Ellis-van Creveld syndrome (EvC) is a recessive disorder characterised by acromesomelic limb shortening, postaxial polydactyly, nail-teeth dysplasia and congenital cardiac defects, primarily caused by pathogenic variants in EVC or EVC2. Weyers acrofacial dysostosis (WAD) is an ultra-rare dominant condition allelic to EvC. The present work aimed to enhance current knowledge on the clinical manifestations of EvC and WAD and broaden their mutational spectrum. We conducted molecular studies in 46 individuals from 43 unrelated families with a preliminary clinical diagnosis of EvC and 3 affected individuals from a family with WAD and retrospectively analysed clinical data. The deleterious effect of selected variants of uncertain significance was evaluated by cellular assays. We identified pathogenic variants in EVC/EVC2 in affected individuals from 41 of the 43 families with EvC. Patients from each of the two remaining families were found with a homozygous splicing variant in WDR35 and a de novo heterozygous frameshift variant in GLI3, respectively. The phenotype of these patients showed a remarkable overlap with EvC. A novel EVC2 C-terminal truncating variant was identified in the family with WAD. Deep phenotyping of the cohort recapitulated 'classical EvC findings' in the literature and highlighted findings previously undescribed or rarely described as part of EvC. This study presents the largest cohort of living patients with EvC to date, contributing to better understanding of the full clinical spectrum of EvC. We also provide comprehensive information on the EVC/EVC2 mutational landscape and add GLI3 to the list of genes associated with EvC-like phenotypes.

Weyers acrofacial dysostosis (WAD) is a rare skeletal dysplasia, which is autosomal-dominant, and the clinical symptoms are presented as dental anomalies, polydactyly, nail dystrophy, and short physical stature. It is also termed "Curry‑Hall syndrome" and reported to be linked to genetic mutations mapped on chromosome 4p16, the region reported being commonly associated with a similar genetic syndrome, Ellis-van Creveld (EVC) syndrome. Most individuals with EVC have congenital heart abnormalities, most often atrial septal defects, unlike WAD. In this case, a 15‑year‑old girl presented with onychodystrophy and polydactyly observed in the hands and feet, microdontia, or agenesis of teeth, which were conical in shape, with a short stature. The patient had dystrophy of nails since birth, and physical growth in terms of height did not match the normal growth parameters with respect to age. The patient also had abnormal dentation with conical-shaped teeth, with the rest of the clinical presentations suggestive of WAD.

Ellis van Creveld syndrome and Weyers acrofacial dysostosis are two rare genetic diseases affecting skeletal development. They are both ciliopathies, as they are due to malfunction of primary cilia, microtubule-based plasma membrane protrusions that function as cellular antennae and are required for Hedgehog signaling, a key pathway during skeletal morphogenesis. These ciliopathies are caused by mutations affecting the EVC-EVC2 complex, a transmembrane protein heterodimer that regulates Hedgehog signaling from inside primary cilia. Despite the importance of this complex, the mechanisms underlying its stability, targeting and function are poorly understood. To address this, we characterized the endogenous EVC protein interactome in control and Evc-null cells. This proteomic screen confirmed EVC's main known interactors (EVC2, IQCE, EFCAB7), while revealing new ones, including USP7, a deubiquitinating enzyme involved in Hedgehog signaling. We therefore looked at EVC-EVC2 complex ubiquitination. Such ubiquitination exists but is independent of USP7 (and of USP48, also involved in Hh signaling). We did find, however, that monoubiquitination of EVC-EVC2 cytosolic tails greatly reduces their protein levels. On the other hand, modification of EVC-EVC2 cytosolic tails with the small ubiquitin-related modifier SUMO3 has a different effect, enhancing complex accumulation at the EvC zone, immediately distal to the ciliary transition zone, possibly via increased binding to the EFCAB7-IQCE complex. Lastly, we find that EvC zone targeting of EVC-EVC2 depends on two separate EFCAB7-binding motifs within EVC2's Weyers-deleted peptide. Only one of these motifs had been characterized previously, so we have mapped the second herein. Altogether, our data shed light on EVC-EVC2 complex regulatory mechanisms, with implications for ciliopathies.

Copy number variations (CNV) are thought to play an important role in causing human diseases, including congenital anomalies, psychiatric disorders, and intellectual disabilities. We report here a one-year-old boy presented to our clinic as developmental delay. He presented a birth weight of 4.5 kg, motor delay, mental retardation, mild hypertonia, and some dysmorphic features (mild frontal bossing, hypertelorism, epicanthus, concave nasal ridge, slightly sparse hair, short hands, and mild nail dysplasia). The brain MRI indicated brain abnormalities; the Gross Motor Function Measure-66 score was 23.37; the Gesell test result showed the development quotient was 50, suggesting mental retardation. Chromosomal microarray analysis showed an approximately 97 kb microdeletion at 4p16.2 (4p16.2 CNV), including part of EVC and EVC2 genes, which were associated with Ellis-van Creveld syndrome (EvC) and Weyers acrofacial dysostosis (WAD). This report suggests 4p16.2 microdeletion may be associated with multiple developmental abnormalities, including motor delay and mental retardation.