Small Molecule Proteostasis Regulator Corrects Cone Photoreceptor Pathology in Retinal Organoids

Abstract

Purpose : The Unfolded Protein Response (UPR) is a conserved intracellular signal transduction mechanism that maintains organismal protein homeostasis and when disrupted, contributes to the pathology of many human diseases. Genetic mutations in the UPR regulator, ATF6, lead to heritable vision loss diseases such as achromatopsia (ACHM), where cone photoreceptors are dysfunctional. The mechanism by which ATF6 mutations cause cone photoreceptor dysfunction is unknown. Furthermore, there are no treatments for people carrying these mutations. Here, we created retinal organoids from stem cells of patients with ATF6 mutations or with CRISPR-edited ATF6 knock-out alleles to investigate pathomechanism and to test ways to prevent disease phenotypes.

Methods : Retinal organoids from ATF6 mutant patient derived iPSCs and ATF6 knock-out human ESCs were generated to investigate the function of ATF6 in developing photoreceptors. Surface-scanning live imaging and confocal immunofluorescent microscopy of sectioned retinal organoids were performed to examine morphology of WT and ATF6 mutant rods and cones. Adaptive optics laser-scanning confocal ophthalmoscopy (AOSLO) was performed to visualize photoreceptor morphology from ATF6 mutant patient stem cell donors. RNAseq analysis was performed to investigate rod and cone gene expression in WT and ATF6 mutant retinal organoids that were also treated with a small molecule ATF6 agonist and characterized by microscopy and gene expression profiling.

Results : Retinal organoids lacking functional ATF6 failed to form cone photoreceptors and instead, developed rod-dominant retinas, as determined by microscopy and gene expression analyses. AOSLO imaging of the fovea of stem cell donor patients carrying ATF6 mutations revealed significant loss of cones with contiguous rod structures that mirrored the surface-scanning phenotypes seen in our organoids. A small molecule proteostasis regulator that enhances ATF6 ER-to-Golgi trafficking potently restored transcriptional function of ATF6 Class 1 mutations and significantly enhanced cone photoreceptor differentiation in retinal organoids derived from these patients.

Conclusions : Our findings reveal that ATF6-associated ACHM arises from disruption of cone photoreceptor differentiation. Our findings identify a novel small molecule strategy to prevent cone photoreceptor disease based on reprogramming of the proteostasis network in the retina.

This is a 2020 ARVO Annual Meeting abstract.