Erythrokeratodermia variabilis et progressiva (EKV-P) is caused by mutations in either theGJB3 (Cx31) or GJB4 gene (Cx30.3). We identified a rare GJB3 missense mutation,c.134G>A (p.G45E), in two unrelated patients and investigated its cellular characteristics.Expression of Cx31G45E-GFP caused previously undescribed changes within HeLa cells andHaCaT cells, a model human keratinocyte cell line. Cx31WT-GFP localised to the plasmamembrane, but expression of Cx31G45E-GFP caused vacuolar expansion of the endoplasmicreticulum (ER), the mutant protein accumulated within the ER membrane and disassembly ofthe microtubular network occurred. No ER stress responses were evoked. Cx31WT-myc-myc-6xHis and Cx31G45E-GFP co-immunoprecipitated, indicative of heteromeric interaction, butco-expression with Cx31WT-mCherry, Cx26 or Cx30.3 did not mitigate the phenotype. Cx31and Cx31G45E both co-immunoprecipated with Cx43, indicating the ability to formheteromeric connexons. WT-Cx31 and Cx43 assembled into large gap junction plaques atpoints of cell to cell contact, Cx31G45E restricted the ability of Cx43 to reach the plasmamembrane in both HaCaT cells and HeLa cells stably expressing Cx43 where the proteinsstrongly co-localised with the vacolourised ER. Cell viability assays identified an increase incell death in cells expressing Cx31G45E-GFP, which FACS analysis determined wasnecrotic. Blocking connexin channel function with 18a-Glycyrrhetinic acid did notcompletely rescue necrosis or prevent propidium iodide uptake, suggesting that expression ofCx31G45E-GFP damages the cellular membrane independent of its channel function. Ourdata suggests that entrapment of Cx43 and necrotic cell death in the epidermis could underliethe EKV skin phenotype.
- cell death
- heteromeric connexins
Easton, J. A., Alboulshi, A. K., Kamps, M. A. F., Brouns, G. H. M. R., Broers, J. L. V., Coull, B. J., Oji, V., Geel, M. V., Steensel, M. A. M. V., & Martin, P. E. (2019). A rare missense mutation in GJB3 (Cx31G45E) is associated with a unique cellular phenotype resulting in necrotic cell death. Experimental Dermatology, 28(10), 1106-1113. https://doi.org/10.1111/exd.13542