Pathogenic variants in the highly conserved OVOL2 promoter region cause posterior polymorphous corneal dystrophy (PPCD) 1 by inducing an ectopic expression of the endothelial OVOL2 mRNA. Here we produced an allelic series of Ovol2 promoter mutations in the mouse model including the heterozygous c.-307T>C variant (RefSeq NM_021220.4) causing PPCD1 in humans. Despite the high evolutionary conservation of the Ovol2 promoter, only some alterations of its sequence had phenotypic consequences in mice. Four independent sequence variants in the distal part of the Ovol2 promoter had no significant effect on endothelial Ovol2 mRNA level or caused any ocular phenotype. In contrast, the mutation c.-307T>C resulted in increased Ovol2 expression in the corneal endothelium. However, only a small fraction of adult mice c.-307T>C heterozygotes developed ocular phenotypes such as irido-corneal adhesions, and corneal opacity. Interestingly, phenotypic penetrance was increased at embryonic stages. Notably, c.-307T>C mutation is located next to the Ovol1/Ovol2 transcription factor binding site. Mice carrying an allele with a deletion encompassing the Ovol2 binding site c.-307_-320del showed significant Ovol2 gene upregulation in the cornea endothelium and exhibited phenotypes similar to the c.-307T>C mutation. In conclusion, although the mutations c.-307T>C and -307_-320del lead to a comparably strong increase in endothelial Ovol2 expression as seen in PPCD1 patients, endothelial dystrophy was not observed in the mouse model, implicating species-specific differences in endothelial cell biology. Nonetheless, the emergence of dominant ocular phenotypes associated with Ovol2 promoter variants in mice implies a potential role of this gene in eye development and disease.

Czech Centre for Phenogenomics and Laboratory of Transgenic Models of Diseases Institute of Molecular Genetics of the CAS Prumyslová 595 252 50 Vestec Czech Republic

Department of Ophthalmology 1st Faculty of Medicine Charles University Prague and General University Hospital Prague U Nemocnice 2 128 08 Prague 2 Prague Czech Republic

Department of Paediatrics and Inherited Metabolic Disorders 1st Faculty of Medicine Charles University and General University Hospital Prague Ke Karlovu 2 121 08 Prague 2 Prague Czech Republic

Laboratory of Transcriptional Regulation Institute of Molecular Genetics of the Czech Academy of Sciences Videnska 1083 142 20 Prague 4 Prague Czech Republic

Program in Craniofacial Biology and Department of Orofacial Sciences University of California 513 Parnassus Avenue CA 94158 San Francisco United States

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