Modulation of Lck function through multisite docking to T cell-specific adapter protein
Modulation of Lck function through multisite docking to T cell-specific adapter protein
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DOI：
10.1074/jbc.m800871200
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发表时间：
2008-08-08
影响因子：
4.8
通讯作者：
Sundvold-Gjerstad, Vibeke
中科院分区：
文献类型：
--
作者：
Granum, Stine;Andersen, Thorny Cesilie Bie;Sundvold-Gjerstad, Vibeke
由SH 2D 2A基因编码的T细胞特异性衔接蛋白（TSAd）通过其C末端与Lck相互作用，从而调节Lck活性。在这里，我们映射Lck磷酸化和相互作用的TSAd网站，并评估其功能的重要性。三个C-末端TSAd酪氨酸Tyr（280）、Tyr（290）和Tyr（305）被Lck磷酸化，并作为Lck Src同源2（SH 2）结构域的对接位点。TSAd Tyr（P）（280）和Tyr（P）（290）磷酸肽与分离的Lck SH 2结构域的结合亲和力与Lck Tyr（P）（505）磷酸肽观察到的亲和力相似，而TSAd Tyr（P）（305）肽显示出10倍更高的亲和力。TSAd上富含脯氨酸的Lck SH 3结合位点以及Lck SH 2结构域是Lck有效地酪氨酸磷酸化TSAd所必需的。TSAd上Lck SH 2和Lck SH 3的相互作用位点对于TSAd介导的近端TCR信号传导事件的调节是必需的。我们发现20-30%的TSAd分子在活化的T细胞中被磷酸化，并且在这样的细胞中TSAd与Lck分子的比例类似于1：1。因此，在活化的T细胞中，相当数量的Lck分子可能潜在地被TSAd接合。总之，Lck结合TSAd脯氨酸和磷酸化，并与三个C-末端TSAd酪氨酸相互作用。我们建议，通过与LCK的多价相互作用，TSAd转移LCK磷酸化其他底物，从而通过底物竞争调节其功能活性。
T cell-specific adapter protein (TSAd), encoded by the SH2D2A gene, interacts with Lck through its C terminus and thus modulates Lck activity. Here we mapped Lck phosphorylation and interaction sites on TSAd and evaluated their functional importance. The three C-terminal TSAd tyrosines Tyr(280), Tyr(290), and Tyr(305) were phosphorylated by Lck and functioned as docking sites for the Lck Src homology 2 (SH2) domain. Binding affinities of the TSAd Tyr(P)(280) and Tyr(P)(290) phosphopeptides to the isolated Lck SH2 domain were similar to that observed for the Lck Tyr(P)(505) phosphopeptide, whereas the TSAd Tyr(P)(305) peptide displayed a 10-fold higher affinity. The proline-rich Lck SH3-binding site on TSAd as well as the Lck SH2 domain were required for efficient tyrosine phosphorylation of TSAd by Lck. Interaction sites on TSAd for both Lck SH2 and Lck SH3 were necessary for TSAd-mediated modulation of proximal TCR signaling events. We found that 20-30% of TSAd molecules are phosphorylated in activated T cells and that the proportion of TSAd to Lck molecules in such cells is similar to 1:1. Therefore, in activated T cells, a considerable number of Lck molecules may potentially be engaged by TSAd. In conclusion, Lck binds to TSAd prolines and phosphorylates and interacts with the three C-terminal TSAd tyrosines. We propose that through multivalent interactions with Lck, TSAd diverts Lck from phosphorylating other substrates, thus modulating its functional activity through substrate competition.