WDR34 Deficiency Disrupts Retrograde Intraflagellar Transport and Induces Unfolded Protein Response-Driven Inflammation and Retinal Degeneration

Retinal degeneration in a mouse model of CRB1 disease rescued by the photoreceptor-specific CRB1-B isoform

Many genes involved in inherited diseases produce alternate mRNA isoforms that remain poorly characterized. Functional assessment of these isoforms could therefore unlock new insights into disease pathobiology or treatment. Here we investigated the function of the newly discovered “B” isoform of CRB1, a gene implicated in inherited retinal degenerations. CRB1-B is the most abundant retinal isoform, and differs from the canonical CRB1-A isoform in several important ways. Crucially, CRB1-B is the only isoform expressed by photoreceptors – the cells vulnerable to degeneration. Using a mouse model of CRB1-associated degeneration with degraded visual acuity, we find that restoring CRB1-B to photoreceptors prevents degeneration and completely rescues visual function, despite the absence of other isoforms. The therapeutic mechanism involves preservation of adherens junctions between photoreceptors and supporting glia: We establish that progressive loss of these junctions is a key pathobiological mechanism underlying photoreceptor death, and that junction loss is prevented by restoring photoreceptor expression of CRB1-B. Our findings nominate CRB1-B replacement as a promising gene therapy strategy. More broadly, they suggest that exploring disease gene isoform diversity offers an untapped opportunity to devise novel therapeutics.

A mutation in Bicdl1 (BICD family-like cargo adaptor 1) leads to mild outer retina thinning and late retinal dysfunction in a mouse model.

Single-cell spatial transcriptomics reveals rewiring of RPE and Müller glia signaling toward photoreceptors following outer segment disruption in Prph2C213Y mice.

PRPH2 mutations cause inherited retinal dystrophies (IRDs), but how photoreceptor outer segment (OS) disruption reshapes the surrounding retina remains unclear. Using a heterozygous Prph2C213Y/+ mouse model generated by CRISPR/Cas9, we characterized age-related retinal pathology and responses of retinal pigment epithelium (RPE) and Müller glia. Independent age- and sex-matched cohorts were examined at 1, 3, and 6 months by electroretinography, optical coherence tomography, and fundus autofluorescence. Mutant mice showed rod dysfunction from 1 month, RPE dysfunction from 3 months, and cone dysfunction by 6 months, accompanied by progressive outer retinal thinning and hyperautofluorescent deposits. Histological and ultrastructural analyses revealed OS disorganization, shortened RPE microvilli, RPE monolayer remodeling, increased RPE autofluorescence, and reactive Müller gliosis. Single-cell spatial transcriptomics of wild-type and mutant retinas at 6 months resolved nine cell populations and identified RPE cells and Müller glia as prominently perturbed non-photoreceptor populations. RPE cells showed an epithelial-mesenchymal transition-related remodeling state linked to a candidate Nfib-Fstl1 module, whereas Müller glia showed activation of activator protein 1 (AP-1) regulons, including Fos, Fosl2, and Junb, with predicted targets Osmr, A2m, and Stat3. Cell-cell communication analyses indicated coordinated changes in neuroprotective, inflammatory, and matrix-related signaling from RPE cells and Müller glia toward photoreceptors. These findings indicate that PRPH2-associated retinal dystrophy is a multicellular process in which OS disruption drives coordinated RPE and Müller glial remodeling with potentially protective or pro-degenerative effects, and nominate the RPE Nfib-Fstl1 program, Müller glial AP-1 responses with predicted STAT3 involvement, and support-cell-derived growth factor signaling as candidate mutation-independent therapeutic targets.

ADORA2A activation restores lysosomal function and photoreceptor outer segment degradation in stressed retinal pigment epithelium

Background Age-related macular degeneration (AMD) involves early retinal pigment epithelium (RPE) dysfunction and impaired processing of photoreceptor outer segments (POS). We investigated whether ADORA2A regulates post-ingestive POS handling and lysosomal recovery under AMD-relevant stress. Methods A2E-stressed ARPE-19 cells, primary porcine RPE cells, and a sodium iodate-induced mouse model were studied. ADORA2A was activated with CGS21680 and inhibited with ZM241385. POS handling, LC3B/Rubicon association with POS, lysosomal function, Rubicon depletion, chronic stress phenotypes, and retinal protection were assessed. Results A2E increased ADORA2A expression while preserving receptor-dependent cAMP responsiveness. CGS21680 had modest effects on early POS binding and uptake but enhanced post-ingestive POS clearance and rhodopsin degradation; these effects were attenuated by ZM241385 and ADORA2A knockdown. CGS21680 increased LC3B and Rubicon association with POS-containing structures, improved lysosomal acidification, and restored DQ Green BSA processing, Cathepsin D activity, and V-ATPase-associated assembly. Bafilomycin A1 increased LC3-II and p62 accumulation and impaired CGS21680-associated POS clearance, supporting lysosome-dependent turnover. Rubicon depletion attenuated CGS21680-associated improvements in POS clearance and lysosomal acidification. CGS21680 also reduced chronic stress-associated autofluorescence, oxidative stress, apoptosis, and junctional disruption. These protective effects were reproduced in primary RPE cells. In sodium iodate-injured mice, CGS21680 preserved outer retinal structure, reduced FITC-BSA leakage, improved electroretinographic responses, and was accompanied by increased CREB phosphorylation and recovery of Cathepsin D proteolytic competence. Conclusion ADORA2A activation promotes Rubicon-associated, lysosome-dependent LC3 processing of internalized POS and restores lysosomal degradative competence, supporting ADORA2A as a potential therapeutic target for early AMD.

Activation of ER stress mediated by HspB4/Cryaa reduction contributes to photoreceptor apoptosis in rd9 mouse model of X-linked Retinitis Pigmentosa

Results identify Cryaa as a critical regulator of ER stress and demonstrate that its reduction promotes UPR activation and subsequent photoreceptor apoptosis in the rd9 model, revealing a key role for Cryaa in XLRP pathology and may provide a novel therapeutic perspective for this disease.

Age- and sex-dependent retinal degeneration in peripherin-2 mutant mice, a model of central areolar choroidal dystrophy

Central areolar choroidal dystrophy (CACD) is a progressive macular dystrophy without treatment. Although PRPH2 mutations are the most common cause of disease, the mechanisms driving their phenotypic variability remain poorly understood. Here, a comprehensive characterization of the pathophysiological consequences of p.Arg195Leu mutation in PRPH2 was carried out in a mouse model (Prph2 KI/WT ). For evaluating the retinal degeneration, this study combines the analysis of electroretinographic responses together with a complete study of bulk RNA-seq transcriptomics and the key cellular and molecular pathways using confocal imaging, flow-cytometry and western blotting. Results from this work demonstrate that ageing and sex influence retinal degeneration. In young mutant mice, retinal functional impairment and structural disorganization of photoreceptor outer segments were accompanied by a reduced expression of Prph2 and Rom1 genes, together with the activation of the immune system and complement pathways. Middle-age stages represent a critical transition point where the increase of cell death and epithelial barrier dysfunction mark the beginning of retinal degeneration. These pathological events become evident at 9 months of age, where visual pathways-related genes were deregulated. Functional, cellular and molecular alterations observed in the mutant mice do not affect males and females equally. From the earliest stages of the disease, females exhibited greater and sustained inflammatory activation, mainly promoted by complement system upregulation and increased CD11b immunoreactivity, and potentially mediated by IL-6/STAT3/ERK signaling. Moreover, females had greater functional decline and photoreceptor loss. Together, these results could explain the high inter- and intrafamilial variability observed in CACD patients carrying the same mutation. Our findings identify inflammatory biomarkers accompanied by visual function loss, prior to evident retinal degeneration, and demonstrate that age and sex critically shape disease onset and severity. These insights underscore the necessity of incorporating sex-specific biology and early anti-inflammatory treatment into the development of targeted therapies for PRPH2-related dystrophies.