Loss of YBX1 regulates apoptotic genes in an m⁶A manner in leukemia cells. (A) Volcano map showing differential expression of Ybx1 targets in WT and Ybx1^cKO leukemia-initiating cells (LICs). (B) GO enrichment analysis of terms enriched in differential pathways with significantly downregulated genes in Ybx1^cKO LICs. qRT-PCR validation of the effect of Ybx1 on the expression levels of (C) Myc, Lgals3, Bcl2l1, P4hb, Rps3, Birc6, Aatf, Tmbim6, Mcl1, Wtap, and Rbm7 and (D) Mxd4, Sival, Phf10, Dynll2, Dapk3, Bag1, Vapb, and Bax in WT and Ybx1^cKO leukemia cells. (E) Distribution pattern for m⁶A peaks in LICs from AML mice as determined by using m⁶A-seq. The distribution (density) across 5′ UTRs, CDS, and 3′ UTRs is identified by an m⁶A site plot. The m⁶A DRACH (D, A/G/U; R, A or G; H, A/C/U) motif was enriched (inset) (P = 10⁻³⁸). (F) Percentages of various RNA species with m⁶A modification for MeRIP-seq of leukemia cells from AML mice. (G) An integrative genomics viewer (IGV) tracks displaying higher enrichment of m⁶A peaks for Bcl2l1, Mcl1, and Wtap transcripts. (H) MeRIP-qPCR analysis of m⁶A enrichment of Bcl2l1, Mcl1, and Wtap. (I) SLAM-seq analysis showing the cumulative distributions of global transcript half-life (t_1/2) in WT and Ybx1^cKO LICs. (J) SLAM-seq analysis showing the comparable half-life of non-m⁶A and m⁶A-tagged transcripts in WT and Ybx1^cKO LICs. (Kolmogorov-Smirnov [K-S] test, P = 5 × 10⁻⁷). (K) The mRNA half-life (t_1/2) of Myc and Bcl2l1 mRNA as determined by SLAM-seq. Two-tailed Student t test: *P < .05; **P < .01; ***P < .001. ATP, adenosine triphosphate; IgG, immunoglobulin G; ncRNA, non-coding RNA; rRNA, ribosome RNA, snoRNA, small nucleolar RNA; snRNA, small nuclear RNA.