A recap of RNA recapping

Abstract

The N7-methylguanosine cap is a hallmark of the 5' end of eukaryotic mRNAs and is required for gene expression. Loss of the cap was believed to lead irreversibly to decay. However, nearly a decade ago, it was discovered that mammalian cells contain enzymes in the cytoplasm that are capable of restoring caps onto uncapped RNAs. In this review, we summarize recent advances in our understanding of cytoplasmic RNA recapping and discuss the biochemistry of this process and its impact on regulating and diversifying the transcriptome. Although most studies focus on mammalian RNA recapping, we also highlight new observations for recapping in disparate eukaryotic organisms, with the trypanosome recapping system appearing to be a fascinating example of convergent evolution. We conclude with emerging insights into the biological significance of RNA recapping and prospects for the future of this evolving area of study. This article is categorized under: RNA Processing > RNA Editing and Modification Translation > Translation Regulation RNA Processing > Capping and 5' End Modifications RNA Turnover and Surveillance > Regulation of RNA Stability.

Keywords: capping; cytoplasm; m7G cap; transcriptome complexity; translational control.

Figure 1.. Biochemistry of RNA recapping.

A. Model of recapping in mammals. Multiple decapping enzymes can convert unmethylated (GpppN) and/or methylated (m⁷GpppN) capped ends to mono- and diphosphate substrates for recapping. 5′-monophosphate RNA is phosphorylated by an ATP-dependent kinase to produce 5′-diphosphate RNA. Guanylation by the guanylyltransferase (GTase) domain of RNGTT yields the GpppN cap structure that is then methylated at the N7 position by the RNMT-RAMAC heterodimeric cap methyltransferase. The 5′-monophosphate RNA kinase and RNGTT bind to adjacent SH3 domains of NCK1, and RNMT-RAMAC is recruited to RNGTT through the C-terminal catalytic domain of RNMT. B. Model of recapping in trypanosomes. Trypanosome decapping enzymes producing both mono- and diphosphate ends from m⁷GpppN caps have been described. Conversion of 5′-monophosphate RNA to the GpppN cap structure is catalyzed by the bifunctional enzyme TbCE1, which has an N-terminal kinase domain and a C-terminal GTase domain. The methyltransferase TbCMT1 is proposed to catalyze N7-methylation.

Figure 2.. Translational control by cap homeostasis.

A. Polysome profiling shows inhibition of recapping by overexpression of a catalytically inactive, cytoplasmically restricted form of RNGTT (K294A) increases transcript accumulation in non-translating messenger ribonucleoprotein complexes (mRNPs, top panel). In the middle and lower panels individual fractions were assayed by RT-qPCR for recapping target and non-target mRNAs. Inhibition of cytoplasmic capping results in the redistribution of target mRNAs into non-translating mRNPs whereas non-target controls remain essentially unchanged. This figure is adapted from Figure 5 in (Mukherjee et al., 2012) and is reproduced here in accordance with Creative Commons License CC-BY. B. Model of cap homeostasis. Because the majority of translation initiation is cap-dependent, the translatability of an mRNA can be regulated by cycles of decapping and recapping. Such cap homeostasis is independent of changes in poly(A) tail length. Translationally silent mRNAs can be stored in stress granules (SGs) and P-bodies (PBs), though it remains to be elucidated how cap homeostasis intersects with RNA granule formation.

Figure 3.

Diversification of the transcriptome and proteome by recapping of 5′-end-processed RNAs.A. Possible avenues for the generation of 5′-end-processed RNA recapping substrates for recapping. New 5′ ends can be created by action of endoribonucleases (1) or by inhibition of XRN1 5′-exoribonuclease activity by strong secondary structure (2) or RNA-binding proteins (3). B. Consequences of recapping at downstream sites. A full-length mRNA with a cap at its canonical transcription start site is shown as (1). Recapping of an mRNA with a shortened 5′ UTR (2) can remove regulatory elements. Recapping within the CDS (3) may facilitate translation from downstream initiation codons, producing N-terminally truncated proteins. Noncoding RNAs with regulatory potential can be generated by recapping within the CDS downstream of alternative translation initiation codons (4) or recapping within the 3’ UTR (5).