Differential insertion of GPI-anchored GFPs into lipid rafts of live cells
| Autor: | Claude Bron, Marie-Agnès Doucey, Pascal Schneider, Florent C. Bender, Laurence Chapatte, Daniel F. Legler |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2005 |
| Předmět: |
Glycosylphosphatidylinositols
Green Fluorescent Proteins GPI-Linked Proteins Cell Cycle Proteins Biology Kidney Biochemistry Green fluorescent protein Membrane Microdomains Genetics Humans Molecular Biology Lipid raft Membrane Glycoproteins Cell Membrane Peripheral membrane protein fungi Membrane Proteins Cell Cycle Proteins/chemistry Cell Cycle Proteins/metabolism Cell Membrane/chemistry Glycosylphosphatidylinositols/chemistry Green Fluorescent Proteins/chemistry Green Fluorescent Proteins/metabolism Indicators and Reagents/chemistry Indicators and Reagents/metabolism Kidney/chemistry Kidney/cytology Membrane Glycoproteins/chemistry Membrane Glycoproteins/metabolism Membrane Microdomains/chemistry Membrane Proteins/chemistry Membrane Proteins/metabolism Neoplasm Proteins/chemistry Neoplasm Proteins/metabolism Fluorescence Gpi anchored protein Neoplasm Proteins Cell biology carbohydrates (lipids) Indicators and Reagents lipids (amino acids peptides and proteins) Biotechnology Glycosylphosphatidyl inositol |
| Zdroj: | FASEB Journal, vol. 19, no. 1, pp. 73-75 |
| Popis: | Partitioning of proteins in cholesterol and sphingolipid enriched plasma membrane microdomains, called lipid rafts, is critical for many signal transduction and protein sorting events. Although raft partitioning of many signaling molecules remains to be determined, glycosylphosphatidyl-inositol (GPI)-anchored proteins possess high affinity for lipid rafts and are currently exploited as markers to investigate fundamental mechanisms in protein sorting and signal transduction events. In this study, we demonstrate that two recombinant GPI-anchored green fluorescent proteins (GFP-GPIs) that differ in their GPI signal sequence confer distinct localization in plasma membrane microdomains. GFP fused to the GPI signal of the decay accelerating factor GFP-GPI(DAF) partitioned exclusively in lipid rafts, whereas GFP fused to the GPI signal of TRAIL-R3, GFP-GPI(TRAIL-R3), associated only minimally with microdomains. In addition, we investigated the unique ability of purified GFP-GPIs to insert into membrane microdomains of primary lymphocytes. This cell surface painting allows rapid, stable, and functional association of the GPI-anchored proteins with the target cell plasma membrane. The distinct membrane localization of the two GFP-GPIs was observed irrespective of whether the GPI-anchored molecules were painted or transfected. Furthermore, we show that painted GFP-GPI(DAF) was totally dependent on the GPI anchor and that the membrane insertion was increased by the addition of raft-associated lipids such as cholesterol, sphingomyelin, and dipalmitoyl-phosphatidylethanolamine. Thus, this study provides evidence that different GPI signal sequences lead to distinct membrane microdomain localization and that painted GFP-GPI(DAF) serves as an excellent fluorescent marker for lipid rafts in live cells. |
| Databáze: | OpenAIRE |
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