Genetically-modified X. laevisindicate distinct roles for prominin-1 and photoreceptor cadherin in outer segment morphogenesis and retinal dystrophy

Abstract

Mutations in prominin-1 (prom1) and photoreceptor cadherin (cdhr1) are associated with inherited retinal degenerative disorders such as retinitis pigmentosa, cone-rod dystrophy, and juvenile macular dystrophy. The proteins encoded by these genes are hypothesized to regulate photoreceptor outer segment disc morphogenesis, but their functions remain unknown. We used CRISPR/Cas9 to generate prom1-, cdhr1-, and prom1 + cdhr1-null X. laevis and then documented the effects of these mutations on photoreceptor structure and function. Prom1-null mutations resulted in dysmorphic photoreceptors comprised of overgrown and disorganized disc membranes. Cones were more severely affected than rods; outer segments were elongated and fragmented, and ERG response was impaired. Autofluorescent deposits in the outer segment layer of aging prom1-null animals indicate that secondary toxic effects to the retina or RPE drive retinal degeneration for this mutation, instead of direct effects on outer segment disc morphogenesis. Cdhr1-null photoreceptors did not appear grossly dysmorphic, but ultrastructural analysis revealed that some disc membranes were overgrown or aligned vertically within the plasma membrane. Prom1 + cdhr1-null mutants did not differ significantly from prom1-null mutants. Our results indicate that neither prom1 nor cdhr1 are necessary for outer segment disc membrane evagination or the membrane fusion event involved in disc sealing. Rather, they are necessary for higher-order organization of the nascent outer segment discs. Prom1 may align and reinforce interactions between the disc leading edges, a function more critical in cone photoreceptors for structural support. Cdhr1 may help to align nascent discs and maintain horizontal disc orientation prior to fusion.