Inhibition of ZAP-70 kinase activity via an analog-sensitive allele blocks T cell receptor and CD28 superagonist signaling.
Inhibition of ZAP-70 kinase activity via an analog-sensitive allele blocks T cell receptor and CD28 superagonist signaling.
复制标题
DOI：
10.1074/jbc.m709000200
复制
发表时间：
2008-05-30
期刊：
影响因子：
--
通讯作者：
Weiss A
中科院分区：
文献类型：
--
作者：
Levin SE;Zhang C;Kadlecek TA;Shokat KM;Weiss A
ZAP-70是T细胞抗原受体（TCR）信号传导所需的细胞质蛋白酪氨酸激酶。缺乏ZAP-70的小鼠和人类都不能发育功能性T细胞，因此证明了其对T细胞发育和功能的必要性。目前还没有高度特异性的，细胞渗透性的，小分子抑制剂ZAP-70，因此，我们产生了一个突变ZAP-70等位基因，保留激酶活性，但对抑制敏感的muplain特异性抑制剂。我们在Jurkat T细胞系中验证了化学遗传抑制剂系统，其中抑制剂阻断了表达类似物敏感等位基因的细胞中的ZAP-70依赖性TCR信号传导。有趣的是，该抑制剂还消除了CD 28超激动剂信号传导，从而证明了该系统在剖析T细胞活化的替代机制中对ZAP-70的需求方面的实用性。因此，我们开发了第一种抑制细胞中ZAP-70的特异性化学手段，这为研究ZAP-70在T细胞中的功能提供了有价值的工具。
ZAP-70 is a cytoplasmic protein tyrosine kinase that is required for T cell antigen receptor (TCR) signaling. Both mice and humans deficient in ZAP-70 fail to develop functional T cells, thus demonstrating its necessity for T cell development and function. There is currently no highly specific, cell-permeable, small molecule inhibitor for ZAP-70; therefore, we generated a mutant ZAP-70 allele that retains kinase activity but is sensitive to inhibition by a mutant-specific inhibitor. We validated the chemical genetic inhibitor system in Jurkat T cell lines, where the inhibitor blocked ZAP-70-dependent TCR signaling in cells expressing the analog-sensitive allele. Interestingly, the inhibitor also ablated CD28 superagonist signaling, thereby demonstrating the utility of this system in dissecting the requirement for ZAP-70 in alternative mechanisms of T cell activation. Thus, we have developed the first specific chemical means of inhibiting ZAP-70 in cells, which serves as a valuable tool for studying the function of ZAP-70 in T cells.