Delivery of raft-associated, GPI-anchored proteins to the apical surface of polarized MDCK cells by a transcytotic pathway

Nat Cell Biol. 2004 Apr;6(4):297-307. doi: 10.1038/ncb1109. Epub 2004 Mar 28.

Abstract

Epithelial cell polarity depends on mechanisms for targeting proteins to different plasma membrane domains. Here, we dissect the pathway for apical delivery of several raft-associated, glycosyl phosphatidylinositol (GPI)-anchored proteins in polarized MDCK cells using live-cell imaging and selective inhibition of apical or basolateral exocytosis. Rather than trafficking directly from the trans-Golgi network (TGN) to the apical plasma membrane as previously thought, the GPI-anchored proteins followed an indirect, transcytotic route. They first exited the TGN in membrane-bound carriers that also contained basolateral cargo, although the two cargoes were laterally segregated. The carriers were then targeted to and fused with a zone of lateral plasma membrane adjacent to tight junctions that is known to contain the exocyst. Thereafter, the GPI-anchored proteins, but not basolateral cargo, were rapidly internalized, together with endocytic tracer, into clathrin-free transport intermediates that transcytosed to the apical plasma membrane. Thus, apical sorting of these GPI-anchored proteins occurs at the plasma membrane, rather than at the TGN.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Biomarkers
  • Cell Line
  • Cell Polarity / physiology*
  • Cricetinae
  • Dogs
  • Endocytosis / physiology
  • Epithelial Cells / metabolism*
  • Epithelial Cells / ultrastructure
  • Exocytosis / drug effects
  • Exocytosis / physiology
  • Glycosylphosphatidylinositols / metabolism*
  • Humans
  • Hydrolyzable Tannins / pharmacology
  • Kidney / metabolism
  • Kidney / ultrastructure
  • Membrane Fusion / physiology
  • Membrane Microdomains / metabolism*
  • Membrane Microdomains / ultrastructure
  • Microscopy, Electron
  • Protein Transport / physiology*
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism
  • Transport Vesicles / drug effects
  • Transport Vesicles / metabolism
  • Transport Vesicles / ultrastructure
  • trans-Golgi Network / drug effects
  • trans-Golgi Network / metabolism
  • trans-Golgi Network / ultrastructure

Substances

  • Biomarkers
  • Glycosylphosphatidylinositols
  • Hydrolyzable Tannins
  • Recombinant Fusion Proteins