Mutations in endothelin 1 cause recessive auriculocondylar syndrome and dominant isolated question-mark ears

Abstract

Auriculocondylar syndrome (ACS) is a rare craniofacial disorder with mandibular hypoplasia and question-mark ears (QMEs) as major features. QMEs, consisting of a specific defect at the lobe-helix junction, can also occur as an isolated anomaly. Studies in animal models have indicated the essential role of endothelin 1 (EDN1) signaling through the endothelin receptor type A (EDNRA) in patterning the mandibular portion of the first pharyngeal arch. Mutations in the genes coding for phospholipase C, beta 4 (PLCB4) and guanine nucleotide binding protein (G protein), alpha inhibiting activity polypeptide 3 (GNAI3), predicted to function as signal transducers downstream of EDNRA, have recently been reported in ACS. By whole-exome sequencing (WES), we identified a homozygous substitution in a furin cleavage site of the EDN1 proprotein in ACS-affected siblings born to consanguineous parents. WES of two cases with vertical transmission of isolated QMEs revealed a stop mutation in EDN1 in one family and a missense substitution of a highly conserved residue in the mature EDN1 peptide in the other. Targeted sequencing of EDN1 in an ACS individual with related parents identified a fourth, homozygous mutation falling close to the site of cleavage by endothelin-converting enzyme. The different modes of inheritance suggest that the degree of residual EDN1 activity differs depending on the mutation. These findings provide further support for the hypothesis that ACS and QMEs are uniquely caused by disruption of the EDN1-EDNRA signaling pathway.

Figure 1

Craniofacial Features of ACS and IQME Individuals Families F1 (A), F2, (B), and F3 (C). The numbering of individuals refers to the pedigrees in Figure 2. In (A), right and left lateral CT scans of individual II:3 are presented (black arrows indicate proximal mandibular hypoplasia, and the white arrow indicates the thickened zygomatic process of the right temporal bone), and an intraoral view of individual II:4 depicts the bifid uvula with adjacent ectopic tissue.

Figure 2

EDN1 Mutations and Their Segregation in ACS- or IQME-Affected Families F1–F3 and Simplex Case S1 “EX” indicates those individuals submitted to exome sequencing. The relevant EDN1 genotype is indicated for all those individuals whose DNA was available for testing.

Figure 3

PreproEDN1 Domain Structure and Positions of Amino Acids Affected by Mutations In the diagram at the top, bigEDN1 is shown in two shades of gray (the mature 21 amino acid peptide is the darker shade). Open arrowheads indicate furin cleavage sites. A filled arrowhead indicates the ECE cleavage site. Red arrows indicate the positions of substitutions identified in this report. The lower part of the figure is a multiple-sequence alignment of big endothelins and surrounding furin cleavage sites from selected species. Sequences used in the alignment were human EDN1 (RefSeq NP_001946.3), mouse EDN1 (RefSeq NP_034234.1), zebrafish Edn1 (RefSeq NP_571594.1), human EDN3 (RefSeq NP_000105.1), and mouse EDN3 (RefSeq NP_031929.1). The numbering above the alignment refers to the 21 amino acid mature peptide. Residues underlined with a red bar are affected by mutations in human EDN1 (this report), zebrafish edn1, human EDN3, and mouse Edn3 (see text for details). Black bars under the alignment indicate the two four-residue furin recognition sites. SP stands for signal peptide.