear factor-κB

TRIF:

TIR domain-containing adaptor inducing IFN-β

MAVS:

Mitochondrial antiviral signaling protein

Myd88:

Myeloid differentiation primary response gene 88

TRAF6:

TNF receptor associated factor 6

RIP3:

Receptor interacting serine/threonine kinase 3

TMEM173:

Transmembrane protein 173

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Acknowledgements

This study was supported by the Chinese Academy of Medical Science (CAMS) Innovation Fund for Medical Sciences (2017-I2M-3-022), the National Natural Science Foundation of China (NSFC grant no. 31500724), the Applied Basic Research Key Project of Yunnan (CN) (202001AS070046), and the Fund for Reserve Talents of Young and Middle-Aged Academic and Technical Leaders of Yunnan Province (2019HB043).

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Correspondence to Jiandong Shi or Yunzhang Hu.

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Mo, L., Zeng, Z., Deng, R. et al. Hepatitis A virus-induced hsa-miR-146a-5p attenuates IFN-β signaling by targeting adaptor protein TRAF6. Arch Virol 166, 789–799 (2021). https://doi.org/10.1007/s00705-021-04952-z

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