Please use this identifier to cite or link to this item: http://theses.ncl.ac.uk/jspui/handle/10443/5360
Title: Improving our understanding of autosomal dominant Retinitis Pigmentosa using PRPF31 patient-specific induced pluripotent stem cells (iPSCs)
Authors: Goes Barbosa Buskin, Adriana
Issue Date: 2019
Publisher: Newcastle University
Abstract: Retinitis pigmentosa (RP) is a genetic condition in which degeneration of photoreceptors, especially rods, gradually leads to visual loss. Patients with RP present symptoms such as dark adaptation or "night blindness," followed by “tunnel vision” and loss of central vision later on in the disease. PRPF31, a widely expressed splicing factor gene causative of RP, encodes a component of the U4/U6.U5 small trinuclear ribonucleic protein (tri-snRNP) complex, which is a constituent of the premRNA processing spliceosome. Mechanisms correlating mutations in splicing factors and retina-specific cell death are still poorly understood. To address this, induced pluripotent stem cells (iPSCs), which can differentiate into any cell types of the three germ layers and are capable of self-renewal, were used to generate patient-specific retinal pigment epithelium (RPE) models to investigate the pathogenesis of the PRPF31 form of RP. PRPF31-RPE patient-derived cells presented defects that impaired normal RPE structure and function, including disrupted apical-basal polarity, reduced transepithelial resistance and phagocytic capacity. Transcriptome profiles from PRPF31- RPE and other cell types revealed that disrupted alternative splicing was more pronounced in the RPE splicing programme. Mis-splicing of genes encoding pre-mRNA splicing proteins was limited to patient-specific RPE and retinal cells. Mis-splicing of genes implicated in ciliogenesis and cellular adhesion was associated with decreased cilia length and incidence, which in turn may have contributed with the severe RPE defects described above. In situ gene correction of a PRPF31 mutation rescued protein expression and key cellular phenotypes in RPE, providing proof-of-concept for future therapeutic strategies. In summary, the results generated by this study highlighted the applicability and importance of patient-specific iPSCs in disease modeling, unraveling more of the underlying molecular and cellular mechanisms responsible for causing RP.
Description: Ph. D. Thesis.
URI: http://hdl.handle.net/10443/5360
Appears in Collections:Institute of Genetic Medicine

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