Misexpression of the constitutive Rpgr(ex1-19) variant leads to severe photoreceptor degeneration

Mutations in the retinitis pigmentosa GTPase regulator (RPGR) gene are a frequent cause of X-linked retinitis pigmentosa. The RPGR transcript undergoes complex alternative splicing to express both constitutive (Rpgr(ex1-19)) and Rpgr(ORF15) variants. Both variants localize to photoreceptor connecting cilia and are believed to play roles in ciliary function. This study examined variability in isoform expression and tested whether the constitutive variant could substitute for Rpgr function in photoreceptors.

Rpgr(ex1-19) and Rpgr(ORF15) expression during retinal development were compared using immunoblot analysis and immunohistochemistry, and ciliary affinity in adult photoreceptors was assessed by protein fractionation. Transgenic mice expressing either the full-length Rpgr(ex1-19) or Rpgr(ORF15) variant were studied using light and electron microscopy and immunofluorescence imaging. The results were compared with those of wild-type and Rpgr(-/-) mice.

Rpgr expression undergoes dynamic temporal regulation during retinal development, and variants exhibit variability for ciliary localization in adult photoreceptors. Transgenic expression of both variants grossly exceeded endogenous Rpgr expression in photoreceptors. Although both variants exhibited normal ciliary localization, overexpression of the Rpgr(ex1-19) variant resulted in atypical accumulation of Rpgr in photoreceptor outer segments, abnormal photoreceptor morphology, and severe retinal degeneration.

The Rpgr isoform ratio in the adult retina is critical to photoreceptor integrity. The utilization of distinct Rpgr variants at different stages of photoreceptor maturation suggests independent roles in photoreceptor function. Finally, misexpression of Rpgr(ex1-19) causes retinal degeneration that is considerably more severe than that caused by Rpgr knockout but photoreceptors tolerate overexpression of Rpgr(ORF15) without evidence of degeneration.