Germ cell-specific Ythdf2 deletion alters the expressions of specific genes modified with m⁶A during spermatogenesis   (A) Scatter plot showing the fold change of transcripts of testis tissue between 2-month-old controls and age-matched Ythdf2-vKO mice. Significantly downregulated and upregulated genes with a fold change greater than 1.5 (FDR<0.1) are shown in blue and red, respectively. Analysis was performed on two biological replicates. (B) qPCR validation of the dysregulated genes in Ythdf2-vKO mouse spermatogenesis. Genes used for analysis are expressed at the following specific stages: Un.S, undifferentiated spermatogonia; A1, type A1 spermatogonia; Prel, preleptotene spermatocytes; DS, diplotene spermatocytes; RS, round spermatids. Data are presented as the mean± SEM (n=3 for each genotype). ***P<0.001. (C) Gene ontology enrichment analysis of the upregulated genes in Ythdf2-vKO mice. The top 10 most enriched biological processes are shown. (D) The ratio of transcripts modified with m⁶A among the dysregulated genes in Ythdf2-vKO mice and the distribution of m⁶A sites were analyzed according to our previously reported m⁶A mRNA methylomes of mouse spermaogenic cells [14]. (E) MeRIP-qPCR analysis using anti-IgG and anti-m⁶A antibodies showed m⁶A enrichment in Tnfrsf12a, Syk, Plch2, Mast1, Inf2, Tspan4, Tubb3, Stard8, Sh2d7, Kl, and Prss40 in adult mouse testis. The relative m⁶A enrichment was normalized to the input. ***P<0.001.