Mutation in retinoic acid receptor β DNA binding domain affects nuclear localization and transcriptional activity causing ocular coloboma

Authors

• VK Kalaskar
• RP Alur
• LK Li
• Y Sergeev
• T Cogliati
• BP Brooks

Abstract

Uveal coloboma is a potentially blinding congenital ocular malformation caused by failure of the optic fissure to close during the fifth week of human gestation. Human genetic screenings have implicated retinoic acid receptor beta (RARβ) gene mutations in causing severe ocular malformations and other syndromic phenotypes. Although the molecular mechanisms causing these multitude of phenotypes are not clear, in this study we elucidate some of the mechanisms underlying a novel mutation in RARβ gene in a family with mainly non-syndromic autosomal dominant uveal coloboma. Through custom capture sequencing of a large cohort of patients with uveal coloboma, we report identification of novel mutations in known coloboma-associated genes. The mutation in RARβ was a missense change in the highly conserved DNA binding domain and predicted to affect the DNA binding ability. Cell culture studies showed that the mutation resulted in lower protein levels with reduced transcriptional activity in the dual luciferase assay. Transfection studies revealed that the mutation partially impaired nuclear localization of the protein, and that mutant RARβ retained other retinoic X receptor (RXR) proteins in the cytoplasm. Functional studies in zebrafish showed that human RARβ mRNA rescued the coloboma phenotype caused by Morpholino knockdown of rarga gene, a zebrafish homolog of human RARβ. Our study reveals how subtle structural changes in RARβ that affect nuclear translocation lead to a non-syndromic uveal coloboma, limiting the phenotype to the eye, the organ most sensitive to retinoic acid signaling.

Scientific Focus Area: Genetics and Genomics

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