SPEN haploinsufficiency causes a neurodevelopmental disorder overlapping proximal 1p36 deletion syndrome with an episignature of X chromosomes in females

Abstract

Deletion 1p36 (del1p36) syndrome is the most common human disorder resulting from a terminal autosomal deletion. This condition is molecularly and clinically heterogeneous. Deletions involving two non-overlapping regions, known as the distal (telomeric) and proximal (centromeric) critical regions, are sufficient to cause the majority of the recurrent clinical features, although with different facial features and dysmorphisms. SPEN encodes a transcriptional repressor commonly deleted in proximal del1p36 syndrome and is located centromeric to the proximal 1p36 critical region. Here, we used clinical data from 34 individuals with truncating variants in SPEN to define a neurodevelopmental disorder presenting with features that overlap considerably with those of proximal del1p36 syndrome. The clinical profile of this disease includes developmental delay/intellectual disability, autism spectrum disorder, anxiety, aggressive behavior, attention deficit disorder, hypotonia, brain and spine anomalies, congenital heart defects, high/narrow palate, facial dysmorphisms, and obesity/increased BMI, especially in females. SPEN also emerges as a relevant gene for del1p36 syndrome by co-expression analyses. Finally, we show that haploinsufficiency of SPEN is associated with a distinctive DNA methylation episignature of the X chromosome in affected females, providing further evidence of a specific contribution of the protein to the epigenetic control of this chromosome, and a paradigm of an X chromosome-specific episignature that classifies syndromic traits. We conclude that SPEN is required for multiple developmental processes and SPEN haploinsufficiency is a major contributor to a disorder associated with deletions centromeric to the previously established 1p36 critical regions.

Keywords: 1p36; DNA methylome analysis; SPEN; X chromosome; distal 1p36 deletion syndrome; episignature; genotype-phenotype correlations; neurodevelopmental disorder; obesity; proximal 1p36 deletion syndrome.

Figure 1

Known critical regions at 1p36, deletions involving SPEN, and facial features of subjects with de novo truncating SPEN variants (A) Cartoon showing the distal and proximal del1p36 critical regions (red boxes) as defined by Wu et al. and Kang et al. and Jordan et al., respectively. Deletions shown by Kang et al. and Rudnik-Schöneborn et al. (estimated on the basis of the data provided) to characterize the phenotypes associated with the proximal 1p36 critical region are shown as orange bars. One of these deletions does not include RERE, and three of these deletions include SPEN (green bars). Previously reported deletions that overlap SPEN and at least one of the critical regions are shown as blue bars. Deletions that affect SPEN but do not include either the distal or proximal 1p36 critical regions are shown as black bars. (B) Facial features of subjects with truncating variants in SPEN. Note the occurrence of broad forehead with frontal bossing, bitemporal narrowing, wide set eyes, arched (childhood) and long bushy (adulthood) eyebrows, synophrys, dysplastic overfolded ears with uplifted ear lobe, wide and depressed nasal bridge, broad nose with prominent/bulbous nasal tip, anteverted nares, long philtrum with thick vermillion, high/narrow palate without cleft, and pointed/rounded chin. A detailed clinical characterization of affected subjects is reported in Table S2 and Supplemental notes.

Figure 2

SPEN is a critical gene for 1p36 deletion syndrome (A) Expression of SPEN during human neocortical development. The expression values of SPEN across cortical samples are grouped and sorted by developmental time points. (B) Scatterplot shows the distribution of Spearman’s correlation with NDD genes in cortical samples for all the genes expressed in human cortex. Dots represent individual genes. The dashed horizontal line at 2.4% indicates the top percentile among which the correlation between NDD genes and SPEN is ranked. (C) Expression of SPEN in ventricular radial glia (vRG) cells and intermediate progenitor cells (IPCs) at gestational week 10. Violin plot shows the median value (point). p value indicates expression difference (one-sided Wilcoxon rank-sum test). (D) Scatterplot shows the distribution of Spearman’s correlation with NDD genes in the vRG-to-IPC transition at gestational week 10 for all the genes expressed in neural progenitor cells. Dots represent individual genes. The dashed horizontal line at 0.46% indicates the top percentile among which the correlation between NDD genes and SPEN is ranked. (E) Scatterplot shows Spearman’s correlation with NDD genes in bulk cortical samples versus the vRG-to-IPC transition for all the genes within the 1p36 region that are expressed in both conditions. Dots represent individual genes. Dot size of a gene is proportional to the number of de novo loss-of-function mutations for the gene in NDDs. Red denotes SPEN with rank 1 and green denotes genes with ranks 2 to 50. Genes ranked within top 50 and harboring at least one de novo loss-of-function mutation are labeled along with the corresponding ranks shown in parentheses. GW, gestational week; IPCs, intermediate progenitor cells; mos, months; NCX, neocortex; NDD, neurodevelopmental disorder; pcw, postconceptional weeks; vRG, ventricular radial glia.

Figure 3

Female subjects with SPEN-truncating mutations exhibit an X chromosome-specific episignature (A) Samples from female individuals were compared to samples from healthy female control individuals processed alongside the SPEN-mutated samples (batch controls) and CpGs on the X chromosome were analyzed. The episignature was able to separate SPEN-truncated samples from control individuals as shown by hierarchical clustering (left) and multidimensional scaling (right) analyses. (B) The X chromosome episignature was able to differentiate between females with SPEN mutations and control samples but not between males with SPEN mutations and control samples. (C) Support vector machine-based methylation variant pathogenicity (MVP) scores showed that the X chromosome signature scored female samples differently from all the other tested samples.

Figure 4

The X chromosome episignature in females with SPEN-truncating mutations differs from subjects with other neurodevelopmental disorders Blue samples (75%), including the SPEN-mutated samples and batch controls, were used for training and red samples (25%) were used for testing. Classification of female samples showed that the identified episignature has high specificity, and no SPEN non-mutated samples have high scores.