The expanding field of non-canonical RNA capping: new enzymes and mechanisms

Abstract

Recent years witnessed the discovery of ubiquitous and diverse 5'-end RNA cap-like modifications in prokaryotes as well as in eukaryotes. These non-canonical caps include metabolic cofactors, such as NAD⁺/NADH, FAD, cell wall precursors UDP-GlcNAc, alarmones, e.g. dinucleotides polyphosphates, ADP-ribose and potentially other nucleoside derivatives. They are installed at the 5' position of RNA via template-dependent incorporation of nucleotide analogues as an initiation substrate by RNA polymerases. However, the discovery of NAD-capped processed RNAs in human cells suggests the existence of alternative post-transcriptional NC capping pathways. In this review, we compiled growing evidence for a number of these other mechanisms which produce various non-canonically capped RNAs and a growing repertoire of capping small molecules. Enzymes shown to be involved are ADP-ribose polymerases, glycohydrolases and tRNA synthetases, and may potentially include RNA 3'-phosphate cyclases, tRNA guanylyl transferases, RNA ligases and ribozymes. An emerging rich variety of capping molecules and enzymes suggests an unrecognized level of complexity of RNA metabolism.

Keywords: RNA processing; capping; non-canonical capping.

Figure 1.

(a) Canonical mechanisms of m⁷G cap formation in eukaryotes. (b) Alternative pathway in vesicular stomatitis virus. Orange ‘P’ symbolizes the phosphorus group, adenosine nucleoside is in green and pink, guanosine nucleoside is in pink-only, pink pentagon represents ribose. Yellow asterisks highlight the new emerging linkage.

Figure 2.

Nucleotide-enzyme, RNA-enzyme or nucleotide-amino acid covalently bound intermediates used during 5′ aminoacylation of RNA by different enzymes (3′ aminoacylation by RtcB). (a) RNA guanylyl transferase (GTase), (b) RNA/DNA ligase, (c) tRNA synthetase (LysU). (d) RNA phosphate cyclase (RtcA), (e) RNA splicing ligase (RtcB), (f) RNA-dependent RNA polymerase L protein from VSV.

Figure 3.

(a) Mechanism of ab initio NAD capping by RNAP. (b) Three possible pathways to produce NAD-RNA post-transcriptionally. (c) Known function of NMNATs in NAD synthesis. Symbols as described in figure 1.

Figure 4.

Pathways capping RNA with Np₄N cap. (a) RNAP ab initio capping. (b) Aminoacyl tRNA synthetase pathway mediating nucleotidyl transfer dependent on amino acid-AMP covalent intermediate. (i) Formation of an amino acid-AMP intermediate, (ii) charging of tRNA, (iii) production of Np₄A alarmone, (iv) Ap₄N capping of RNA. Symbols as described in figure 1.

Figure 5.

(a) Proposed mechanism for 5′ RNA capping with ADPR by transcription initiation by RNA polymerase. (b) Tpt1-mediated transfer of an internal RNA 2′-monophosphate (2′ p) to NAD+ to form a 2′- OH RNA, ADP-ribose 1″,2″ cyclic phosphate, and nicotinamide [36]. (c) Proposed reaction mechanism for Tpt1-dependent 5′ RNA capping with ADPR. (d) Mechanism of decapping of NADylated RNA by ADP ribosyl cyclase (CD38) producing ADPR-capped RNA. Symbols as described in figure 1.

Figure 6.

Mechanism of RNA ligation by ATP-dependent RNA ligases. Symbols as described in figure 1.

Figure 7.

(a) Mechanism of 5′ RNA/DNA adenylylation by RtcA. (b) Mechanism of RtcA 2′,3′ phosphate cyclization. Symbols as described in figure 1.

Figure 8.

Splicing mechanism of RtcB. Symbols as described in figure 1.

Figure 9.

Mechanism of 3′-5′ GTP (ppp-G) addition by tRNA guanylyl transferases. Symbols as described in figure 1.