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Global analysis of dynamic changes in lipid raft proteins during T‐cell activation
Author(s) -
Kobayashi Michimoto,
Katagiri Takuya,
Kosako Hidetaka,
Iida Naoyuki,
Hattori Seisuke
Publication year - 2007
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.200600675
Subject(s) - lipid raft , microbiology and biotechnology , pleckstrin homology domain , biology , t cell receptor , immunological synapse , endocytosis , lipid microdomain , signal transduction , t cell , biochemistry , cell , membrane , immune system , immunology
Abstract Lipid rafts are considered as specialized microdomains within the plasma membrane with unique lipid compositions different from surrounding membranes. Following T‐cell receptor (TCR) stimulation, lipid rafts assemble in T‐cell/antigen‐presenting cell (APC) contact site known as the immunological synapse, inner leaflets of which serve as activation or docking sites for downstream signaling components. To understand the signaling events occurring in lipid rafts, we globally analyzed dynamic changes in lipid raft proteins during TCR/CD28 costimulation using 2‐D fluorescence difference gel electrophoresis. We detected multiple spots whose intensities were enhanced after costimulation, and identified proteins in these spots by PMF. Identified proteins include Src family tyrosine kinases, tyrosine phosphatase, phosphatidylinositol 3‐kinase (PI3‐kinase), actin‐binding proteins, and regulators for small GTPases. Of particular interest, a number of pleckstrin homology (PH) domain‐containing proteins were identified. Biochemical and histochemical analyses confirmed the translocation of these proteins from cytosol to lipid rafts. We also demonstrated that these proteins assembled at the T‐cell/APC interface. These results indicate the efficacy of our system to systematically analyze dynamics of lipid raft proteins during extracellular stimulation.