Heterozygous mutations in the C-terminal domain of COPA underlie a complex autoinflammatory syndrome
Selket Delafontaine, … , Jérôme Delon, Isabelle Meyts
Selket Delafontaine, … , Jérôme Delon, Isabelle Meyts
Published January 4, 2024
Citation Information: J Clin Invest. 2024;134(4):e163604. https://doi.org/10.1172/JCI163604.
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Heterozygous mutations in the C-terminal domain of COPA underlie a complex autoinflammatory syndrome

Abstract

Mutations in the N-terminal WD40 domain of coatomer protein complex subunit α (COPA) cause a type I interferonopathy, typically characterized by alveolar hemorrhage, arthritis, and nephritis. We described 3 heterozygous mutations in the C-terminal domain (CTD) of COPA (p.C1013S, p.R1058C, and p.R1142X) in 6 children from 3 unrelated families with a similar syndrome of autoinflammation and autoimmunity. We showed that these CTD COPA mutations disrupt the integrity and the function of coat protein complex I (COPI). In COPAR1142X and COPAR1058C fibroblasts, we demonstrated that COPI dysfunction causes both an anterograde ER-to-Golgi and a retrograde Golgi-to-ER trafficking defect. The disturbed intracellular trafficking resulted in a cGAS/STING-dependent upregulation of the type I IFN signaling in patients and patient-derived cell lines, albeit through a distinct molecular mechanism in comparison with mutations in the WD40 domain of COPA. We showed that CTD COPA mutations induce an activation of ER stress and NF-κB signaling in patient-derived primary cell lines. These results demonstrate the importance of the integrity of the CTD of COPA for COPI function and homeostatic intracellular trafficking, essential to ER homeostasis. CTD COPA mutations result in disease by increased ER stress, disturbed intracellular transport, and increased proinflammatory signaling.

Authors

Selket Delafontaine, Alberto Iannuzzo, Tarin M. Bigley, Bram Mylemans, Ruchit Rana, Pieter Baatsen, Maria Cecilia Poli, Daisy Rymen, Katrien Jansen, Djalila Mekahli, Ingele Casteels, Catherine Cassiman, Philippe Demaerel, Alice Lepelley, Marie-Louise Frémond, Rik Schrijvers, Xavier Bossuyt, Katlijn Vints, Wim Huybrechts, Rachida Tacine, Karen Willekens, Anniek Corveleyn, Bram Boeckx, Marco Baggio, Lisa Ehlers, Sebastian Munck, Diether Lambrechts, Arnout Voet, Leen Moens, Giorgia Bucciol, Megan A. Cooper, Carla M. Davis, Jérôme Delon, Isabelle Meyts

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Figure 4

COPAR1142X and COPAR1058C disrupt COPI integrity and impair anterograde ER-to-Golgi and retrograde Golgi-to-ER trafficking.

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COPAR1142X and COPAR1058C disrupt COPI integrity and impair anterograde ...
(A) Western blot analysis of FLAG, N-terminal COPA, COPB2, COPE, and β-actin antibody in whole-cell extract (input) or eluate of IP of HEK293T cells cotransfected with both WT or mutant COPA and WT or EV of COPB2 and COPE. IP was performed with an antibody against FLAG (IP COPA). (B) Quantification of COPB2 and COPE protein levels coimmunoprecipitated with COPA-FLAG (COPA IP) compared with the input signal, as observed in A. (C) Immunofluorescence analysis of PCI transport assay in 4 different control fibroblasts (1 representative control is shown), COPAR1142X fibroblasts, derived from A.I.1 and A.II.3, and COPAR1058C fibroblasts, derived from B.II.1, at 0 and 60 minutes. Scale bars: 10 μm. (D) Graphs represent quantification of the ER exit (top) and Golgi entry (bottom) of PCI, as shown in C. (E) Immunofluorescence analysis of CtxB transport assay in 4 different control fibroblasts, COPAR1142X fibroblasts, derived from A.I.1 and A.II.3, and COPAR1058C fibroblasts, derived from B.II.1. Analysis was performed 2 hours and 10 hours after exposure to CtxB. Scale bars: 10 μm. (F) Graphs represent quantification of the Golgi release (top) and ER entry (bottom) of CtxB, as observed in E. In B, D, and F, results are shown as mean ± SEM, and significance levels were calculated using 1-way (D and F) or 2-way (B) ANOVA (*P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001). Data are representative of 2–3 independent experiments. (G) Electron microscopy of COPAR1142X fibroblasts, derived from A.II.3, compared with fibroblasts from a healthy control. Original magnification, ×15,000; ×40,000 (bottom right); scale bars: 200 nm. The images demonstrate a fragmented and disorganized Golgi apparatus (blue box) and an accumulation of vesicles (red box) in the cytosol of COPAR1142X fibroblasts.