Figure 7.

Transcriptome-wide identification of YTHDF2-binding targets in NK cells.(A) Top 10 GO clusters from GO analysis of differentially expressed genes from RNA-seq data. (B–D) Heat maps of differentially expressed genes between NK cells from Ythdf2^WT and Ythdf2^ΔNK mice from RNA-seq grouped by cell cycle and division (B), spindle and chromosome (C), and cell survival and NK cell function (D). (E) The m⁶A motif detected by the HOMER motif discovery toll with m⁶A-seq data. (F) Density distribution of the m⁶A peaks across mRNA transcriptome from m⁶A-seq data. (G) Overlapping analysis of genes identified by RNA-seq, m⁶A-seq, and RIP-seq. 12 upregulated and 17 downregulated differentially expressed transcripts bound by YTHDF2 and marked with m⁶A are listed on the right. (H) Distribution of m⁶A peaks and YTHDF2-binding peaks across the indicated transcripts by Integrative Genomics Viewer. (I and J) RIP using either an m⁶A (I) or a YTHDF2 antibody (J) followed by qPCR to validate the target genes identified by m⁶A-seq and RIP-seq. Rabbit IgG was used as control. Enrichment of indicated genes was normalized to the input level. (K–M) The mRNA half-life (t_1/2) of Mdm2 (K), Tardbp (L), and Crebzf (M) transcripts in NK cells from Ythdf2^WT and Ythdf2^ΔNK mice. Data represent two independent experiments. (N and O) IL-2–expanded NK cells were transfected with Tardbp-specific siRNA cells under the stimulation of IL-15 (50 ng/ml). 3 d after transfection, cell proliferation or apoptosis were analyzed by Ki67 staining or annexin V staining, respectively, followed by a flow cytometric analysis (n = 3 per group, at least two independent experiments). Data are shown as mean ± SD and were analyzed by unpaired two-tailed t test (I and J) or one-way ANOVA (N and O). *, P < 0.05; **, P < 0.01; ***, P < 0.001.