International Immunology Advance Access originally published online on September 26, 2008
International Immunology 2008 20(11):1427-1437; doi:10.1093/intimm/dxn100
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Impaired TCR signaling through dysfunction of lipid rafts in sphingomyelin synthase 1 (SMS1)-knockdown T cells
1 Department of Hematology and Immunology, Kanazawa Medical University, 1-1 Daigaku, Uchinada, Ishikawa 920-0293, Japan
2 Department of Hematology and Immunology, Tongji Hospital, Huazhong University of Science and Technology, Wuhan, Hubei 430030, China
3 Department of Biochemistry, Osaka Dental University, Kuzuha, Osaka, Japan
4 Department of Rheumatology and Clinical Immunology, Kyoto University Graduate School of Medicine, Sakyo-ku, Kyoto 606-8507, Japan
5 Department of Biochemistry and Molecular Biology, Osaka University Graduate School of Medicine, Osaka 565-0871, Japan
6 Division of Cellular and Gene Therapies (HFM-725), Center for Biologics Evaluation and Research, Food and Drug Administration, Bethesda, MD 20892, USA
7 Department of Clinical Laboratory, Medicine/Hematology, Faculty of Medicine, Tottori University, Yonago, Tottori 683-8504, Japan
Correspondence to: H. Umehara; E-mail: umehara{at}kanazawa-med.ac.jp
During T cell activation, TCRs cluster at the center of the T cell–antigen-presenting cell interface forming the central supramolecular activation cluster. Although it has been suggested that sphingolipid- and cholesterol-rich microdomains, termed lipid rafts, form platforms for the regulation and transduction of TCR signals, an actual role for membrane sphingomyelin (SM), a key component of lipid rafts, has not been reported. After cloning a gene responsible for SM synthesis, sphingomyelin synthase (SMS) 1, we established a SM-knockdown cell line (Jurkat-SMS1/kd) by transfection of SMS1-short-interfering RNA into Jurkat T cells, which is deficient in membrane expression of SM. Upon CD3 stimulation, expression of CD69 (the earliest leukocyte activation antigen), activation-induced cell adhesion and proliferation as well as TCR clustering was severely impaired in Jurkat-SMS1/kd cells. CD3-induced tyrosine phosphorylation and association of linker for activation of T cell with ZAP-70 and Grb2 and phosphorylation of protein kinase C (PKC) 
were also severely impaired in Jurkat-SMS1/kd cells. Finally, translocation of TCR, ZAP-70 and PKC into lipid rafts was markedly decreased in Jurkat-SMS1/kd cells. These findings indicate that membrane SM is crucial for TCR signal transduction, leading to full T cell activation through lipid raft function.
Keywords: LAT, lipid rafts, microdomain, sphingomyelin, TCR
Transmitting editor: K. Okumura
Received 28 June 2008, accepted 11 August 2008.