Fig 2: TgCPEB4?4:CPEB4 KOGT/+ mice, but not CPEB4KOGT/+ mice, show anatomical and behavioral alteration.a, Brain weight in 6-week-old control (n = 45), CPEB4 KOGT/+ (n = 25), TgCPEB4?4 (n = 13) and TgCPEB4?4:CPEB4 KOGT/+ (n = 6) mice and evolution of body weight of control (n = 74), CPEB4 KOGT/+ (n = 27), TgCPEB4?4 (n = 18) and TgCPEB4?4:CPEB4 KOGT/+ (n = 6) mice. b, Immunohistochemistry against anti-cleaved caspase-3 in Cx (n = 3 slices from six controls and six TgCPEB4?4 mice). Scale bars represent 250 µm. c, Striatal neuronal cell density in Control (n = 19) and TgCPEB4?4 mice (n = 5). d, Spine density (spines/µm) in cortical layers II/III of pyramidal neurons in CPEB4 KOGT/+ mice (n = 5 cells from three controls, and n = 5 cells from four CPEB4 KOGT/+ mice). e, Amplitude (pA) and frequency (Hz) of mEPSCs recorded from pyramidal neurons of the somatosensory Cx, in CPEB4 KOGT/+ mice (n = 13 cells from five controls, and n = 17 cells from six CPEB4 KOGT/+ mice). f, Ultrasonic calls of pups during 5 min after separation from their mothers as mean of data from postnatal days 6 and 12 in control (n = 36), CPEB4 KOGT/+(n = 22), TgCPEB4?4 (n = 17) and TgCPEB4?4:CPEB4 KOGT/+ (n = 4) pups. g, Stereotypical running represented as distance travelled (cm) in the periphery in the OF-test in control (n = 74), CPEB4 KOGT/+ (n = 25), TgCPEB4?4 (n = 19) and TgCPEB4?4:CPEB4 KOGT/+ (n = 6) mice. h, Time interacting with empty cage or an unfamiliar mouse during 10 min. Control (n = 40), CPEB4 KOGT/+ (n = 24), TgCPEB4?4 (n = 11) and TgCPEB4?4:CPEB4 KOGT/+ (n = 4) mice. a, One-way ANOVA followed by Games-Howell post hoc test. b, Two-sided Mann-Whitney-Wilcoxon test. c-e, Two-sided unpaired t-test. f, g, Kruskal-Wallis one-way ANOVA test. h, Two-sided Wilcoxon signed-rank test. Data are mean ± s.e.m. 95% CIs. n.s non-significative, *P < 0.05, **P < 0.01, ***P < 0.001.

Fig 3: mRNA and protein levels of CPEBs in Cx of idiopathic ASD individuals and features of CPEB4 mis-splicing.a, CPEB1-3 mRNA expression levels according to RNA-seq data (n = 63 for control, n = 43 for ASD). b, CPEB1-3 protein levels (n = 10) and c, CPEB4 protein levels (n = 20 for control and n = 19 for ASD). d, Diagram representing the alternative splicing events of CPEB4 by rMATS. Percent Spliced in (PSI) is shown under each event (n = 81 for CTRL and n = 82 ASD cortical prefrontal and temporal samples). e, CPEB4 exon 4 inclusion level (PSI) in all (left panel) and over 35-year-old (right panel) individuals, and f, percentage of each CPEB4 splicing isoform by vast-tools analysis of isoform-specific EEJs (exon-exon junctions). Percentage of each CPEB4 splicing isoform by g, Digital Droplet PCR and h, Absolute qRT-PCR, i, ?4/Ex4+ CPEB4 isoform ratio in Cx of idiopathic ASD cases (n = 11) and CTRL (n = 10) under 35-year-old. For gel source data, see Supplementary Figure 1. a, d-g, i, Two-sided Mann -Whitney-Wilcoxon test. b, h, Two-sided unpaired t-test. Box plots show median, 25th, 75th percentiles. Data are mean ± s.e.m. 95% CIs. *P < 0.05, **P < 0.01.

Fig 4: Supplemental characterization of TgCPEB4?4 mice.a, Breeding protocol to obtain TgCPEB4?4 mice. Number of mice and percentages of births observed and expected for the four experimental genotypes. b, Kaplan-Meier curve for cumulative survival (continuous line) and probability of developing cranial dysmorphology (dashed line), (n = 44 for control, n = 39 for TgCPEB4?4 mice). c, Evolution of mice body weight (grams). Males (continuous line), n = 25 controls, n = 9 TgCPEB4?4 mice. Females (dashed line), n = 26 control, n = 7 TgCPEB4?4 mice. d, ß-GAL nuclear staining in forebrain neurons from 1.5-month-old controls (n = 6) and TgCPEB4?4 mice (n = 4). Cx, cortex; St, striatum; Hipp, hippocampus; LV, lateral ventricle. Scale bars represent 250 µm. e, St CPEB4 immunohistochemistry shows cytoplasm pattern in control (n = 6), no staining in CPEB4 KO (n = 2) and overexpressing neurons in TgCPEB4?4 mice (n = 4). Scale bars represent 50 µm. f, Protein and g, mRNA expression levels of CPEB1-4 in forebrain at embryonic day 18 (n = 3) and Cx at 1.5 months (n = 6), 1 year (n = 4) and 2 years (n = 5) of control and TgCPEB4?4 mice. For gel source data, see Supplementary Figure 1. a, Pearson’s chi-squared test. c, f, g, Two-sided unpaired t-test. Data are mean ± s.e.m. 95% CIs. *P < 0.05, **P < 0.01, ***P < 0.001.

Fig 5: Enrichment in ASD-risk genes among CPEB1-4 binding transcripts whose poly(A)-tail is shortened in an HD mouse model with altered CPEBs.a, Experimental design of RIP from WT and HD mice (with altered CPEB1 and CPEB4, see methods). b, Percentage of CPE sequences in the 3’ UTR of total genome, brain genes and CPEB1 and CPEB4 binders from RIP experiment. c, Percentage of CPEB1- or CPEB4-only binders with shortened (red), lengthened (blue) or unaltered (purple) poly(A) tail. d, Symbol and gene names of CPEB4 binders in WT St with the most shortened poly(A)-tail (FC = -3.0) in HD mice. The last column indicates whether they are also CPEB1 binders (Y, yes; N, no). High-confidence ASD-risk genes (SFARI cat. 1–3) are highlighted in pink. e, Fold-change enrichment of high-confidence ASD genes (SFARI cat. 1–3 and cat. 1–2) in CPEB4 binders whose poly(A) tail is shortened in HD mice (FC = -3.0). Heatmaps of CPEB4/CPEB1 binders f, in SFARI ASD genes or removing intellectual disability genes (ASD only) and g, in weighted gene co-expression network analysis (WGCNA) modules involved in ASD. h, Fold change enrichment of percentage of CPE sequences and CPEB4 binders of ASD genes (SFARI cat. 1-2, n = 63) vs. total genome stratified by 5’UTR, 3’UTR, CDS, gDNA length and ratio neuronal/glial expression. b, e-g, One-sided Fisher’s exact test. c, Pearson’s chi-squared test. h, Statistical details in simulations in method section. **P < 0.01, ***P < 0.001.