EphA2 proteomics in human keratinocytes reveals a novel association with afadin and epidermal tight junctions

Bethany E. Perez White, Rosa Ventrella, Nihal Kaplan, Calvin J. Cable, Paul M. Thomas, Spiro Getsios*

*Corresponding author for this work

Research output: Contribution to journalArticle

12 Scopus citations

Abstract

EphA2 is a receptor tyrosine kinase that helps to maintain epidermal tissue homeostasis. A proximity-dependent biotin identification (BioID) approach was used to identify proteins in close proximity to EphA2 within primary human keratinocytes and three-dimensional (3D) reconstituted human epidermis (RHE) cultures to map a putative protein interaction network for this membrane receptor that exhibits a polarized distribution in stratified epithelia. Although a subset of known EphA2 interactors were identified in the BioID screen, > 97% were uniquely detected in keratinocytes with over 50% of these vicinal proteins only present in 3D human epidermal culture. Afadin (AFDN), a cytoskeletal and junction-associated protein, was present in 2D and 3D keratinocyte cultures, and validated as a so-far-unknown EphA2- interacting protein. Loss of EphA2 protein disrupted the subcellular distribution ofafadinandoccludin indifferentiated keratinocytes, leading to impairment of tight junctions. Collectively, these studies illustrate the use of the BioID approach in order to map receptor interaction networks in 3D human epithelial cultures, and reveal a positive regulatory role for EphA2 in the organization of afadin and epidermal tight junctions.

Original languageEnglish (US)
Pages (from-to)111-118
Number of pages8
JournalJournal of cell science
Volume130
Issue number1
DOIs
StatePublished - 2017

Keywords

  • 3D culture
  • Afadin
  • EphA2
  • Keratinocytes
  • Proteomics
  • Tight junction

ASJC Scopus subject areas

  • Cell Biology

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