Keratin 12 missense mutation induces the unfolded protein response and apoptosis in Meesmann epithelial corneal dystrophy

Hum Mol Genet. 2016 Mar 15;25(6):1176-91. doi: 10.1093/hmg/ddw001.

Edwin H A Allen ¹, David G Courtney ², Sarah D Atkinson ², Johnny E Moore ³, Laura Mairs ², Ebbe Toftgaard Poulsen ⁴, Davide Schiroli ², Eleonora Maurizi ², Christian Cole ⁵, Robyn P Hickerson ⁵, John James ⁶, Helen Murgatroyd ⁷, Frances J D Smith ⁵, Carrie MacEwen ⁷, Jan J Enghild ⁸, M Andrew Nesbit ², Deena M Leslie Pedrioli ⁵, W H Irwin McLean ⁹, C B Tara Moore ¹⁰

Affiliations

1 School of Biomedical Sciences, University of Ulster, Coleraine BT52 1SA, Northern Ireland, UK, Centre for Dermatology and Genetic Medicine, Division of Biological Chemistry and Drug Discovery, School of Life Sciences, University of Dundee, Scotland DD1 5EH, UK.
2 School of Biomedical Sciences, University of Ulster, Coleraine BT52 1SA, Northern Ireland, UK.
3 School of Biomedical Sciences, University of Ulster, Coleraine BT52 1SA, Northern Ireland, UK, Cathedral Eye Clinic, Academy Street, Belfast BT15 1ED, UK.
4 Department of Molecular Biology and Genetics.
5 Centre for Dermatology and Genetic Medicine, Division of Biological Chemistry and Drug Discovery, School of Life Sciences, University of Dundee, Scotland DD1 5EH, UK.
6 Microscopy Facility, College of Life Sciences, University of Dundee, Dundee DD1 5EH, UK.
7 Department of Ophthalmology, Ninewells Hospital and Medical School, Dundee DD1 9SY, UK.
8 Department of Molecular Biology and Genetics, Interdisciplinary Nanoscience Center (iNANO) and Center for Insoluble Protein Structures (inSPIN), Science Park, Aarhus University, Aarhus, Denmark and.
9 Centre for Dermatology and Genetic Medicine, Division of Biological Chemistry and Drug Discovery, School of Life Sciences, University of Dundee, Scotland DD1 5EH, UK, t.moore@ulster.ac.uk w.h.i.mclean@dundee.ac.uk.
10 School of Biomedical Sciences, University of Ulster, Coleraine BT52 1SA, Northern Ireland, UK, t.moore@ulster.ac.uk w.h.i.mclean@dundee.ac.uk.

PMID: 26758872 DOI: 10.1093/hmg/ddw001

Abstract

Meesmann epithelial corneal dystrophy (MECD) is a rare autosomal dominant disorder caused by dominant-negative mutations within the KRT3 or KRT12 genes, which encode the cytoskeletal protein keratins K3 and K12, respectively. To investigate the pathomechanism of this disease, we generated and phenotypically characterized a novel knock-in humanized mouse model carrying the severe, MECD-associated, K12-Leu132Pro mutation. Although no overt changes in corneal opacity were detected by slit-lamp examination, the corneas of homozygous mutant mice exhibited histological and ultrastructural epithelial cell fragility phenotypes. An altered keratin expression profile was observed in the cornea of mutant mice, confirmed by western blot, RNA-seq and quantitative real-time polymerase chain reaction. Mass spectrometry (MS) and immunohistochemistry demonstrated a similarly altered keratin profile in corneal tissue from a K12-Leu132Pro MECD patient. The K12-Leu132Pro mutation results in cytoplasmic keratin aggregates. RNA-seq analysis revealed increased chaperone gene expression, and apoptotic unfolded protein response (UPR) markers, CHOP and Caspase 12, were also increased in the MECD mice. Corneal epithelial cell Apoptosis was increased 17-fold in the mutant cornea, compared with the wild-type (P < 0.001). This elevation of UPR marker expression was also observed in the human MECD cornea. This is the first reporting of a mouse model for MECD that recapitulates the human disease and is a valuable resource in understanding the pathomechanism of the disease. Although the most severe phenotype is observed in the homozygous mice, this model will still provide a test-bed for therapies not only for corneal dystrophies but also for other keratinopathies caused by similar mutations.

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