Pre-mRNA modifications and their role in nuclear processing

Abstract

Background: Cellular non-coding RNAs are extensively modified post-transcriptionally, with more than 100 chemically distinct nucleotides identified to date. In the past five years, new sequencing based methods have revealed widespread decoration of eukaryotic messenger RNA with diverse RNA modifications whose functions in mRNA metabolism are only beginning to be known.

Results: Since most of the identified mRNA modifying enzymes are present in the nucleus, these modifications have the potential to function in nuclear pre-mRNA processing including alternative splicing. Here we review recent progress towards illuminating the role of pre-mRNA modifications in splicing and highlight key areas for future investigation in this rapidly growing field.

Conclusions: Future studies to identify which modifications are added to nascent pre-mRNA and to interrogate the direct effects of individual modifications are likely to reveal new mechanisms by which nuclear pre-mRNA processing is regulated.

Keywords: RNA-modifying enzymes; mRNA modification; pre-mRNA modification; splicing.

Figure 1.. Human nuclear mRNA modifying enzymes.

(A) Chemical structures of the modified nucleotides installed by known nuclear human mRNA modifying enzymes. (B) Illustration of the potential function of mRNA modifications in all stages of the mRNA life cycle if they are added co-transcriptionally to nascent pre-mRNA. Depicted are the human mRNA modifying enzymes that are nuclearlocalized PUS1 (Ψ), TRUB1 (Ψ), NSUN2 (m⁵C), TET (hm⁵C), TRMT10C (m¹A), METTL16 (m⁶A) and those that are chromatinassociated- PUS7 (Ψ), or interact with Pol II- METTL3 (m⁶A) suggesting that the modification is added to pre-mRNA.

Figure 2.. Molecular mechanisms by which pre-mRNA modifications regulate splicing.

RNA modifications (mod) located in pre-mRNA introns could: (A) alter snRNA-pre-mRNA interactions to change inclusion of an alternative exon (dark grey). U1 snRNA and the 5’ splice site are shown as an example, but modifications could similarily alter pre-mRNA interactions with the U2 or U6 snRNAs; (B) enhance interaction between a splicing factor and its regulatory binding site on the pre-mRNA to change splicing outcome; (C) stabilize or disrupt structures that occlude or expose the binding site of a splicing factor or accessibility of a splice site sequence to the snRNAs. SF-Splicing factor, mod-RNA modification, gold line-SF binding site. (D) Mechanisms by which m⁶A regulates splicing. Left to right. By modulating structure to increase accessibility of hnRNPC to its binding site in introns. YTHDC1 is a direct m⁶A reader that binds to m⁶A in exons, recruits splicing factor SRSF3 to the exon and inhibits SRSF10 binding to enhance exon inclusion. YT521-B (Drosophila YTHDC1) binds the intron of the Sex-lethal pre-mRNA along with the Sex-lethal protein itself to repress exon inclusion and promote female specific splicing. In SAM depleted conditions METTL16 binding to the 3’ UTR is stabalized and METTL16 is sufficient to promote splicing of the upstream intron.