INTERACTIONS OF P59(FYN) AND ZAP-70 WITH T-CELL RECEPTOR ACTIVATION MOTIFS - DEFINING THE NATURE OF A SIGNALING MOTIF
INTERACTIONS OF P59(FYN) AND ZAP-70 WITH T-CELL RECEPTOR ACTIVATION MOTIFS - DEFINING THE NATURE OF A SIGNALING MOTIF
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DOI：
10.1128/mcb.14.6.3729
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发表时间：
1994-06-01
影响因子：
5.3
通讯作者：
SHAW, AS
中科院分区：
文献类型：
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作者：
GAUEN, LKT;ZHU, YX;SHAW, AS
基于酪氨酸的激活基序是一个20-25个氨基酸的序列，包含在许多造血受体的细胞质结构域中，它本身就足以重建信号。该基序由两个YXXL/I序列组成，间隔约10个残基。这个基序发出信号的分子基础尚不清楚。在这里，我们证明了基于酪氨酸的激活基序是与酪氨酸激酶P59(Fyn)和ZAP-70结合所必需的，也是充分的，这表明与这些激酶的结合是该基序的一个一般特征。针对epsilon中存在的单一激活基序，我们分析了哪些基序对P59(Fyn)与ZAP-70的结合至关重要。令人惊讶的是，我们发现没有任何一个epsilon残基的单一突变导致P59(Fyn)关联的丧失。相反，epsilon激活基序的五个残基的单一突变取消了ZAP-70的结合。酪氨酸和紧随其后的亮氨酸或异亮氨酸残基都是关键的。酪氨酸之间的间距也很重要，因为两个残基的缺失破坏了ZAP-70的结合，尽管P59(Fyn)结合没有中断。因此，酪氨酸激活基序的大多数已定义特征都是ZAP-70结合所必需的。有趣的是，ZAP-70与基序的相互作用依赖于ZAP-70 SH2结构域和两个酪氨酸残基的存在，这表明ZAP-70与两个磷酸酪氨酸残基相互作用，两个SH2结构域的结合是协同的。此外，我们还证明了酪氨酸活化蛾之间的相互作用是直接的，并且需要事先对基序进行酪氨酸磷酸化。我们认为，酪氨酸激活基序对细胞的激活经历了四个不同的步骤：P59(Fyn)的结合、基序的磷酸化、ZAP-70的结合和ZAP-70激酶活性的激活。
The tyrosine-based activation motif is a 20- to 25-amino-acid sequence contained in the cytoplasmic domains of many hematopoietic receptors which is sufficient by itself to reconstitute signalling. This motif is characterized by two YXXL/I sequences separated by approximately 10 residues. The molecular basis of signalling by this motif is unknown. Here we demonstrate that the tyrosine-based activation motif is required and sufficient for association with the tyrosine kinases p59(fyn) and ZAP-70, suggesting that association with these kinases is a general feature of this motif. Focusing on the single activation motif present in epsilon, we analyzed which residues of the motif were critical for binding of p59(fyn) and ZAP-70. Surprisingly, we found that no single mutation of any residue of epsilon resulted in the loss of p59(fyn) association. In contrast, single mutations at five residues of the epsilon activating motif abrogated ZAP-70 binding. Both of the tyrosines and the leucine or isoleucine residues that follow them were critical. The spacing between the tyrosines was also important, as deletion of two residues disrupted binding of ZAP-70, although p59(fyn) binding was not disrupted. Most of the defined features of the tyrosine activation motif are therefore requirements for ZAP-70 binding. Interestingly, the interaction of ZAP-70 with the motif was dependent on the presence of both ZAP-70 SH2 domains and both of the tyrosine residues in the moth, suggesting that ZAP-70 interacts with two phosphotyrosine residues and that the binding of the two SH2 domains is cooperative. In addition, we demonstrate that the interaction between the tyrosine activation moth is direct and requires prior tyrosine phosphorylation of the motif. We propose that the activation of cells by the tyrosine activating motif occurs in four discrete steps: binding of p59(fyn), phosphorylation of the motif, binding of ZAP-70, and activation of ZAP-70 kinase activity.