The CHD4-related syndrome: a comprehensive investigation of the clinical spectrum, genotype-phenotype correlations, and molecular basis

Abstract

Purpose: Sifrim-Hitz-Weiss syndrome (SIHIWES) is a recently described multisystemic neurodevelopmental disorder caused by de novo variants inCHD4. In this study, we investigated the clinical spectrum of the disorder, genotype-phenotype correlations, and the effect of different missense variants on CHD4 function.

Methods: We collected clinical and molecular data from 32 individuals with mostly de novo variants in CHD4, identified through next-generation sequencing. We performed adenosine triphosphate (ATP) hydrolysis and nucleosome remodeling assays on variants from five different CHD4 domains.

Results: The majority of participants had global developmental delay, mild to moderate intellectual disability, brain anomalies, congenital heart defects, and dysmorphic features. Macrocephaly was a frequent but not universal finding. Additional common abnormalities included hypogonadism in males, skeletal and limb anomalies, hearing impairment, and ophthalmic abnormalities. The majority of variants were nontruncating and affected the SNF2-like region of the protein. We did not identify genotype-phenotype correlations based on the type or location of variants. Alterations in ATP hydrolysis and chromatin remodeling activities were observed in variants from different domains.

Conclusion: The CHD4-related syndrome is a multisystemic neurodevelopmental disorder. Missense substitutions in different protein domains alter CHD4 function in a variant-specific manner, but result in a similar phenotype in humans.

Keywords: 12p13.31; ATPase; chromatin remodeling; intellectual disability; missense.

Figure 1

A: CHD4 domains and the location of variants identified in this cohort. Each variant is marked by an asterisk (*). Dark grey asterisk represents a truncating variant at the marked position. 1B: A table demonstrating the amino acid positions and the clinical findings of CHD4 variants detected in 32 individuals. The variants in bold have not been previously published. The domain color corresponds to A. Clinical findings marked in grey represent patients with truncating variants. *Hypogonadism refers to undescended testis, micropenis or abnormal gonadotropin levels. 1C: 3D protein modeling of the different CHD4 domains. The domain color corresponds to A, and the variants are depicted in gold. Pathogenic variants were found in all highly conserved domains. The box shows that the Cys467 in the PHD2 domain is involved in Zinc binding (purple sphere).

Figure 2

A: The frequency of different clinical features in the CHD4-related syndrome cohort. The n refers to the number of individuals for which we had data on the specific feature. For instance, only 23 individuals in the cohort had a brain MRI and 29 had an echocardiogram. 2B: Histogram of types of brain and heart anomalies seen in individuals in this cohort.

Figure 3

A: Photographs of 17 individuals with the CHD4-related syndrome at different ages. Photo 14,19 and 23 was previously published in . Photo 8 and 17 were published in . 3B: A composite of 16 photographs of participants harboring variants within the SNF2 like domain compared to healthy controls. (FDNA Inc. USA)

Figure 4:

Radiometric ATPase assays were performed in the presence of recombinant nucleosome, naked DNA, or in the absence of CHD4 (no protein). The percent of ATP hydrolyzed was quantified for each mutant and then normalized to wild-type (WT) activity. Experimental data are presented as means with standard deviation. Individual data points are represented by black (nucleosome) or grey (naked DNA) squares. Three individual experiments from two individual purifications (n=6) were performed. The significance of observed differences in activity was assessed by two-way ANOVA with Dunnett’s multiple test correction. (*) indicates P values < 0.05.

Figure 5

Chromatin Remodeling Assay. A: Remodeling activity of CHD4 WT and mutants was assessed and quantified. 12, 6, and 3 nM enzyme was incubated with 12nM recombinant nucleosome in the presence of HhaI. Digested DNA fragments were analyzed on 6% native polyacrylamide gel. B: Quantitation of remodeling activity. Experimental data are presented as means with standard deviation. Three individual experiments from two individual purifications (N = 6) were performed. C: A two-way ANOVA with Dunnett’s multiple test correction was performed to assess the significance of observed differences in activity. Table shows the significant P values for each mutation and at each dose. Ns = not significant.