Specific sDMA Modifications on the RGG/RG Motif of METTL14 Regulate Its Function in AML

Protein arginine methylations are crucial post-translational modifications (PTMs) in eukaryotes, playing a significant regulatory role in diverse biological processes. Different forms of arginine methylations are known to co-exist within densely modified arginine-rich protein regions. However, pinpointing the precise pattern of arginine methylation and their functional implications have been challenging. In this study, we employed high-resolution mass spectrometry and in vitro enzyme reactions to uncover that the RGG/RG motifs located in the C-terminal region of METTL14 undergo both asymmetric and symmetric dimethylations (aDMA and sDMA). Specifically, PRMT5 catalyses the symmetric dimethylation of residues R425 and R445 within the extensively methylated region of METTL14, whereas PRMT1 demonstrates a broader substrate specificity. Our results indicate that mutating R425 and R445 to lysines reduces the catalytic efficiency of the METTL3:METTL14 complex, implying that sDMA modifications at these residues are important for MELL14’s function. Moreover, our findings underscore the role of sDMA modifications on METTL14 in regulating m6A-specific gene expression, including BRD4, SP1, and c-MYC, in acute myeloid leukaemia (AML) cell lines. Finally, the proliferation of AML cells is more sensitive to the combined inhibition of METTL3 and PRMT5 activity than inhibiting either alone. In summary, our study delineates the methylation landscape of the C terminal region of METTL14 and shows that sDMA modifications at residues R425 and R445 modulate the m6A deposition activity of the methyltransferase complex. These insights suggest that sDMA modifications in METTL14 could represent a promising therapeutic target for the treatment of AML.