Fig. 1: RNA methylation machinery controls L1 retrotransposition.
From: L1 retrotransposons exploit RNA m6A modification as an evolutionary driving force
a A schematic of the L1 construct and an overview of the L1 retrotransposition assay using engineered human L1 construct. b Retrotransposition assay in HeLa cells treated with siRNA that targets METTL3, ALKBH5, or FTO. A nontargeting siRNA (siCtrl) was used as a control. c Retrotransposition assays performed by co-transfecting the pL1Hs expression cassette with the indicated m6A enzyme-expressing vectors into HeLa cells. d L1 retrotransposition assays were performed in ALKBH5, ALKBH5 catalytically inactive mutant (H204A), or AcGFP(control)-overexpressing cells. Cells treated with 50 μM stavudine (d4T) served as a reverse transcription negative control. (n = 3 independent samples, mean ± s.d., one-way ANOVA and Tukey’s multiple comparisons test; ****p < 0.0001, ***p < 0.001, **p < 0.01, in comparison to control, ns: not significant). e Immunoblot assay of lysates from pL1Hs-transfected HeLa cells treated with indicated siRNAs that target m6A enzymes. Vinculin served as a loading control. f, g Immunoblot assay using pL1Hs-expressing HeLa cells. AcGFP, ALKBH5, FTO, or ALKBH5H204A overexpression plasmids were co-transfected with pL1Hs. FH-AcGFP served as transfection control. HSP70 served as a loading control. The predicted molecular weight of FLAG-HA-tagged proteins are 34 kDa for FH-AcGFP, 51 kDa for FH-ALKBH5, and 65 kDa for FH-FTO. The immunoblot images (e–g) are representative of three independent experiments. Source data are provided as a Source data file.
