Mapping Keratoconus Molecular Substrates by Multiplexed High-Resolution Proteomics of Unpooled Corneas

Vishal Shinde, Nan Hu, Santosh Renuse, Alka Mahale, Akhilesh Pandey, Charles Eberhart, Donald Stone, Samar A. Al-Swailem, Azza Maktabi, Shukti Chakravarti

Research output: Contribution to journalArticle

Abstract

Keratoconus (KCN) is a leading cause for cornea grafting worldwide. Keratoconus is a multifactorial disease that causes progressive thinning of the cornea and whose etiology is poorly understood. Several studies have used proteomics on patient tear fluids to identify potential biomarkers. However, proteome of the cornea itself has not been investigated fully. We report here new findings from a case-control study using multiplexed mass spectrometry (MS) on individual (unpooled) corneas to gain deeper insights into proteins and biomarkers relevant to keratoconus. We employed a high-pressure approach to extract total protein from individual corneas from five cases and five controls, followed by trypsin digestion and tandem mass tag (TMT) labeling. The MS-derived data were searched using the Human NCBI RefSeq protein database v92, with peptides and proteins filtered at 1% false discovery rate. A total of 3132 proteins were detected, of which 627 were altered significantly (p ≤ 0.05) in keratoconus corneas. The increases were overwhelmingly in the mTOR/PI3/AKT signal-mediated regulations of cell survival and proliferation, nonsense-mediated decay of transcripts, and proteasomal pathways. The decreases were in several extracellular matrix proteins and in many members of the complement system. Importantly, this multiplexed proteomic study of keratoconus corneas identified, to our knowledge, the largest number of corneal proteins. The novel findings include changes in pathways that regulate transcript stability, proteasomal degradation, and the complement system in corneas with keratoconus. These observations offer new prospects toward future discovery of novel molecular targets for diagnostic and therapeutic innovations for patients with keratoconus.

Original languageEnglish (US)
Pages (from-to)583-597
Number of pages15
JournalOMICS A Journal of Integrative Biology
Volume23
Issue number11
DOIs
StatePublished - Nov 2019

Keywords

  • ER stress
  • complement
  • cornea
  • keratoconus
  • proteasomal degradation
  • proteomics

ASJC Scopus subject areas

  • Biotechnology
  • Biochemistry
  • Molecular Medicine
  • Molecular Biology
  • Genetics

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  • Cite this

    Shinde, V., Hu, N., Renuse, S., Mahale, A., Pandey, A., Eberhart, C., Stone, D., Al-Swailem, S. A., Maktabi, A., & Chakravarti, S. (2019). Mapping Keratoconus Molecular Substrates by Multiplexed High-Resolution Proteomics of Unpooled Corneas. OMICS A Journal of Integrative Biology, 23(11), 583-597. https://doi.org/10.1089/omi.2019.0143