NSUN2-mediated m ⁵ C methylation of IRF3 mRNA negatively regulates type I interferon responses during various viral infections

Abstract

5-Methylcytosine (m 5 C) is a widespread post-transcriptional RNA modification and is reported to be involved in manifold cellular responses and biological processes through regulating RNA metabolism. However, its regulatory role in antiviral innate immunity has not yet been elucidated. Here, we report that NSUN2, a typical m 5 C methyltransferase, negatively regulates type I interferon responses during various viral infections, including SARS-CoV-2. NSUN2 specifically mediates m 5 C methylation of IRF3 mRNA and accelerates its degradation, resulting in low levels of IRF3 and downstream IFN-β production. Knockout or knockdown of NSUN2 enhanced type I interferon and downstream ISGs during various viral infection in vitro . And in vivo , the antiviral innate response is more dramatically enhanced in Nsun2 +/− mice than in Nsun2 +/+ mice. The highly m 5 C methylated cytosines in IRF3 mRNA were identified, and their mutation enhanced cellular IRF3 mRNA levels. Moreover, infection with Sendai virus (SeV), vesicular stomatitis virus (VSV), herpes simplex virus 1 (HSV-1), or Zika virus (ZIKV) resulted in a reduction of endogenous NSUN2 levels. Especially, SARS-CoV-2 infection (WT strain and BA.1 omicron variant) also decreased endogenous levels of NSUN2 in COVID-19 patients and K18-hACE2 KI mice, further increasing type I interferon and downstream ISGs. Together, our findings reveal that NSUN2 serves as a negative regulator of interferon response by accelerating the fast turnover of IRF3 mRNA, while endogenous NSUN2 levels decrease during SARS-CoV-2 and various viral infections to boost antiviral responses for effective elimination of viruses.

RNA m5C methylation modification: a potential therapeutic target for SARS-CoV-2-associated myocarditis

“…For EBV infection, NSUN2 catalyzes the m5C modification of EBER1, which impacts its metabolism and defense against the virus ( 102 ). Flaviviruses ( 103 ), hepatitis C virus (HCV) ( 104 ), Dengue Virus (DENV) ( 105 ), Zika Virus (ZIKV) ( 103 ) and others regulated by NSUN2; the methyl donor inhibitor A9145 has been found to inhibit the replication of DENV and ZIKV ( 105 ). It has been reported that the knockdown of NSUN2 does not significantly alter the m5C methylation of viral RNAs, but reduces the replication and gene expression of a variety of viruses (RSV, VSV, hMPV, SeV, and HSV) ( 22 ).…”

“…Through transcriptome sequencing of RNA isolated from bronchoalveolar lavage fluid (BALF) of two SARS-CoV-2 patients, NSUN2 mRNA level is found decreased in SARS-CoV-2 patients compared with that in healthy individuals. In comparison to the uninfected group, the infected mice with the SARS-CoV-2 WT strain or the K18-hACE2 KI mouse model with BA.1 omicron variant-activated innate immune response show significantly lower endogenous NSUN2 mRNA levels ( 103 ). These results could be also observed in the SARS-CoV-2-infected Caco-2 cell model and SARS-CoV-2 patients, suggesting that NSUN2 plays an important regulatory role in SARS-CoV-2 infection.…”

“…RNA m5C modification is crucial in the antiviral innate immune response of host cells ( 103 ) ( Figure 3 ). Kariko et al.…”

“…NSUN2 negatively regulates IFN-I responses during multiple viral infections, including SARS-CoV-2 infection ( 103 ). During infection by RNA viruses (RSV, VSV, hMPV, and SeV) and DNA viruses (HSV), NSUN2 alters m5C levels and either directly or indirectly regulates the RIG-I signaling pathway-mediated IFN-I response through downregulating levels of specific ncRNAs (particularly RPPH1 and 7SL RNAs), thereby enhancing the antiviral response ( 22 ).…”

“…During infection by RNA viruses (RSV, VSV, hMPV, and SeV) and DNA viruses (HSV), NSUN2 alters m5C levels and either directly or indirectly regulates the RIG-I signaling pathway-mediated IFN-I response through downregulating levels of specific ncRNAs (particularly RPPH1 and 7SL RNAs), thereby enhancing the antiviral response ( 22 ). In addition, NSUN2 specifically mediates m5C methylation of IRF3 mRNA and speeds up its degradation, resulting in lower levels of IRF3 and downstream IFN-β ( 103 ). NSUN2 knockout or knockdown can enhance the expression of IFN-I and downstream ISGs during various viral infections in vitro ( 103 ).…”