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Activation of autophagy reverses progressive and deleterious protein aggregation in PRPF31 patient iPSC-derived retinal pigment epithelium cells

Lookup NU author(s): Dr Maria Georgiou, Dr Chunbo Yang, Rob Atkinson, Dr Adriana BuskinORCiD, Dr Joseph Collin, Tracey DaveyORCiD, Professor Lyle Armstrong, Dr Viktor Korolchuk, Professor Majlinda LakoORCiD



This work is licensed under a Creative Commons Attribution 4.0 International License (CC BY 4.0).


Mutations in pre-mRNA processing factor 31 (PRPF31), a core protein of the spliceosomal tri-snRNP complex, cause autosomal-dominant retinitis pigmentosa (adRP). It has remained an enigma why mutations in ubiquitously expressed tri-snRNP proteins result in retina-specific disorders, and so far, the underlying mechanism of splicing factors-related RP is poorly understood. Here, we used iPSC technology to generate retinal organoids and RPE models from four patients with severe and very severe PRPF31-adRP, unaffected individuals and a CRISPR/Cas9 isogenic control. To fully assess the impacts of PRPF31 mutations, quantitative proteomics analyses of retinal organoids and RPE cells was carried out showing RNA splicing, autophagy and lysosome, unfolded protein response (UPR) and visual cycle-related pathways to be significantly affected. Strikingly, the patient-derived RPE and retinal cells were characterised by the presence of large amounts of cytoplasmic aggregates containing the mutant PRPF31 and misfolded, ubiquitin-conjugated proteins including key visual cycle and other RP-linked tri-snRNP proteins, which accumulated progressively with time. The mutant PRPF31 variant was not incorporated into splicing complexes, but reduction of PRPF31 wildtype levels led to tri-snRNP assembly defects in Cajal bodies of PRPF31 patient retinal cells, altered morphology of nuclear speckles and reduced formation of active spliceosomes giving rise to global splicing dysregulation. Moreover, the impaired waste disposal mechanisms further exacerbated aggregate formation, and targeting these by activating the autophagy pathway using Rapamycin reduced cytoplasmic aggregates, leading to improved cell survival. Our data demonstrate that it is the progressive aggregate accumulation that overburdens the waste disposal machinery rather than direct PRPF31-initiated mis-splicing, and thus relieving the RPE cells from insoluble cytoplasmic aggregates presents a novel therapeutic strategy that can be combined with gene therapy studies to fully restore RPE and retinal cell function in PRPF31-adRP patients.

Publication metadata

Author(s): Georgiou M, YAng C, Atkinson R, Pan K, Buskin A, Molina M, Collin C, Alaama J, Goertler F, Ludwig S, Davey T, Luhrmann R, Grellscheid S, Johnson C, Ali R, Armstring L, Korolchuk V, Urlaub H, Mozzafari J, LAko M

Publication type: Article

Publication status: Published

Journal: Clinical and Translational Medicine

Year: 2022

Volume: 12

Issue: 3

Online publication date: 16/03/2022

Acceptance date: 23/02/2022

Date deposited: 20/05/2022

ISSN (electronic): 2001-1326

Publisher: Wiley


DOI: 10.1002/ctm2.759


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Funder referenceFunder name
MR/T017503/1Medical Research Council (MRC)