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The Journal of Membrane Biology

, Volume 189, Issue 1, pp 35–43 | Cite as

Evidence for Segregation of Heterologous GPI-anchored Proteins into Separate Lipid Rafts within the Plasma Membrane

  • J. Wang
  • W. Gunning
  • K.M.M. Kelley
  • M. Ratnam

Cholesterol and glycosphingolipid-rich membrane rafts, which are rich in GPI-anchored proteins and are distinct from caveolae, are believed to serve as platforms for signal transduction events and protein recycling. GPI-anchored proteins with diverse functions as well as caveolin may be recovered in a membrane fraction insoluble in cold non-ionic detergent. This study tests for possible heterogeneity in the protein composition of the lipid rafts and detergent-insoluble membrane complexes by examining the two GPI-anchored homologous human folate receptors (FR)-a and -b, the GPI-anchored human placental alkaline phosphatase (PLAP), and caveolin (control) in transfected CHO cells. Both FR and PLAP showed the equal distribution of cell-surface vs. sequestered (recycling) protein typical of GPI-proteins. Quantitative affinity purification of detergent-insoluble complexes using biotinylated folate or specific antibodies demonstrated a strong association of the homologous FR-a and FR-b in the same detergent-insoluble complex and separate complexes containing either PLAP or caveolin. Immunogold localization experiments using antibody crosslinking to produce larger aggregates of GPI-anchored proteins for visualization by electron microscopy also showed a clear separation between FR- and PLAP-rich membrane microdomains. Thus, even though functionally diverse and heterologous GPI-anchored proteins are known to share endocytic and recycling vesicles, they may be segregated in distinct lipid rafts on the basis of their ecto(protein) domains facilitating clustering, compartmentalization and homotypic protein interactions.

Keywords

Lipid Raft Folate Receptor Membrane Microdomains Membrane Raft Signal Transduction Event 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

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Copyright information

© Springer-Verlag New York Inc. 2002

Authors and Affiliations

  • J. Wang
    • 1
  • W. Gunning
    • 2
  • K.M.M. Kelley
    • 1
  • M. Ratnam
    • 1
  1. 1.Department of of Biochemistry and Molecular Biology, Medical college of Ohio, 3035 Arlington Avenue, Toledo, OH 43614-5804, USAUS
  2. 2.Department of Pathology, Medical College of Ohio, 3035 Arlington Avenue, Toledo, OH 43614-5804, USAUS

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