Fig. 3

ALKBH5-mediated m⁶A demethylation maintains RNA stability of MYH1G-AS. A, B Relative ALKBH5 expression in PEM and SOL of 7-week-old Xinghua chicken detect by RNA-seq (A) and qPCR (B). C–G Relative ALKBH5 expression (C), RNA dot blot assay (D), relative m⁶A methylation level (E), relative SELECT product at the 263 site of MYH1G-AS (F), and relative MYH1G-AS expression (G) after ALKBH5 overexpression or interference. H, I MYH1G-AS RNA stability assay after ALKBH5 overexpression (H) and inhibition (I). J Relative ALKBH5 expression after SMAD3 (J) and SP2 (K) overexpression or interference. L Left: Schematic of four truncated ALKBH5-promoter constructs used for luciferase assays. Right: Dual-luciferase reporter assays of four reporter constructs. M The transcriptional activity of the ALKBH5 core promoter region. N ChIP analysis of the binding capacity of SP2 to ALKBH5 promoter. O The transcriptional activity of ALKBH5 core promoter region after SP2 overexpression or knockdown. P–R RNA dot blot assay (P), relative m⁶A methylation level (Q), and relative SELECT product at the 263 site of MYH1G-AS (R) with SP2 overexpression or interference. Results are showed as mean ± SEM. In panels A–C, E–O, and Q, R statistical significance of differences between means was assessed using an independent sample t-test. (*P < 0.05; **P < 0.01; NS, no significant difference)