Rapid Variant Pathogenicity Analysis by CRISPR Activation of CRB1 Gene Expression in Patient-Derived Fibroblasts

Sang Yoon Moon, Dan Zhang, Shang Chih Chen, Tina M. Lamey, Jennifer A. Thompson, Terri L. McLaren, Fred K. Chen, Samuel McLenachan

Research output: Contribution to journalArticlepeer-review

Abstract

Inherited retinal diseases (IRDs) are a heterogeneous group of blinding genetic disorders caused by pathogenic variants in genes expressed in the retina. In this study, we sought to develop a method for rapid evaluation of IRD gene variant pathogenicity by inducing expression of retinal genes in patient-derived fibroblasts using CRISPR-activation (CRISPRa). We demonstrate CRISPRa of CRB1 expression in fibroblasts derived from patients with retinitis pigmentosa, enabling investigation of pathogenic mechanisms associated with specific variants. We show the CRB1 c.4005 + 1G>A variant caused exon 11 skipping in CRISPR-activated fibroblasts and retinal organoids (ROs) derived from the same RP12 patient. The c.652 + 5G>C variant was shown to enhance exon 2 skipping in CRISPR-activated fibroblasts and differentially affected CRB1 isoform expression in fibroblasts and ROs. Our study demonstrates an accessible platform for transcript screening of IRD gene variants in patient-derived fibroblasts, which can potentially be applied for rapid pathogenicity assessments of any gene variant.

Original languageEnglish
Pages (from-to)100-110
Number of pages11
JournalCRISPR Journal
Volume7
Issue number2
DOIs
Publication statusPublished - Apr 2024

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