Mediators of Inflammation (Jan 2023)

The Spliceosome Factor EFTUD2 Promotes IFN Anti-HBV Effect through mRNA Splicing

  • Pingping Hu,
  • Yuwen Li,
  • Wen Zhang,
  • Rui Liu,
  • Linya Peng,
  • Ruirui Xu,
  • Jinyuan Cai,
  • Hui Yuan,
  • Tiantong Feng,
  • Anran Tian,
  • Ming Yue,
  • Jun Li,
  • Wenting Li,
  • Chuanlong Zhu

DOI
https://doi.org/10.1155/2023/2546278
Journal volume & issue
Vol. 2023

Abstract

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Background/Aims. The underlying mechanisms of differential responsiveness to IFN remain poorly understood. The elongation factor Tu GTP binding domain-containing protein 2 (EFTUD2), involved in RNA splicing and pre-mRNA processing, was recognized as a novel innate immune regulator of the IFN antihepatitis C virus (HCV) effect. The aim of this study is to elucidate the mechanisms underlying EFTUD2’s involvement in IFN-mediated anti-HBV responses. Methods. Using a CRISPR/Cas9 gene-editing system, EFTUD2 single allele knockout HepG2.2.15 cells were constructed. Subsequently, the HBV biomarkers in EFTUD2+/– HepG2.2.15 cells and wild-type (WT) cells with or without IFN-α treatment were detected. And the EFTUD2-regulated genes were then identified using mRNA sequence. Selected gene mRNA variants and their proteins were examined by qRT-PCR and Western blotting. To confirm the effects of EFTUD2 on HBV replication and IFN-stimulated gene (ISG) expression, a rescue experiment in EFTUD2+/– HepG2.2.15 cells was performed by EFTUD2 overexpression. Results. IFN-induced anti-HBV activity was found to be restricted in EFTUD2+/– HepG2.2.15 cells. The mRNA sequence showed that EFTUD2 could regulate classical IFN and virus response genes. Mechanistically, EFTUD2 single allele knockout decreased the expression of ISG-encoded proteins, comprising Mx1, OAS1, and PKR (EIF2AK2), through mediated gene splicing. However, EFTUD2 did not affect the expression of Jak-STAT pathway genes. Furthermore, EFTUD2 overexpression could restore the attenuation of IFN anti-HBV activity and the reduction of ISG resulting from EFTUD2 single allele knockout. Conclusion. EFTUD2, the spliceosome factor, is not IFN-inducible but is an IFN effector gene. EFTUD2 mediates IFN anti-HBV effect through regulation of gene splicing for certain ISGs, including Mx1, OAS1, and PKR. EFTUD2 does not affect IFN receptors or canonical signal transduction components. Therefore, it can be concluded that EFTUD2 regulates ISGs using a novel, nonclassical mechanism.