Fig. 4: Drosophila Mettl1 methylates tRNA in gonads.
a Schematic diagram of m7G site-specific reduction and cleavage. Cleaved 3′ fragments were detected by northern blotting. b Northern blot of chemically-treated total RNAs from wild-type control (yw) and Mettl1-KO (Mettl1KO1) testes. The probe was designed at the 3′ end of TrpCCA. Arrowhead denotes the position of the cleaved 3′ fragment. c Northern blot of chemically-treated total RNAs from WT (FM7c) and Wh-KO (Wh56) testes. The probe was designed around the 3′ end of tRNA TrpCCA. Arrowhead denotes the position of the cleaved 3′ fragment. d TRAC-seq scheme using Drosophila gonads. e Mettl1-dependent m7G modified tRNAs identified in testes by TRAC-seq. f Sequence motif of m7G modification sites identified in testes by TRAC-seq. The arrowhead corresponds to the m7G site. g Representative plot of the cleavage score (difference between cleavage score of WT and Mettl1-KO) of tRNA CysGCA (Cys-GCA-2) between WT and Mettl1-KO, which showed the highest score difference by TRAC-seq. Pink shading represents the variable loop region of tRNA CysGCA. h Quantitative comparison of cleavage scores (CleavageScore) of identified m7G-modified tRNAs (n = 36) between WT and Mettl1-KO testes. yw is treated as a wild-type control. Two-sided P-value was calculated by the Mann–Whitney U-test are indicated. Central bands, upper and lower edges of box plots represent median, first and third quartiles, respectively. Upper and lower whiskers represent maximum, and minimum values. In our TRAC-seq, 3 biological replicates were set for each sample. Each northern blot was reproduced for three times (Fig. 4b, c). Source data of Fig. 4b, c, g and h are provided as a Source Data file.
