Post-transcriptional regulation of antiviral gene expression by N6-methyladenosine

Abstract

SUMMARY Type I interferons (IFNs) induce hundreds of IFN-stimulated genes (ISGs) in response to viral infection. Induction of these ISGs must be regulated for an efficient and controlled antiviral response, but post-transcriptional of these genes have not been well defined. Here, we identify a role for the RNA base modification N 6-methyladenosine (m 6 A) in the regulation of ISGs. Using ribosome profiling and quantitative mass spectrometry, coupled with m 6 A-immunoprecipitation and sequencing, we identify a subset of ISGs, including IFITM1, whose translation is enhanced by m 6 A and the m 6 A methyltransferase proteins METTL3 and METTL14. We further determine that the m 6 A reader YTHDF1 increases the expression of IFITM1 in an m 6 A-binding-dependent manner. Importantly, we find that the m 6 A methyltransferase complex promotes the antiviral activity of type I IFN. Thus, these studies identify m 6 A as having a role in post-transcriptional control of ISG translation during the type I IFN response for antiviral restriction.

FTO suppresses STAT3 activation and modulates proinflammatory interferon-stimulated gene expression

“…To test this hypothesis, we used siRNAs to deplete METTL3/14 or FTO in Huh7 cells and induced the expression of ISGs by treatment with IFN-β, a type I IFN. Similar to our previous results, depletion of METTL3/14 resulted in reduced protein levels of IFITM1, GBP1, IFIT3, and MX1 [5]. Depletion of FTO showed the opposite result in that its depletion resulted in increased protein expression of the METTL3/14-regulated ISGs, expect for MX1 whose expression was only modestly affected by METTL3/14 depletion and not altered by FTO depletion (Figure 1A).…”

“…We next determined whether FTO depletion changed the mRNA induction of these ISGs. Interestingly, while METTL3/14 depletion did not impact the mRNA levels of these ISGs, as expected [5], FTO depletion did increase the mRNA levels of the regulated ISGs, including IFITM1, GBP1, and IFIT3 (Figure 1B). These results show that FTO negatively regulates specific ISGs at the mRNA level.…”

“…Having previously shown that the m 6 A-methyltransferase complex proteins METTL3/14 promote the translation of specific ISGs via the addition of m 6 A to these ISGs [5], we hypothesized that the major m 6 A-demethylase FTO would suppress the translation of these ISGs by removal of m 6 A. To test this hypothesis, we used siRNAs to deplete METTL3/14 or FTO in Huh7 cells and induced the expression of ISGs by treatment with IFN-β, a type I IFN.…”

“…However, whereas METTL3/14 promotes the translation of a subset of ISGs without affecting their mRNA levels [5], FTO regulates a distinct subset of ISGs at the mRNA level. By labeling nascent RNA, we found that FTO inhibits the transcription of these ISGs and that FTO-depleted cells are primed to respond to type I IFN.…”

“…Specifically, the functions of the transcription factor STAT3 in the type I IFN response are not clear, although it has been shown to have functions either in suppressing this response or promoting the expression of certain ISGs, likely in a cell type-specific fashion [4]. We previously established that the addition of the mRNA modification m 6 A to a subset of ISGs by the m 6 A-methyltransferase complex proteins METTL3 and METTL14 (METTL3/14) increases their translation by recruiting the m A-binding protein YTHDF1 and that this promotes an antiviral cellular state [5]. However, the role of the m 6 A demethylase FTO in the type I IFN response has not yet been described.…”