Mandibulofacial dysostosis with alopecia results from ETAR gain-of-function mutations via allosteric effects on ligand binding
Yukiko Kurihara, … , Jeanne Amiel, Hiroki Kurihara
Yukiko Kurihara, … , Jeanne Amiel, Hiroki Kurihara
Published January 13, 2023
Citation Information: J Clin Invest. 2023;133(4):e151536. https://doi.org/10.1172/JCI151536.
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Research Article Development Genetics

Mandibulofacial dysostosis with alopecia results from ETAR gain-of-function mutations via allosteric effects on ligand binding

Abstract

Mutations of G protein–coupled receptors (GPCRs) cause various human diseases, but the mechanistic details are limited. Here, we establish p.E303K in the gene encoding the endothelin receptor type A (ETAR/EDNRA) as a recurrent mutation causing mandibulofacial dysostosis with alopecia (MFDA), with craniofacial changes similar to those caused by p.Y129F. Mouse models carrying either of these missense mutations exhibited a partial maxillary-to-mandibular transformation, which was rescued by deleting the ligand endothelin 3 (ET3/EDN3). Pharmacological experiments confirmed the causative ETAR mutations as gain of function, dependent on ET3. To elucidate how an amino acid substitution far from the ligand binding site can increase ligand affinity, we used molecular dynamics (MD) simulations. E303 is located at the intracellular end of transmembrane domain 6, and its replacement by a lysine increased flexibility of this portion of the helix, thus favoring G protein binding and leading to G protein–mediated enhancement of agonist affinity. The Y129F mutation located under the ligand binding pocket reduced the sodium-water network, thereby affecting the extracellular portion of helices in favor of ET3 binding. These findings provide insight into the pathogenesis of MFDA and into allosteric mechanisms regulating GPCR function, which may provide the basis for drug design targeting GPCRs.

Authors

Yukiko Kurihara, Toru Ekimoto, Christopher T. Gordon, Yasunobu Uchijima, Ryo Sugiyama, Taro Kitazawa, Akiyasu Iwase, Risa Kotani, Rieko Asai, Véronique Pingault, Mitsunori Ikeguchi, Jeanne Amiel, Hiroki Kurihara

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Figure 1

A patient with MFDA harboring the EDNRA mutation p.E303K, that occurred postzygotically.

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A patient with MFDA harboring the EDNRA mutation p.E303K, that occurred ...
(A–F) Dysmorphic facial features and alopecia of the patient (see text for details). (G–J) High resolution craniofacial 3D CT-scans of the patient with MFDA (G and I) and an age-matched control (H and J). (G and H) Frontal views show hypertrophic, dysplastic, and asymmetric malar bones and an abnormal orbital floor in the patient (G) compared with the control (H). (I and J) Volume-rendering sagittal reconstructions of the temporomandibular articulations show a novel, tripartite temporo-mandibulo-malar articulation on the left side (red arrow) with increased malar bone volume (blue star) in the patient (I) compared with the control (J). (K and L) Oblique axial reconstructions of CT scans of the incus and malleus showing hypoplasia of the long process of the incus in the patient (K) compared with a control (L). (M) Next-generation sequencing reads showing mosaicism for the EDNRA variant c.907G>A (8%–11% of reads, in green) in the patient’s peripheral blood DNA. (N) Sanger sequencing of EDNRA amplified by PCR from leukocyte DNA of the patient and her parents, demonstrating low -evel mosaicism for the variant c.907G>A (green arrow) in the patient. (O) Sequencing of PCR products amplified from individual bacterial colonies, following transformation with cloned EDNRA PCR fragments from the patient, indicating 3/32 colonies (9 %) harboring the variant (green arrow) while 29 colonies harbored the WT allele (black arrow). Abbreviations: orb, orbital floor; mal, malar bone; cond, condyle of the mandible; ZPT, zygomatic process of the temporal bone; mall, malleus; incus lp, long process of the incus. An asterisk indicates structures that are dysmorphic compared with normal.