July 2019
Volume 60, Issue 9
Open Access
ARVO Annual Meeting Abstract  |   July 2019
Mapping interleukin-33 to metabolic function in the retinal pigment epithelium
Author Affiliations & Notes
  • Louis Scott
    Ophthalmology, University of Bristol, United Kingdom
  • Emma E Vincent
    Cellular and Molecular Medicine, University of Bristol, Bristol, United Kingdom
  • Natalie Hudson
    Genetics, Trinity College Dublin, Ireland
  • David Copland
    Ophthalmology, University of Bristol, United Kingdom
    UCL Institute of Ophthalmology, United Kingdom
  • Kate Heesom
    Proteomics Facility, University of Bristol, United Kingdom
  • Matthew Campbell
    Genetics, Trinity College Dublin, Ireland
  • Andrew Halestrap
    Biochemistry, University of Bristol, United Kingdom
  • Andrew D Dick
    Ophthalmology, University of Bristol, United Kingdom
    UCL Institute of Ophthalmology, United Kingdom
  • Sofia Theodoropoulou
    Ophthalmology, University of Bristol, United Kingdom
  • Footnotes
    Commercial Relationships   Louis Scott, None; Emma Vincent, None; Natalie Hudson, None; David Copland, None; Kate Heesom, None; Matthew Campbell, None; Andrew Halestrap, None; Andrew Dick, None; Sofia Theodoropoulou, None
  • Footnotes
    Support  Macular Society (L.M.SCOTT and A.D. DICK)
Investigative Ophthalmology & Visual Science July 2019, Vol.60, 1685. doi:
  • Views
  • Share
  • Tools
    • Alerts
      ×
      This feature is available to authenticated users only.
      Sign In or Create an Account ×
    • Get Citation

      Louis Scott, Emma E Vincent, Natalie Hudson, David Copland, Kate Heesom, Matthew Campbell, Andrew Halestrap, Andrew D Dick, Sofia Theodoropoulou; Mapping interleukin-33 to metabolic function in the retinal pigment epithelium. Invest. Ophthalmol. Vis. Sci. 2019;60(9):1685.

      Download citation file:


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

      ×
  • Supplements
Abstract

Purpose : Interleukin-33 (IL-33), is an IL-1 family cytokine that is constitutively expressed in the inner retina, RPE and choroid. IL-33 functions as an “alarmin” molecule released from barrier cells following cellular stress. We have previously demonstrated a role for IL-33 in the homeostatic regulation and maintenance of mitochondrial metabolism. The aim of this study was to further determine the role of IL-33 in the regulation of RPE metabolism, utilizing a proteomics approach to map the molecular interactions (interactome) of IL-33.

Methods : Using retinal pigment epithelium (RPE) whole cell lysates derived from both human (ARPE-19) and mouse (primary isolated RPE), immunoprecipitation of IL-33 was performed. Proteomic analysis of the IL-33 “pull-down” was assessed using tandem mass-tagging (mass spectrometry-based quantification). Interactions were mapped to human or murine UniProt databases, and ranked analysis performed using STRING (https://string-db.org/).The role of cellular IL-33 was further assessed in vitro using both knock-down (siRNA) and over-expression (CRISPRcas9) approaches. Cellular metabolism was assessed using seahorse extracellular flux, C13 glucose pathway tracing and immunoblotting/RT-PCR quantification of metabolic pathway intermediates.

Results : Pathway analysis of the IL-33 interactome demonstrated an enrichment of targets involved with protein translation, RNA splicing, metabolism and innate immune regulation. Importantly for cellular metabolism and regulation of RPE glycolytic potential, IL-33 interacts with the splicing factors heterogeneous nuclear ribonucleoproteins A1 + A2 (hnRNPA1/2) and serine/arginine-rich splicing factor 3 (SRSF3) which regulate alternative splicing of the pyruvate kinase gene (Pkm).In IL-33 deficient RPE, altered ratios of Pkm splice variants are observed, alongside with reduced mitochondrial function and structural and morphological differences. Overexpression of IL-33 lead to increased glycolytic flux and mitochondrial activity, dependent on pyruvate import into the mitochondria.

Conclusions : These findings highlight a role for intracellular IL-33 in the regulation of metabolic enzyme expression, serving as a molecular check-point to license mitochondrial respiration. This has implications for a degenerative retina, where the strain on mitochondrial function with age may subvert the ability of RPE to respond appropriately to the metabolic demands of inflammation.

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

×
×

This PDF is available to Subscribers Only

Sign in or purchase a subscription to access this content. ×

You must be signed into an individual account to use this feature.

×