Translating the epitranscriptome

Abstract

RNA modifications are indispensable for the translation machinery to provide accurate and efficient protein synthesis. Whereas the importance of transfer RNA (tRNA) and ribosomal RNA (rRNA) modifications has been well described and is unquestioned for decades, the significance of internal messenger RNA (mRNA) modifications has only recently been revealed. Novel experimental methods have enabled the identification of thousands of modified sites within the untranslated and translated regions of mRNAs. Thus far, N⁶ -methyladenosine (m⁶ A), pseudouridine (Ψ), 5-methylcytosine (m⁵ C) and N¹ -methyladenosine (m¹ A) were identified in eukaryal, and to some extent in prokaryal mRNAs. Several of the functions of these mRNA modifications have previously been reported, but many aspects remain elusive. Modifications can be important factors for the direct regulation of protein synthesis. The potential diversification of genomic information and regulation of RNA expression through editing and modifying mRNAs is versatile and many questions need to be addressed to completely elucidate the role of mRNA modifications. Herein, we summarize and highlight some recent findings on various co- and post-transcriptional modifications, describing the impact of these processes on gene expression, with emphasis on protein synthesis. WIREs RNA 2017, 8:e1375. doi: 10.1002/wrna.1375 For further resources related to this article, please visit the WIREs website.

Figure 1

Schematic representation of mRNA editing and its effect on translation. Editing of pre‐mRNA transcripts can generate start codons (green) and stop codons (red) by insertions of nucleotides or by base conversions. Base conversions potentially remove stop codons causing a prolonged open reading frame (purple). mRNA editing in the coding sequences can lead to non‐synonymous codon substitutions (blue). In addition, editing within the coding sequences or in the 3′ UTR of the mRNA can induce alternative splicing (yellow) and altered mRNA stabilities (dashed frame), respectively. Insertions or deletions of nucleotides can cause a restoration or even a creation of an ORF (gray arrow). Edited mRNAs are subsequently subjected to translation and result in shortened/extended protein products (red and purple, respectively) or functionally altered proteins (blue) (E: exon; ORF: open reading frame).

Figure 2

The dynamics of the m⁶A methylome. A METTL3‐METTL14‐WTAP methyltransferase complex (blue) mediates adenosine‐to‐m⁶A conversion of mRNAs.95, 96, 97 Once deposited, m⁶A fulfills distinct functions dependent on its localization within a transcript and the reader proteins (green) interacting with the m⁶A mark (blue triangle). m⁶A positioned within UTR sequences stimulates translational initiation via YTHDF1 or YTHDF2.98, 99 Alternatively, an YTHDF2‐m6A interaction in the 3′ UTR also induces transfer of mRNAs to decay sites.4 Other reader proteins affect alternative splicing100, 101 or processing and nuclear export of mRNAs.102 Eraser proteins, i.e., FTO or ALKBH5 (red), dynamically demethylate m⁶As83, 84 (E: exon; I: intron).

Figure 3

Pseudouridylation directly affects ribosomal translation. (a) Uridine isomerization to Ψ in mRNAs is achieved by two independent mechanisms. Either H/ACA box snoRNAs guide the catalytically active pseudouridine synthase Cbf5/dyskerin to a cognate target sequence, or pseudouridine synthases directly modify a target RNA independent of guide RNAs. Thereby, a second hydrogen bond donor (d) is liberated at the non‐Watson‐Crick edge of Ψ, whereas the Watson–Crick edge is unchanged (a: hydrogen bond acceptor). (b) The pseudouridylation of stop codons leads to stop codon read‐through.123, 124 In more detail, ΨAG/ΨAA stop codons can be recognized by tRNA^Ser or tRNA^Thr, whereas ΨGA stop codons interact with tRNA^Tyr or with tRNA^Phe thereby competing with release factors. (c) Ψ interpretation by the elongating ribosome is not universally conserved. Whereas randomly pseudouridylated mRNAs yield higher protein levels in rabbit reticulocyte lysates, translational rates are reduced in wheat germ extracts and are nearly abolished in E. coli lysates.3 The extent of translational inhibition by single Ψs in bacteria depends on the position of Ψ within a codon (ref: unmodified mRNA).116