TCR Triggering Induces the Formation of Lck-RACK1-Actinin-1 Multiprotein Network Affecting Lck Redistribution.
TCR Triggering Induces the Formation of Lck-RACK1-Actinin-1 Multiprotein Network Affecting Lck Redistribution.
复制标题
DOI：
10.3389/fimmu.2016.00449
复制
发表时间：
2016
影响因子：
7.3
通讯作者：
Filipp D
中科院分区：
文献类型：
--
作者：
Ballek O;Valečka J;Dobešová M;Broučková A;Manning J;Řehulka P;Stulík J;Filipp D
T细胞信号传导的启动关键取决于Src家族酪氨酸激酶成员Lck的功能。在T细胞抗原受体（TCR）触发后，Lck激酶活性诱导信号转导枢纽的成核，所述信号转导枢纽调节复杂信号网络的形成和细胞骨架重排。此外，Lck功能的传递需要快速和有针对性的膜再分布，但支撑这一过程的机制在很大程度上是未知的。为了深入了解这一过程，我们考虑了先前描述的蛋白质，这些蛋白质可以通过与激酶相互作用并调节其细胞内易位的能力来帮助这一过程。一个衔接蛋白，受体活化C激酶1（RACK 1），被选为一个可行的选择，其能力结合LCK和援助的过程中激活诱导的再分配LCK进行了评估。我们的显微镜观察表明，T细胞活化诱导Lck和RACK 1快速，伴随和短暂的共同重新分布到形成的免疫突触。与该观察结果一致，在原代CD 4 + T细胞中可检测到瞬时RACK 1-Lck复合物的形成，其最大水平在TCR-CD 4共聚集后10秒达到峰值。此外，RACK 1优先结合激酶活性pY 394 Lck的库，其与高分子量细胞级分共纯化。RACK 1-Lck复合物的形成依赖于Lck的功能性SH 2和SH 3结构域，并且包括瞬时结合复合物的几种其他信号传导和细胞骨架元件。值得注意的是，F-肌动蛋白交联蛋白α-辅肌动蛋白-1仅在存在激酶活性Lck的情况下与RACK 1结合，这表明RACK 1-pY 394 Lck-α-辅肌动蛋白-1复合物的形成充当将肌动蛋白细胞骨架捆绑与有效的TCR/CD 4触发偶联的信号模块。此外，用破坏微管网络的诺考达唑处理CD 4 + T细胞也阻断了RACK 1-Lck复合物的形成。重要的是，激活诱导的Lck再分布减少在原代CD 4 + T细胞的腺病毒介导的敲低RACK 1。这些结果表明，在T细胞中，RACK 1，作为一个多蛋白复合物的必要组成部分，在TCR接合，链接激酶活性Lck的细胞骨架网络的元素的结合，并影响Lck的亚细胞再分布。
The initiation of T-cell signaling is critically dependent on the function of the member of Src family tyrosine kinases, Lck. Upon T-cell antigen receptor (TCR) triggering, Lck kinase activity induces the nucleation of signal-transducing hubs that regulate the formation of complex signaling network and cytoskeletal rearrangement. In addition, the delivery of Lck function requires rapid and targeted membrane redistribution, but the mechanism underpinning this process is largely unknown. To gain insight into this process, we considered previously described proteins that could assist in this process via their capacity to interact with kinases and regulate their intracellular translocations. An adaptor protein, receptor for activated C kinase 1 (RACK1), was chosen as a viable option, and its capacity to bind Lck and aid the process of activation-induced redistribution of Lck was assessed. Our microscopic observation showed that T-cell activation induces a rapid, concomitant, and transient co-redistribution of Lck and RACK1 into the forming immunological synapse. Consistent with this observation, the formation of transient RACK1–Lck complexes were detectable in primary CD4+ T-cells with their maximum levels peaking 10 s after TCR–CD4 co-aggregation. Moreover, RACK1 preferentially binds to a pool of kinase active pY394Lck, which co-purifies with high molecular weight cellular fractions. The formation of RACK1–Lck complexes depends on functional SH2 and SH3 domains of Lck and includes several other signaling and cytoskeletal elements that transiently bind the complex. Notably, the F-actin-crosslinking protein, α-actinin-1, binds to RACK1 only in the presence of kinase active Lck suggesting that the formation of RACK1–pY394Lck–α-actinin-1 complex serves as a signal module coupling actin cytoskeleton bundling with productive TCR/CD4 triggering. In addition, the treatment of CD4+ T-cells with nocodazole, which disrupts the microtubular network, also blocked the formation of RACK1–Lck complexes. Importantly, activation-induced Lck redistribution was diminished in primary CD4+ T-cells by an adenoviral-mediated knockdown of RACK1. These results demonstrate that in T cells, RACK1, as an essential component of the multiprotein complex which upon TCR engagement, links the binding of kinase active Lck to elements of the cytoskeletal network and affects the subcellular redistribution of Lck.
登录
查看更多内容
影响因子：
7.3
作者：
Filipp D;Ballek O;Manning J
通讯作者：
Manning J
DOI：
10.1083/jcb.84.2.305
发表时间：
1980-02
期刊：
The Journal of cell biology
影响因子：
--
作者：
Hoessli D;Rungger-Brändle E;Jockusch BM;Gabbiani G
通讯作者：
Gabbiani G
影响因子：
5.3
作者：
Chang, BY;Conroy, KB;Cartwright, CA
通讯作者：
Cartwright, CA
影响因子：
4.4
作者：
Filipp, D;Leung, BL;Julius, M
通讯作者：
Julius, M
影响因子：
4.2
作者：
Henry, Whitney I.;Dubois, Juwen;Quick, Quincy A.
通讯作者：
Quick, Quincy A.