The molecular language of RNA 5' ends: guardians of RNA identity and immunity

Abstract

RNA caps are deposited at the 5' end of RNA polymerase II transcripts. This modification regulates several steps of gene expression, in addition to marking transcripts as self to enable the innate immune system to distinguish them from uncapped foreign RNAs, including those derived from viruses. Specialized immune sensors, such as RIG-I and IFITs, trigger antiviral responses upon recognition of uncapped cytoplasmic transcripts. Interestingly, uncapped transcripts can also be produced by mammalian hosts. For instance, 5'-triphosphate RNAs are generated by RNA polymerase III transcription, including tRNAs, Alu RNAs, or vault RNAs. These RNAs have emerged as key players of innate immunity, as they can be recognized by the antiviral sensors. Mechanisms that regulate the presence of 5'-triphosphates, such as 5'-end dephosphorylation or RNA editing, prevent immune recognition of endogenous RNAs and excessive inflammation. Here, we provide a comprehensive overview of the complexity of RNA cap structures and 5'-triphosphate RNAs, highlighting their roles in transcript identity, immune surveillance, and disease.

Keywords: 5′ cap; CMTR1; CMTR2; IFIT; RIG-I; antiviral; capping; m7G; type I interferon.

Figure 1.

5′ RNA cap structures. (A) Structure of m7G cap linked to the 5′-5′ triphosphate bridge (or cap 0) and m6A methylation if the first transcribed nucleotide is an adenosine. Additionally, both the first (N1, cap 1) and the second (N2, cap 2) can be 2′-O-methylated. Enzymes involved in each of these modifications are illustrated in blue circles and methyl groups are marked in orange. (RNTM) RNA guanine-7 methyltransferase, (CAPAM) cap-specific adenosine N6-methyltransferase, and (CMTR) cap 2′-O-methyltransferase 1 and 2. (B) Tri-methylguanosine (TMG) cap structure. Methyl groups at the N2 position on the cap 0 structure are marked in orange. (C) Structure of the noncanonical RNA cap, diadenosine tetraphosphate (Ap₄ A). (D) Metabolite-derived cap structures. (Left to right) Nicotinamide adenine dinucleotide (NAD) caps can exist in the oxidized form, as NAD⁺ (shown), or in the reduced form, NADH. Flavin adenine dinucleotide (FAD) caps structures. UDP-glucose-derived caps include uridine diphosphate-glucose (UDP-Glc) and UDP-N-acetylglucosamine (UDP-GlcNAc).

FIGURE 2.

Innate immune sensors for 5′-end RNA recognition. (A) Functional domains of the human RLR-pathway receptors, RIG-I, MDA5, and LGP2 (top), and IFIT members, IFIT1 and IFIT5 (bottom). (CARD) Caspase activation and recruitment domains; (Hel) helicase domain; (CTD) C-terminal domain; (TRP) tetratricopeptide repeat motif. Numbers indicate residue position. (B) The RIG-I-like receptor RIG-I recognizes single-stranded RNA (ssRNA) or dsRNA containing 5′-PPP, or 5′-diphosphate ends (5′-PP). RIG-I can also bind m7G-RNAs that are unmethylated in positions N1 and N2. The other major RLR receptor MDA5 is specialized in recognizing long dsRNAs. Upon binding to viral RNAs, RIG-I and MDA5 signal through the mitochondrial-associated protein, MAVS. Next, the kinases TBK1 and IKKe are activated, promoting the nuclear translocation of the IRF3/7 and NFkB transcription factors, which drive expression of type I IFNs and proinflammatory cytokines.