June 2022
Volume 63, Issue 7
Open Access
ARVO Annual Meeting Abstract  |   June 2022
Dual mutational mechanism of the KIZ c.226C>T mutation as the cause of autosomal recessive retinitis pigmentosa
Author Affiliations & Notes
  • Yogapriya Sundaresan
    Department of Ophthalmology, Hadassah Medical Center, Jerusalem, Jerusalem, Israel
  • Maya Ross
    Koret School of Veterinary Medicine, The Hebrew University, Rehovot, Israel
  • Ron Ofri
    Koret School of Veterinary Medicine, The Hebrew University, Rehovot, Israel
  • Eyal Banin
    Department of Ophthalmology, Hadassah Medical Center, Jerusalem, Jerusalem, Israel
  • Dror Sharon
    Department of Ophthalmology, Hadassah Medical Center, Jerusalem, Jerusalem, Israel
  • Footnotes
    Commercial Relationships   Yogapriya Sundaresan None; Maya Ross None; Ron Ofri None; Eyal Banin None; Dror Sharon None
  • Footnotes
    Support  This study was supported by the Foundation Fighting Blindness USA (grant BR- GE-0214-0734 to DS and EB) and the Israel Science Foundation (grant 1778/20 to DS and EB) within the Israel Precision Medicine Partnership program
Investigative Ophthalmology & Visual Science June 2022, Vol.63, 1599 – A0388. doi:
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      Yogapriya Sundaresan, Maya Ross, Ron Ofri, Eyal Banin, Dror Sharon; Dual mutational mechanism of the KIZ c.226C>T mutation as the cause of autosomal recessive retinitis pigmentosa. Invest. Ophthalmol. Vis. Sci. 2022;63(7):1599 – A0388.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose : Mutations in the Kizuna (KIZ) gene, that encodes the centrosomal protein Kizuna, have been reported to cause autosomal recessive retinitis pigmentosa (ARRP), including a nonsense mutation (c.226C>T, p.R76*), located in exon 3. There is currently no approved treatment for KIZ-associated disease. Our aim is to characterize the expression pattern of KIZ in patient-derived skin fibroblasts as well as in normal mouse and sheep retinas by a novel method of alternative splicing analysis of RNA using next generation sequencing (NGS).

Methods : Skin biopsies were procured from four controls and three RP patients homozygous for KIZ-c.226C>T. In addition, retinas dissected from normal mice (n=6) and sheep (n=1) were included in the analysis. Primers for RT-PCR and NGS amplifying exons 2 to 5 were designed using Primer3. Following NGS, the aligned BAM files were analyzed using Integrative Genomics Viewer (IGV). In addition, potential exonic splicing enhancer (ESEs) sites were determined in WT and mutant human sequence using ESEFinder 3.0.

Results : RT-PCR and NGS-based analyses of RNA isolated from primary fibroblasts of patients and controls revealed the presence of four different transcripts: (i) the full-length transcript, (ii) skipping of exon 3, (iii) skipping of exon 3 and inclusion of an alternative exon, and (iv) skipping of both exons 3 and 4. Subsequent analysis of the expression pattern in the mouse and sheep retinas identified only two transcripts: (i) the full-length transcript and (ii) skipping of exon 4. The analyses revealed a lower expression level of the full-length transcript in patients compared to control (22% versus 58%). ESE analysis identified a 7 bp long sequence that includes the c.226 position with a higher score for the human WT sequence (4.411) compared to the mutant sequence (2.694).

Conclusions : KIZ- c.226C>T is a relatively frequent cause of ARRP in the Jewish population. Our data indicate that this variant might affect a putative ESE, that results in pronounced skipping of exon 3. Therefore, mutation-specific therapies, such as readthrough therapy and RNA editing, might show lower than expected efficacy since many transcripts do not contain this mutation. The KIZ nonsense variant is one of the few IRD mutations reported to affect an ESE, but we predict that ESE-affecting mutations are more common and need RNA-based analyses to be identified.

This abstract was presented at the 2022 ARVO Annual Meeting, held in Denver, CO, May 1-4, 2022, and virtually.

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