Ghost authors revealed: The structure and function of human N6 -methyladenosine RNA methyltransferases.

Despite the discovery of modified nucleic acids nearly 75 years ago, their biological functions are still being elucidated. N6 -methyladenosine (m6 A) is the most abundant modification in eukaryotic messenger RNA (mRNA) and has also been detected in non-coding RNAs, including long non-coding RNA, ribosomal RNA, and small nuclear RNA. In general, m6 A marks can alter RNA secondary structure and initiate unique RNA-protein interactions that can alter splicing, mRNA turnover, and translation, just to name a few. Although m6 A marks in human RNAs have been known to exist since 1974, the structures and functions of methyltransferases responsible for writing m6 A marks have been established only recently. Thus far, there are four confirmed human methyltransferases that catalyze the transfer of a methyl group from S-adenosylmethionine (SAM) to the N6 position of adenosine, producing m6 A: methyltransferase-like protein (METTL) 3/METTL14 complex, METTL16, METTL5, and zinc-finger CCHC-domain-containing protein 4. Though the methyltransferases have unique RNA targets, all human m6 A RNA methyltransferases contain a Rossmann fold with a conserved SAM-binding pocket, suggesting that they utilize a similar catalytic mechanism for methyl transfer. For each of the human m6 A RNA methyltransferases, we present the biological functions and links to human disease, RNA targets, catalytic and kinetic mechanisms, and macromolecular structures. We also discuss m6 A marks in human viruses and parasites, assigning m6 A marks in the transcriptome to specific methyltransferases, small molecules targeting m6 A methyltransferases, and the enzymes responsible for hypermodified m6 A marks and their biological functions in humans. Understanding m6 A methyltransferases is a critical steppingstone toward establishing the m6 A epitranscriptome and more broadly the RNome. This article is categorized under: RNA Interactions with Proteins and Other Molecules > Protein-RNA Recognition RNA Interactions with Proteins and Other Molecules > RNA-Protein Complexes RNA Interactions with Proteins and Other Molecules > Protein-RNA Interactions: Functional Implications.

Elucidating the Kinetic Mechanism of Human METTL16.

Methyltransferase-like protein 16 (METTL16) is one of four catalytically active, S-adenosylmethionine (SAM)-dependent m6A RNA methyltransferases in humans. Well-known methylation targets of METTL16 are U6 small nuclear RNA (U6 snRNA) and the MAT2A mRNA hairpins; however, METTL16 binds to other RNAs, including the 3' triple helix of the metastasis-associated lung adenocarcinoma transcript 1 (MALAT1). Herein, we investigated the kinetic mechanism and biochemical properties of METTL16. METTL16 is a monomer in complex with either the MALAT1 triple helix or U6 snRNA and binds to these RNAs with respective dissociation constants of 31 nM and 18 nM, whereas binding to the methylated U6 snRNA product is 1.1 µM. The MALAT1 triple helix, on the other hand, is not methylated by METTL16 under in vitro conditions. Using the U6 snRNA to study methylation steps, preincubation and isotope partitioning assays indicated an ordered-sequential mechanism, whereby METTL16 binds U6 snRNA before SAM. The apparent dissociation constant for the METTL16·U6 snRNA·SAM ternary complex is 126 µM. Steady-state kinetic assays established a kcat of 0.07 min-1, and single-turnover assays established a kchem of 0.56 min-1. Furthermore, the methyltransferase domain of METTL16 methylated U6 snRNA with an apparent dissociation constant of 736 µM and a kchem of 0.42 min-1, suggesting that the missing vertebrate conserved regions weaken the ternary complex but do not induce any rate-limiting conformational rearrangements of the U6 snRNA. This study helps us to better understand the catalytic activity of METTL16 in the context of its biological functions.

Naturally occurring modified ribonucleosides.

The chemical identity of RNA molecules beyond the four standard ribonucleosides has fascinated scientists since pseudouridine was characterized as the "fifth" ribonucleotide in 1951. Since then, the ever-increasing number and complexity of modified ribonucleosides have been found in viruses and throughout all three domains of life. Such modifications can be as simple as methylations, hydroxylations, or thiolations, complex as ring closures, glycosylations, acylations, or aminoacylations, or unusual as the incorporation of selenium. While initially found in transfer and ribosomal RNAs, modifications also exist in messenger RNAs and noncoding RNAs. Modifications have profound cellular outcomes at various levels, such as altering RNA structure or being essential for cell survival or organism viability. The aberrant presence or absence of RNA modifications can lead to human disease, ranging from cancer to various metabolic and developmental illnesses such as Hoyeraal-Hreidarsson syndrome, Bowen-Conradi syndrome, or Williams-Beuren syndrome. In this review article, we summarize the characterization of all 143 currently known modified ribonucleosides by describing their taxonomic distributions, the enzymes that generate the modifications, and any implications in cellular processes, RNA structure, and disease. We also highlight areas of active research, such as specific RNAs that contain a particular type of modification as well as methodologies used to identify novel RNA modifications. This article is categorized under: RNA Processing > RNA Editing and Modification.