Role of ACK1 (activated Cdc42-associated kinase 1) in adaptor SLP-76 signalling in T-cells

我们NSERC研究计划的目标是了解控制下游功能的抗原-受体近端信号事件，如钙动员和接头和激酶的聚集。介导T细胞激活的最初事件是由近端的酪氨酸激酶如p56lck和ZAP-70以及它们对接头蛋白的磷酸化所介导的。转接子反过来调节细胞内整合信号所需的复合体的形成。SLP-76(含76 kDa SH2结构域的白细胞蛋白)是一种关键的适配蛋白，它在促进T细胞的活化和发育中起着积极的作用。SLP-76含有一个无菌α基序(SAM)结构域，我们先前发现该结构域介导SLP-76二聚体和寡聚体的形成。我们还表明，酪氨酸激酶AcK1CDC42(细胞分裂周期42)激活的酪氨酸激酶1(AcK1或TNK2)与SLP-76结合，SLP-76 SAM结构域的丢失破坏了结合。TcR诱导磷脂酶C(PLCZAP-1)活化和钙动员的经典模型涉及另一种激酶γ-70及其磷酸化及其对白细胞介素2诱导的T细胞激酶的募集到SLP-76。我们发现，AcK1的表达诱导了SLP-76的磷酸化和钙内流，因此，SLP-76-AcK1模块可能是激活T细胞内钙动员和F-肌动蛋白组装的第二个模块途径。为了验证这一模型，我们提出：目标1：检测AcK1与SLP-76结合是否需要SAM结构域介导的寡聚。我们发现AcK1可以磷酸化这些关键酪氨酸上的SLP-76，并通过其SAM结构域与SLP-76结合。本文介绍了调节T细胞中SLP-76功能的新机制。

The goal of our NSERC research program is to understand the antigen-receptor proximal signaling events that control downstream functions such as calcium mobilization and the aggregation of adaptors and kinases. The initial events that mediate activation of T-cells are mediated by proximal tyrosine kinases such as p56lck and ZAP-70 and their phosphorylation of adaptor proteins. Adaptors in turn mediate the formation of complexes needed to integrate signalling in cells. One key adaptor protein is SLP-76 (SH2 domain containing leukocyte protein of 76kDa) which plays a positive role in promoting T cell activation and development. SLP-76 contains a sterile­ alpha motif (SAM) domain that we previously showed mediates SLP-76 dimer and oligomer formation. We have also shown that the tyrosine kinase ACK1 Cdc42 (cell division cycle 42)-activated tyrosine kinase 1 (ACK1 or Tnk2) binds to SLP-76 and that the loss of the SLP-76 SAM domain disrupts binding. The classic model for TCR induction of phospholipase C (PLCγ1) activation and calcium mobilization involves another kinase ZAP-70 and its phosphorylation and its recruitment of interleukin-2-inducible T-cell kinase (ITK) to SLP-76. We found that ACK1 expression induced SLP-76 phosphorylation and calcium influx, and as such, the SLP-76-ACK1 module could represent a second modular pathway for the activation calcium mobilization and F-actin assembly in T-cells. To test this model, we propose: Aim 1: to examine whether the SAM domain mediated oligomerization is needed for the binding of ACK1 to SLP-76. We have discovered that ACK1 can phosphorylate SLP-76 on these key tyrosines and binds to SLP-76 via its SAM domain. This introduces are novel mechanism to regulate SLP-76 function in T-cells.