Original Article

Aggregation of Lipid Rafts Accompanies Signaling Via the T Cell Antigen Receptor

Correspondence to: Anthony I. Magee, National Institute for Medical Research, The Ridgeway, Mill Hill, London NW7 1AA, UK. Tel:44-181-959-3666 Fax:44-181-906-4477 E-mail:t-magee{at}nimr.mrc.ac.uk or s-ley@nimr.mrc.ac.uk.

The role of lipid rafts in T cell antigen receptor (TCR) signaling was investigated using fluorescence microscopy. Lipid rafts labeled with cholera toxin B subunit (CT-B) and cross-linked into patches displayed characteristics of rafts isolated biochemically, including detergent resistance and colocalization with raft-associated proteins. LCK, LAT, and the TCR all colocalized with lipid patches, although TCR association was sensitive to nonionic detergent. Aggregation of the TCR by anti-CD3 mAb cross-linking also caused coaggregation of raft-associated proteins. However, the protein tyrosine phosphatase CD45 did not colocalize to either CT-B or CD3 patches. Cross-linking of either CD3 or CT-B strongly induced tyrosine phosphorylation and recruitment of a ZAP-70(SH2)₂–green fluorescent protein (GFP) fusion protein to the lipid patches. Also, CT-B patching induced signaling events analagous to TCR stimulation, with the same dependence on expression of key TCR signaling molecules. Targeting of LCK to rafts was necessary for these events, as a nonraft- associated transmembrane LCK chimera, which did not colocalize with TCR patches, could not reconstitute CT-B–induced signaling. Thus, our results indicate a mechanism whereby TCR engagement promotes aggregation of lipid rafts, which facilitates colocalization of LCK, LAT, and the TCR whilst excluding CD45, thereby triggering protein tyrosine phosphorylation.

Key Words: lipid rafts, T cell antigen receptor, LCK, cholera toxin-B, signal transduction

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• Tai, Y.-T., Podar, K., Catley, L., Tseng, Y.-H., Akiyama, M., Shringarpure, R., Burger, R., Hideshima, T., Chauhan, D., Mitsiades, N., Richardson, P., Munshi, N. C., Kahn, C. R., Mitsiades, C., Anderson, K. C. (2003). Insulin-like Growth Factor-1 Induces Adhesion and Migration in Human Multiple Myeloma Cells via Activation of {beta}1-Integrin and Phosphatidylinositol 3'-Kinase/AKT Signaling. Cancer Res. 63: 5850-5858 [Abstract] [Full Text]  
• Hare, K. J., Pongracz, J., Jenkinson, E. J., Anderson, G. (2003). Modeling TCR Signaling Complex Formation in Positive Selection. J. Immunol. 171: 2825-2831 [Abstract] [Full Text]  
• Park, J., Cho, N.-H., Choi, J.-K., Feng, P., Choe, J., Jung, J. U. (2003). Distinct Roles of Cellular Lck and p80 Proteins in Herpesvirus Saimiri Tip Function on Lipid Rafts. J. Virol. 77: 9041-9051 [Abstract] [Full Text]  

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