July 2019
Volume 60, Issue 9
Open Access
ARVO Annual Meeting Abstract  |   July 2019
PPIP5K2 mutations & Keratoconus in Human Patients and Mouse Models
Author Affiliations & Notes
  • Yutao Liu
    Cellular Biology and Anatomy, Augusta University, Augusta, Georgia, United States
    James & Jean Culver Vision Discovery Institute, Augusta University, Augusta, GA, Georgia
  • Mariam Khaled
    Cellular Biology and Anatomy, Augusta University, Augusta, Georgia, United States
  • Yelena Bykhovskaya
    Surgery, Cedars-Sinai Medical Center, Los Angeles, California, United States
  • Khaled Abu-Amero
    Department of Ophthalmology, College of Medicine, King Saud University, Riyadh, Saudi Arabia
  • Sylvia B Smith
    Cellular Biology and Anatomy, Augusta University, Augusta, Georgia, United States
    James & Jean Culver Vision Discovery Institute, Augusta University, Augusta, GA, Georgia
  • Mitchell A Watsky
    Cellular Biology and Anatomy, Augusta University, Augusta, Georgia, United States
    James & Jean Culver Vision Discovery Institute, Augusta University, Augusta, GA, Georgia
  • Amy Estes
    Ophthalmology, Augusta University, Augusta, Georgia, United States
    James & Jean Culver Vision Discovery Institute, Augusta University, Augusta, GA, Georgia
  • Stephen B. Shears
    Signal Transduction Laboratory, National Institute of Environmental Health Sciences, Research Triangle Park, North Carolina, United States
  • Anthony N Kuo
    Ophthalmology & Biomedical Engineering, Duke University Medical Center, Durham, North Carolina, United States
  • Yaron S Rabinowitz
    Surgery, Cedars-Sinai Medical Center, Los Angeles, California, United States
  • Footnotes
    Commercial Relationships   Yutao Liu, None; Mariam Khaled, None; Yelena Bykhovskaya, None; Khaled Abu-Amero, None; Sylvia Smith, None; Mitchell Watsky, None; Amy Estes, None; Stephen Shears, None; Anthony Kuo, None; Yaron Rabinowitz, None
  • Footnotes
    Support  This work was supported in part by National Institutes of Health (NIH; Bethesda, MD, USA) grants R01EY023242 and R21028671 (YL), R01EY021747 (MAW), R01EY009052 (YSR), R01EY024312 (ANK), and R01EY028103 (SBS).
Investigative Ophthalmology & Visual Science July 2019, Vol.60, 4784. doi:
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    • Get Citation

      Yutao Liu, Mariam Khaled, Yelena Bykhovskaya, Khaled Abu-Amero, Sylvia B Smith, Mitchell A Watsky, Amy Estes, Stephen B. Shears, Anthony N Kuo, Yaron S Rabinowitz; PPIP5K2 mutations & Keratoconus in Human Patients and Mouse Models. Invest. Ophthalmol. Vis. Sci. 2019;60(9):4784.

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

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Abstract

Purpose : Genetic factors contribute to the pathogenesis of Keratoconus (KC). A linkage locus (Chr5q14.3-q21.1) was reported in a four-generation KC pedigree with an autosomal dominant mode; however, no mutations have been identified. We aimed to identify and characterize the underlying mutation and the related molecular mechanism.

Methods : We selected 10 subjects for whole exome sequencing (WES) and 5 subjects for whole genome sequencing. WES was also performed in other 20 multiplex families. We filtered and focused on exonic, non-synonymous variants with good quality and read depth, segregated with the disease phenotype, and with MAF≤1% in public databases. We validated the variants using Sanger sequencing. The biochemical impact of the identified PPIP5K2 mutations was evaluated using an established HPLC-based in vitro assay with purified overexpressed proteins carrying these mutations. We assessed the gene expression in human primary corneal epithelial and stromal cells using droplet digital PCR and the protein expression in mouse and human corneas using immunofluorescence. We examined a Ppip5k2 gene trap mouse model with no phosphatase activity, but elevated kinase activity (Ppip5k2K^) using slit lamp, OCT (Optical Coherence Tomography) scanning, and histology staining for KC-related corneal phenotypes.

Results : We identified a missense variant in PPIP5K2 (c.1255T>G, p.Ser419Ala) in the linkage family and another variant (c.2528A>G, p.Asn843Ser) in a second family. PPIP5K2 encodes a bifunctional kinase/phosphatase that controls metabolism and signaling of inositol phosphates. In vitro biochemical assays indicate reduced phosphatase activity of PPIP5K2 carrying these two mutations. Both human corneal epithelial and stromal cells showed higher expression of PPIP5K2 mRNA. Protein staining indicated significantly higher expression of PPIP5K2 than PPIP5K1 in human and mouse cornea. Mouse corneal OCT-derived tomography and histology demonstrated the presence of KC-related corneal surface irregularities in the Ppip5k2K^/Ppip5k2K^ mice compared to littermate controls at 3-4 months of age. Slit lamp data indicated varying degrees of corneal opacity and shallow anterior chamber compared to littermate controls at 3 months.

Conclusions : We identified two potential mutations in the phosphatase domain of PPIP5K2 gene in multiplex KC families. Our in vitro and in vivo data strongly suggest an important role of PPIP5K2 in KC pathogenesis.

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.

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