- 作者: Dipayan Bose, Xiang Lin, Le Gao, Zhi Wei, Yonggang Pei & Erle S. Robertson
- 作者服務機構: 1.Department of Computer Science, New Jersey Institute of Technology, 07102, New Jersey, United States of America 2.Department of Otorhinolaryngology-Head and Neck Surgery, and Tumor Virology Program, Perelman School of Medicine, University of Pennsylvania, 19104, Philadelphia, PA, USA 3.School of Public Health and Emergency Management, Southern University of Science and Technology, 518055, Shenzhen, Guangdong, China
- 中文摘要:
- 英文摘要:
Background
Reactivation of Epstein Barr virus (EBV) leads to modulation of the viral and cellular epitranscriptome. N6-methyladenosine (m6A) modification is a type of RNA modification that regulates metabolism of mRNAs. Previous reports demonstrated that m6A modification affects the stability and metabolism of EBV encoded mRNAs. However, the effect of reactivation on reprograming of the cellular mRNAs, and how this contributes to successful induction of lytic reactivation is not known.
Methods
Methylated RNA immunoprecipitation sequencing (MeRIP-seq), transcriptomic RNA sequencing (RNA-seq) and RNA pull-down PCR were used to screen and validate differentially methylated targets. Western blotting, quantitative real-time PCR (RT-qPCR) and immunocytochemistry were used to investigate the expression and localization of different proteins. RNA stability and polysome analysis assays were used to detect the half-lives and translation efficiencies of downstream genes. Insertion of point mutation to disrupt the m6A methylation sites was used to verify the effect of m6A methylation on its stability and expression levels.
Results
We report that during EBV reactivation the m6A eraser ALKBH5 is significantly downregulated leading to enhanced methylation of the cellular transcripts DTX4 and TYK2, that results in degradation of TYK2 mRNAs and higher efficiency of translation of DTX4 mRNAs. This resulted in attenuation of IFN signaling that promoted progression of viral lytic replication. Furthermore, inhibition of m6A methylation of these transcripts led to increased production of IFN, and a substantial reduction in viral copy number, which suggests abrogation of lytic viral replication.
Conclusion
Our findings illuminate the significance of m6A modification in overcoming the innate immune response during EBV reactivation. We now report that during lytic reactivation EBV targets the RNA methylation system of the host to attenuate the innate immune response by suppressing the interferon signaling which facilitates successful lytic replication of the virus. - 中文關鍵字:
- 英文關鍵字: N6-methyladenosine, Epitranscriptome, Lytic reactivation, Innate Immune response, RNA immunoprecipitation