Dual role of SLP-76 in mediating T cell receptor-induced activation of phospholipase C-γ1
Dual role of SLP-76 in mediating T cell receptor-induced activation of phospholipase C-γ1
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DOI：
10.1074/jbc.m606697200
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发表时间：
2007-02-02
影响因子：
4.8
通讯作者：
Yablonski, Deborah
中科院分区：
文献类型：
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作者：
Beach, Dvora;Gonen, Ronnie;Yablonski, Deborah
磷脂酶C-γ 1（PLC-γ 1）的激活依赖于由LAT、Gads和SLP-76组成的接头蛋白的异源三聚体复合物。在T细胞受体刺激后，PLC-γ 1的一部分被募集到被称为鞘糖脂富集膜微结构域（GEM）或脂筏的抗洗涤剂膜部分，LAT组成性地定位于该膜部分。除了LAT之外，PLC-γ 1 GEM募集依赖于SLP-76，并且特别地，需要SLP-76的Gads结合结构域。SLP-76的N-末端酪氨酸磷酸化位点和P-I区对于PLC-γ 1 GEM募集不是必需的，但是对于PLC-γ 1在Tyr（783）处的磷酸化是必需的。因此，在不存在第二SLP-76介导的事件的情况下，GEM募集可能不足以完全激活PLC-γ 1。事实上，PLC-γ 1的GEM靶向衍生物依赖于SLP-76进行T细胞受体诱导的Tyr（783）磷酸化和随后的NFAT活化。在生物化学水平上，SLP-76可诱导地与Vav和催化活性ITK结合，其在体外有效地磷酸化Tyr（783）处的PLC-γ 1片段。这两个协会被破坏后，突变的N-末端酪氨酸磷酸化位点的SLP-76。P-I区缺失破坏了Vav结合并降低了SLP-76相关激酶活性。P-I区域内较小的缺失不会损害PLC-γ 1活化，不会损害与Vav的关联，但会降低SLP-76相关激酶活性。这些结果为SLP-76的多种作用及其与其他信号蛋白相互作用的功能重要性提供了新的见解。
Phospholipase C-gamma 1 (PLC-gamma 1) activation depends on a heterotrimeric complex of adaptor proteins composed of LAT, Gads, and SLP-76. Upon T cell receptor stimulation, a portion of PLC-gamma 1 is recruited to a detergent-resistant membrane fraction known as the glycosphingolipid-enriched membrane microdomains (GEMs), or lipid rafts, to which LAT is constitutively localized. In addition to LAT, PLC-gamma 1 GEM recruitment depended on SLP-76, and, in particular, required the Gads-binding domain of SLP-76. The N-terminal tyrosine phosphorylation sites and P-I region of SLP-76 were not required for PLC-gamma 1 GEM recruitment, but were required for PLC-gamma 1 phosphorylation at Tyr(783). Thus, GEM recruitment can be insufficient for full activation of PLC-gamma 1 in the absence of a second SLP-76-mediated event. Indeed, a GEM-targeted derivative of PLC-gamma 1 depended on SLP-76 for T cell receptor-induced phosphorylation at Tyr(783) and subsequent NFAT activation. On a biochemical level, SLP-76 inducibly associated with both Vav and catalytically active ITK, which efficiently phosphorylated a PLC-gamma 1 fragment at Tyr(783) in vitro. Both associations were disrupted upon mutation of the N-terminal tyrosine phosphorylation sites of SLP-76. The P-I region deletion disrupted Vav association and reduced SLP-76-associated kinase activity. A smaller deletion within the P-I region, which does not impair PLC-gamma 1 activation, did not impair the association with Vav, but reduced SLP-76-associated kinase activity. These results provide new insight into the multiple roles of SLP-76 and the functional importance of its interactions with other signaling proteins.