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Interactions, localization, and phosphorylation of the m6A generating METTL3–METTL14–WTAP complex.
RNA 24, 499-512 (2018)
N6-methyladenine (m6A) is found on many eukaryotic RNAs including mRNAs. m6A modification has been implicated in mRNA stability and turnover, localization, or translation efficiency. A heterodimeric enzyme complex composed of METTL3 and METTL14 generates m6A on mRNAs. METTL3/14 is found in the nucleus where it is localized to nuclear speckles and the splicing regulator WTAP is required for this distinct nuclear localization pattern. Although recent crystal structures revealed how the catalytic MT-A70 domains of METTL3 and METTL14 interact with each other, a more global architecture including WTAP and RNA interactions has not been reported so far. Here, we used recombinant proteins and mapped binding surfaces within the METTL3/14-WTAP complex. Furthermore, we identify nuclear localization signals and identify phosphorylation sites on the endogenous proteins. Using an in vitro methylation assay, we confirm that monomeric METTL3 is soluble and inactive while the catalytic center of METTL14 is degenerated and thus also inactive. In addition, we show that the C-terminal RGG repeats of METTL14 are required for METTL3/14 activity by contributing to RNA substrate binding. Our biochemical work identifies characteristic features of METTL3/14-WTAP and reveals novel insight into the overall architecture of this important enzyme complex.
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Scopus SNIP
Web of Science
Times Cited
Times Cited
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4.490
0.940
128
202
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Publication type
Article: Journal article
Document type
Scientific Article
Keywords
M6a ; Methyltransferase ; Mettl14 ; Mettl3 ; Rna Modification; Gene-expression Regulation; Messenger-rna Methylation; N-6-adenosine Methylation; Nuclear-rna; Translation; Protein; N6-methyladenosine; Methyltransferase; Identification; Demethylase
Language
english
Publication Year
2018
HGF-reported in Year
2018
ISSN (print) / ISBN
1355-8382
e-ISSN
1469-9001
Journal
RNA
Quellenangaben
Volume: 24,
Issue: 4,
Pages: 499-512
Publisher
Cold Spring Harbor Laboratory Press
Publishing Place
Cold Spring Harbor
Reviewing status
Peer reviewed
Institute(s)
CF Monoclonal Antibodies (CF-MAB)
POF-Topic(s)
30201 - Metabolic Health
Research field(s)
Helmholtz Diabetes Center
PSP Element(s)
G-502210-001
WOS ID
WOS:000428085200006
Scopus ID
85044966647
PubMed ID
29348140
Erfassungsdatum
2018-03-20