Probing Proteostatic Stress in Degenerating Photoreceptors Using Two ComplementaryIn VivoReporters of Proteasomal Activity

Abstract

Inherited retinal degenerations originate from mutations in Ͼ300 genes, many of which cause the production of misfolded mutant photoreceptor proteins that are ultimately degraded by the ubiquitin-proteasome system (UPS). It was previously shown that rod photoreceptors in multiple mouse models of retinal degeneration suffer from proteostatic stress consisting of an insufficient cellular capacity for degrading UPS substrates. In this study, we focused on a specific UPS component required for the degradation of a subset of proteasome targets: the substrate-processing complex formed by the AAAϩ ATPase P97/VCP and associated cofactors. To assess whether P97 capacity may be insufficient in degenerating rods, we employed two complementary in vivo proteasomal activity reporters whose degradation is either P97-dependent or P97-independent. Retinal accumulation of each reporter was measured in two models of retinal degeneration: the transducin ␥-subunit knockout (G␥ 1-/-) and P23H rhodopsin knock-in (P23H) mice. Strikingly, the patterns of reporter accumulation differed between these models, indicating that the proteostatic stress observed in G␥ 1-/and P23H rods likely originates from different pathobiological mechanisms, in which UPS substrate degradation may or may not be limited by P97-dependent substrate processing. Further, we assessed whether P97 overexpression could ameliorate pathology in G␥ 1-/mice, in which proteostatic stress appears to result from P97 insufficiency. However, despite P97 overexpression being aphenotypic in other tissues, the ϳ2.4-fold increase in retinal P97 content was toxic to rods, which complicated the interpretation of the observed phenotype. Our results highlight the complexity of pathophysiological mechanisms related to degrading misfolded proteins in mutant photoreceptors.

Nonretinoid chaperones improve rhodopsin homeostasis in a mouse model of retinitis pigmentosa

“…Notably, RHO protein misfolding leads to dominant-negative effects, which include: (a) coaggregation of WT and mutant RHO in the endoplasmic reticulum (ER; refs. 14,15) leading to their degradation and shorter OS, (b) activated ER stress (16)(17)(18), and (c) a chronically overwhelmed proteolytic system (19,20). These factors are considered to contribute to the death of rod cells carrying class II RHO mutations.…”

Inhibition of VCP preserves retinal structure and function in autosomal dominant retinal degeneration

“…In these conditions, the dysfunction of VCP is considered to result in the deregulation of the proteostasis network (26, 31). Conversely, excessive ATPase activity of VCP is also pathogenic (32), resulting in mitochondrial fragmentation, cell death in neurons (33), and photoreceptor cell degeneration (34).…”

Retinal homeostasis and metformin-induced protection are not affected by retina-specific Pparδ knockout

“…The RNAseq datasets included ages P22, P25, P33, and P45 for P23H mice and P30 for the Gγ1 −/− mice. These ages cover times early through late degeneration in Rho P23H/+ , and mid-degeneration in Gγ1 −/− mice [ 20 , 27 ].…”

“…The libraries were prepared and sequenced by GENEWIZ. RNA-sequencing data was obtained from retinal tissue of control mice and Rho P23H /+ mice at P22, P25, P33, and P45 ages and from Gγ1 −/− mice [ 20 , 27 ] at P30. Sequencing data were mapped to the mm10 genome using the STAR RNA-seq aligner [ 28 ].…”