METTL16: An RNA Methyltransferase That Wears Many Hats

Methyltransferase-like protein 16 (METTL16) is a recently characterized human RNA methyltransferase in the class I SAM-methyltransferase family. It methylates adenosine at the N⁶ position in a consensus sequence on U6 small nuclear RNA (U6 snRNA) and S-adenosylmethionine (SAM) synthetase (MAT2A) pre-mRNA to control the activity of these molecules. By regulating SAM homeostasis in a feedback mechanism, METTL16 also exerts functional control over cellular methylation in general, affecting even its more well-known family members, METTL3 and METTL14. METTL16 is currently being investigated for its association with various mRNA, non-coding RNA, long non-coding RNA (lncRNA), and ribosomal RNA (rRNA).¹

M6A is the most prevalent epigenetic marking in mRNA. The realm of N⁶-methyladenosine (m6A) modifications is orchestrated by adenosine methyltransferases (writers), proteins that recognize m6A (readers), and enzymes that demethylate m6A marks (erasers). M6A marking influences many areas of RNA processing, especially transcript stability/longevity, pre-mRNA splicing, and translation efficiency. METTL16 functions primarily as a writer that places over 80% of its m6A marks within introns or at intron/exon boundaries. However, as a regulator of SAM, METTL16 can also function as a reader to regulate the splicing of the SAM synthetase transcript.¹

METTL16’s Unique Form and Multiple Functions

METTL16 is a 562 amino acid protein of just under 64 kDa. From front to back, it contains a distinct N-terminal domain (NTD), a methyltransferase domain with characteristic Rossmann fold (MTD), and two C-terminal vertebrate conserved domains (VCR), separated from each other by a disordered region. The NTD secures RNA substrates to enable methylation by the MTD. The MTD binds SAM and RNA at specific internal sites to catalyze methylation. The MTD contains a unique, autocatalytically inhibitory K-loop structure that lessens its affinity for SAM under nominal circumstances. Even though METTL16 is broadly conserved, only vertebrates possess the VCR domains. These domains, in concert with their disordered hinge region, modulate U6 snRNA and MAT2A methylation by greatly increasing METTL16 binding.²

While METTL16 methylates the (underlined) adenosine at the consensus UACAGARAA sequence, proper RNA conformation at this site is also critically important. A recognition loop, transition region, and stem that produce a hairpin bulge with the target adenosine prominently exposed are essential for both the MAT2A and U6 snRNA methylations.²

While METTL16 has only a few confirmed targets, its regulation of SAM homeostasis profoundly affects all SAM-mediated methylation. When SAM is abundant in the cell, METTL16’s increased methylation of hairpins in MAT2A mRNA’s 3’UTR causes intron retention, transcript destabilization, and degradation. The interaction is relatively brief, but catalytically efficient. This limits the production of MAT2A protein and, consequently, SAM. However, when SAM supplies dwindle below a critical point, METTL16’s residence time at MAT2A mRNA hairpins lengthens, waiting for the arrival of the methyl-group donor. As the pertinent hairpins are proximal to critical splice sites for MAT2A, METTL16’s longer interaction effects proper splicing, producing more mature MAT2A mRNA and, subsequently, higher MAT2A protein production. As a result, more SAM is made. This splicing does not require active METTL16 methyltransferase activity.³

Independent of its methyltransferase function, METTL16 prevents DNA end resection in the nucleus and interacts with eukaryotic translation initiation factor 3a/b to promote efficient translation in the cytosol. Most intriguingly, METTL16’s interaction with lncRNA MALAT1 seems to indicate an ability to recognize triple RNA helical structures.³

A Complex Role in Disease

While METTL16 is absolutely required for embryogenesis, its contributions to disease remain to be resolved. Mutations have been found in METTL16 that contribute to the incidence and severity of colon cancer. Low expression of METTL16 correlates with ovarian and hepatocellular carcinoma, while high expression predicts a poor prognosis in breast cancer. METTL16 seems to bind and stabilize the triple helical configuration of MALAT1, without effecting its methylation. This enhanced stability may potentiate MALAT1’s oncogenic influence.⁴

The principal problem in sorting METTL16’s diverse functions is isolating its indispensable SAM regulation from the interactions it seems to have with a great number of other experimentally suggested (but unconfirmed) partners. Only further investigation can eventually remedy this issue.

References

1. Ruszkowska, A. (2021) METTL16, Methyltransferase-Like Protein 16: Current Insights into Structure and Function. Int. J. Mol. Sci. 22(4), 2176. Review. https://doi.org/10.3390/ijms22042176.
2. Satterwhite, E. R., & Mansfield, K. D. (2022). RNA methyltransferase METTL16: Targets and function. Wiley Interdisciplinary Reviews: RNA. 13(2), e1681. Review. https://doi.org/10.1002/wrna.1681.
3. Mermoud, J.E. (2022) The Role of the m6A RNA Methyltransferase METTL16 in Gene Expression and SAM Homeostasis. Genes. 13, 2312. Review. https://doi.org/10.3390/genes13122312.
4. Fazi, F. and Fatica, A. (2019) Interplay Between N6-Methyladenosine (m6A) and Non-coding RNAs in Cell Development and Cancer. Front. Cell Dev. Biol. 7. Mini Review. https://doi.org/10.3389/fcell.2019.00116.