Structural insights into the RNA methyltransferase domain of METTL16

Agnieszka Ruszkowska,Milosz Ruszkowski,Zbigniew Dauter,Jessica A Brown

doi:10.1038/s41598-018-23608-8

Abstract

N6-methyladenosine (m6A) is an abundant modification in messenger RNA and noncoding RNAs that affects RNA metabolism. Methyltransferase-like protein 16 (METTL16) is a recently confirmed m6A RNA methyltransferase that methylates U6 spliceosomal RNA and interacts with the 3′-terminal RNA triple helix of MALAT1 (metastasis-associated lung adenocarcinoma transcript 1). Here, we present two X-ray crystal structures of the N-terminal methyltransferase domain (residues 1–291) of human METTL16 (METTL16_291): an apo structure at 1.9 Å resolution and a post-catalytic S-adenosylhomocysteine-bound complex at 2.1 Å resolution. The structures revealed a highly conserved Rossmann fold that is characteristic of Class I S-adenosylmethionine-dependent methyltransferases and a large, positively charged groove. This groove likely represents the RNA-binding site and it includes structural elements unique to METTL16. In-depth analysis of the active site led to a model of the methyl transfer reaction catalyzed by METTL16. In contrast to the major m6A methyltransferase heterodimer METTL3/METTL14, full-length METTL16 forms a homodimer and METTL16_291 exists as a monomer based on size-exclusion chromatography. A native gel-shift assay shows that METTL16 binds to the MALAT1 RNA triple helix, but monomeric METTL16_291 does not. Our results provide insights into the molecular structure of METTL16, which is distinct from METTL3/METTL14.

Highlights

Most m6A RNA marks are catalyzed by a heterodimeric ‘writer’ complex comprised of methyltransferase-like protein 3 and methyltransferase-like protein 14 (METTL3/METTL14), which methylates adenosine within the RRACH motif with no obvious structural preferences[10]
RNA MTases bind to SAM and RNA, transfer a methyl group from SAM to adenosine, and the methylated RNA and SAH products dissociate from the MTase (Fig. 1A)
The MTase domain of Methyltransferase-like protein 16 (METTL16) has a canonical Class I MTase structure, METTL16_291 has several unique structural regions: residues 1–78 that precede the Rossmann fold as well as residues 95–97 and 188–222 within the Rossmann fold (Fig. 1C,D). These structural elements unique to METTL16 likely contribute to its RNA substrate specificity that is distinctly different from the METTL3/METTL14 complex[6,7,10,11,12,22,23,24]

Summary

Introduction

Most m6A RNA marks are catalyzed by a heterodimeric ‘writer’ complex comprised of methyltransferase-like protein 3 and methyltransferase-like protein 14 (METTL3/METTL14), which methylates adenosine within the RRACH motif with no obvious structural preferences[10]. Preliminary studies suggest that METTL16 uses a combination of sequence and structure to recognize its RNA substrates[11]. MTases are structurally similar, they use a variety of mechanisms to recognize their targets, such as oligomerization and unique structural elements. These unique structural elements include variable loop regions within the Rossmann fold and auxiliary domains within and/or flanking the Rossmann fold[20,21,22]. We investigated the oligomeric state of METTL16 and its ability to bind to the MALAT1 RNA triple helix

Methods

Results

Conclusion