Fig 1: Splicing dysregulation of Mdm2 mRNA in CNS-specific Ddx20 cKO mice.A Western blotting for SMN, Gemin2, Ddx20, Gemin6, and α-Tubulin in Control and Nestin-Cre; Ddx20 cKO spinal cords at E13.5. α-Tubulin was used as a loading control. B RT-qPCR for components of the SMN complex in control and Nestin-Cre;Ddx20 cKO spinal cords at E14.5. Actb gene was used as an internal control. n = 3 mice per group. C RT-qPCR for spliceosomal U snRNAs in control and Nestin-Cre;Ddx20 cKO spinal cords at E14.5. 5S rRNA was used as an internal control. n = 3 mice per group. D Scatter plot represents the rate of exon inclusion in control and Nestin-Cre;Ddx20 cKO mRNA-seq significant hits. Each point indicates the mean, obtained from two biological replicates, for an individual alternative splicing event. E Sashimi plot showing the differential usage of Mdm2 exon 3 between control and Nestin-Cre;Ddx20 cKO mice. F Semi-quantitative RT-PCR for the alternative splicing of Mdm2 exon 3, in control and Nestin-Cre;Ddx20 cKO mice. n = 4 mice per group. Arrows indicate the Mdm2 mRNA primer recognition sites. G RT-qPCR for the exclusion of Mdm2 exon 3, in control and Nestin-Cre;Ddx20 cKO mice. Arrows indicate the Mdm2 mRNA primer recognition sites. n = 4 mice per group. Actb gene was used as an internal control. H Schematic of splice-modifying MOs, targeting the 5’ splice site of Mdm2 exon 3, and the induction strategy for in vivo Mdm2 splice-switching. MOs were injected into the ventricles of E14.5 mouse brains. Ninety-six hours after injection, the embryos were analyzed. I RT-PCR gel images showing the effective induction of Mdm2 exon 3 splice-skipping, in control and Mdm2 MO-injected brains, at E18.5. J RT-qPCR for the splice-skipping of Mdm2 exon 3, in control and Mdm2 MO-injected brains. n = 3 mice per group. Actb gene was used as an internal control. K Double-immunostaining for p53 and Olig2 in the cerebral cortex and ganglionic eminence of control and Mdm2 MO-injected brains. Yellow arrowheads indicate cells double-positive for p53 and Olig2. L RT-qPCR for p53-related genes in control and Mdm2 MO-injected brains. n = 3 mice per group. Actb gene was used as an internal control. Bar charts represent the mean ± SD. Statistical analysis was performed by two-tailed, unpaired t-test. *p < 0.05; **p < 0.01, ***p < 0.001, ****p < 0.0001; n.s., not significant.