Retinal Degeneration Slow (RDS) Glycosylation Plays a Role in Cone Function and in the Regulation of RDS·ROM-1 Protein Complex Formation

Abstract

Background: Mutations in retinal degeneration slow (RDS) lead to rod and cone-dominant retinal degeneration by mechanisms that remain unknown. Results: Knockin mice carrying unglycosylated RDS have reduced RDS levels and functional defects in cones. Conclusion: RDS glycosylation is important for cone but not rod function. Significance: Differential reliance on glycosylation may help explain why some RDS disease mutations target cones and others target rods.The photoreceptor-specific glycoprotein retinal degeneration slow (RDS, also called PRPH2) is necessary for the formation of rod and cone outer segments. Mutations in RDS cause rod and cone-dominant retinal disease, and it is well established that both cell types have different requirements for RDS. However, the molecular mechanisms for this difference remain unclear. Although RDS glycosylation is highly conserved, previous studies have revealed no apparent function for the glycan in rods. In light of the highly conserved nature of RDS glycosylation, we hypothesized that it is important for RDS function in cones and could underlie part of the differential requirement for RDS in the two photoreceptor subtypes. We generated a knockin mouse expressing RDS without the N-glycosylation site (N229S). Normal levels of RDS and the unglycosylated RDS binding partner rod outer segment membrane protein 1 (ROM-1) were found in N229S retinas. However, cone electroretinogram responses were decreased by 40% at 6 months of age. Because cones make up only 3-5% of photoreceptors in the wild-type background, N229S mice were crossed into the nrl ؊/؊ background (in which all rods are converted to cone-like cells) for biochemical analysis. In N229S/nrl ؊/؊ retinas, RDS and

The Interplay between Peripherin 2 Complex Formation and Degenerative Retinal Diseases

“…The higher order complexes are localized at the closed rim of discs indicating an important role for them in the membrane folding necessary to close the disc rim [25,74]. Biochemical evidence supported the idea that during the formation of Prph2/Rom1 complexes, intermolecular disulfide bonding between Prph2 and Rom1 are formed [72,75,76].…”

PRPH2/RDS and ROM-1: Historical context, current views and future considerations

“…Non-glycosylated PRPH2/RDS retains the ability to bind ROM-1, but the lack of glycosylation leads to a readily detectable downshift in apparent molecular weight for both PRPH2/RDS monomer and covalently-linked dimer on non-reducing SDS-PAGE gels. We find that under non-reducing conditions in the N229S retina, ROM-1 dimer also significantly downshifts suggesting it is covalently linked to the smaller N229S PRPH2/RDS (Stuck et al, 2015). As expected, ROM-1 monomer does not shift in size, since non-covalent interactions between N229S PRPH2/RDS and ROM-1 would be interrupted by the denaturing conditions of the gels.…”

“…We generated a knockin mouse model carrying the N229S mutation in the endogenous Rds locus (Stuck et al, 2015). Consistent with previous results, we found no adverse effect of non-glycosylated PRPH2/RDS in rods; rds N229S/N229S animals exhibited normal PRPH2/RDS and ROM-1 levels and no defects in retinal structure or function.…”

“…However, by six months of age, cone electroretinogram responses were decreased by ~40%. To study the biochemical defect in cones, the N229S animals were crossed onto the nrl −/− background, and we found that in cones expressing non-glycosylated PRPH2/RDS, PRPH2/RDS and ROM-1 levels were each reduced by ~60% (Stuck et al, 2015). These results suggest that PRPH2/RDS glycosylation is important for cone photoreceptors.…”

Tetraspanins Function as Regulators of Cellular Signaling

“…Interestingly, the photoreceptor-specific tetraspanin retinal degeneration slow (RDS) can also be glycosylated (Kedzierski et al, 1999; Conley et al, 2012). More recently, the function of RDS glycosylation was re-examined by expressing a glycosylation deficient version of RDS in mice, which identified differential functional outcomes in cones vs. rod photoreceptor cells (Stuck et al, 2015). Moreover, the authors determined that glycosylation regulates the formation of RDS complexes with another tetraspanin ROM-1, demonstrating that glycosylation can modulate tetraspanin complex formation.…”