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Review
. 2019 Jan 30;10(2):102.
doi: 10.3390/genes10020102.

Eukaryotic 5-methylcytosine (m⁵C) RNA Methyltransferases: Mechanisms, Cellular Functions, and Links to Disease

Katherine E Bohnsack  1 , Claudia Höbartner  2 , Markus T Bohnsack  3   4
Affiliations
Review

Eukaryotic 5-methylcytosine (m⁵C) RNA Methyltransferases: Mechanisms, Cellular Functions, and Links to Disease

Katherine E Bohnsack et al. Genes (Basel). .

Abstract

5-methylcytosine (m⁵C) is an abundant RNA modification that's presence is reported in a wide variety of RNA species, including cytoplasmic and mitochondrial ribosomal RNAs (rRNAs) and transfer RNAs (tRNAs), as well as messenger RNAs (mRNAs), enhancer RNAs (eRNAs) and a number of non-coding RNAs. In eukaryotes, C5 methylation of RNA cytosines is catalyzed by enzymes of the NOL1/NOP2/SUN domain (NSUN) family, as well as the DNA methyltransferase homologue DNMT2. In recent years, substrate RNAs and modification target nucleotides for each of these methyltransferases have been identified, and structural and biochemical analyses have provided the first insights into how each of these enzymes achieves target specificity. Functional characterizations of these proteins and the modifications they install have revealed important roles in diverse aspects of both mitochondrial and nuclear gene expression. Importantly, this knowledge has enabled a better understanding of the molecular basis of a number of diseases caused by mutations in the genes encoding m⁵C methyltransferases or changes in the expression level of these enzymes.

Keywords: 5-methylcytosine; RNA methyltransferase; RNA modification; epitranscriptome; gene expression; messenger RNA (mRNA); mitochondria; ribosome; transfer RNA (tRNA).

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
Mechanisms of C5-methylation of cytosine by m5C RNA methyltransferases. (a) Amino acid sequence alignment of regions forming the active sites of human m5C methyltransferases. Top: NSUN family of m5C RNA methyltransferases, bottom DNMT family containing DNMT2 as RNA methyltransferase and DNMT1 and DNMT3A/B as DNA methyltransferases. The conserved motifs IV and VI are boxed. The catalytic cysteine that forms a covalent bond with C6 of the target cytosine is marked with magenta background, and is located in motif VI in NSUN methyltransferases, and in motif IV in DNMT methyltransferases. (b) The catalytic mechanism is depicted in detail for NSUN6 (see text for description). (c) The active site in the crystal structure of NSUN6 with target RNA is presented, showing the arrangement and key contacts between the amino acids in the active site and the target cytosine (C72) in RNA (PDB 5WWS).
Figure 2
Figure 2
Schematic views of the positions of m5C modifications in cytoplasmic rRNAs. A cryo-EM structure of the human 80S ribosome is shown with the positions of 28S-m5C3761 and 28S-m5C4413, the enzymes that install them as well as key ribosomal features indicated. The ribosomal protein eL41 is highlighted in yellow, tRNAs in the P-site and E-site in blue and green, respectively, and an mRNA fragment in magenta. PTC – peptidyl transferase center; DS – decoding site; eB – eukaryotic inter-subunit bridge.
Figure 3
Figure 3
The m5C modifications in cytoplasmic and mitochondrial tRNAs. (a) Schematic secondary structure and three-dimensional L-shape structure of a tRNA with the positions of m5C modifications and the cognate methyltransferases responsible for installing them marked. The interaction sites of NSUN6 with the discriminator base and additional base pairs in the acceptor stem and the D-loop are indicated as observed by X-ray crystallography. (b) Chemical structures of m5C-containing base pairs in Watson-Crick orientation (with the D-stem G65-m5C49) and the reverse-Watson-Crick orientation of the Levitt base pair G15:m5C48.
Figure 4
Figure 4
Hypermodification of m5C34 in anticodon of cytoplasmic and mitochondrial tRNAs. (a) During the maturation of the cytoplasmic tRNALeu(CAA), an m5C34 modification is installed by NSUN2. The methylated RNA is spliced, followed by oxidation to hm5C and f5C, which occurs in the nucleus. The oxidation is catalyzed by ALKBH1. The RNA is exported to the cytoplasm, where the methyltransferase FTSJ1 installs an additional methyl group on the ribose 2’-OH to produce hm5Cm and f5Cm. (b) Structures of modified nucleotides at position 34 and the modification pathway. (c) Modification of mt-tRNAMet by NSUN3 and ALKBH1 occurs in the mitochondria. SAM = S-adenosylmethionine, SAH = S-adenosylhomocysteine, α-KG = alpha-ketoglutarate.
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