Extended Data Fig. 5. The direct interaction between METTL16 and eIF3a/b in HEK293T cells.

a, The experimental scheme for Far-Western blotting showing how to detect the potential direct interaction between METTL16 protein and eIF3 family members. Note: the * indicates the C-term truncated eIF3 family member. b, Agarose gel electrophoresis of the RNA samples recovered from Co-IP samples with or without RNase digestion. Samples of eIF3a were used for Extended Data Fig. 5c and d; while samples of eIF3b were used for Extended Data Fig. 5e and f. c, d, Co-IP (c) and reciprocal Co-IP (d) showing the potent interaction between eIF3a and METTL16 is RNA independent. Representative images from two biological replicates with similar results were presented. e, f, Co-IP (e) and reciprocal Co-IP (f) showing the robust association between eIF3b and METTL16 is also RNA independent. Representative images from two biological replicates with similar results were displayed. g, Western blotting of Co-IP assays to test the binding of METTL16 wild type (WT) and mutants (F187G, PP185/186AA, and R200Q) with eIF3a and eIF3b. The 3 × Flag tag was fused to the N-terminal of WT and mutant METTL16 proteins. Representative images from two independent Co-IP assays with similar results were presented.