Department of Molecular Biology, Cell Biology, and Biochemistry, Brown University , Providence, Rhode Island 02912, United States.
Department of Chemistry, Brown University , Providence, Rhode Island 02912, United States.
J Proteome Res. 2017 Aug 4;16(8):2729-2742. doi: 10.1021/acs.jproteome.6b01026. Epub 2017 Jul 6.
Phospholipase C gamma 1 (PLC-γ1) occupies a critically important position in the T-cell signaling pathway. While its functions as a regulator of both Ca signaling and PKC-family kinases are well characterized, PLC-γ1's role in the regulation of early T-cell receptor signaling events is incompletely understood. Activation of the T-cell receptor leads to the formation of a signalosome complex between SLP-76, LAT, PLC-γ1, Itk, and Vav1. Recent studies have revealed the existence of both positive and negative feedback pathways from SLP-76 to the apical kinase in the pathway, Lck. To determine if PLC-γ1 contributes to the regulation of these feedback networks, we performed a quantitative phosphoproteomic analysis of PLC-γ1-deficient T cells. These data revealed a previously unappreciated role for PLC-γ1 in the positive regulation of Zap-70 and T-cell receptor tyrosine phosphorylation. Conversely, PLC-γ1 negatively regulated the phosphorylation of SLP-76-associated proteins, including previously established Lck substrate phosphorylation sites within this complex. While the positive and negative regulatory phosphorylation sites on Lck were largely unchanged, Tyr phosphorylation was elevated in Jgamma1. The data supports a model wherein Lck's targeting, but not its kinase activity, is altered by PLC-γ1, possibly through Lck Tyr phosphorylation and increased association of the kinase with protein scaffolds SLP-76 and TSAd.
磷酸脂酶 C 伽马 1（PLC-γ1）在 T 细胞信号通路中占据至关重要的位置。尽管它作为 Ca 信号和 PKC 家族激酶的调节剂的功能已经得到很好的描述，但 PLC-γ1 在调节早期 T 细胞受体信号事件中的作用还不完全清楚。T 细胞受体的激活导致 SLP-76、LAT、PLC-γ1、Itk 和 Vav1 之间信号体复合物的形成。最近的研究揭示了 SLP-76 到通路顶端激酶 Lck 之间存在正向和负向反馈途径。为了确定 PLC-γ1 是否有助于调节这些反馈网络，我们对 PLC-γ1 缺陷型 T 细胞进行了定量磷酸化蛋白质组学分析。这些数据揭示了 PLC-γ1 在正向调节 Zap-70 和 T 细胞受体酪氨酸磷酸化中的先前未被认识的作用。相反，PLC-γ1 负调节 SLP-76 相关蛋白的磷酸化，包括该复合物中先前建立的 Lck 底物磷酸化位点。虽然 Lck 的正向和负向调节磷酸化位点变化不大，但 Jgamma1 中的 Tyr 磷酸化水平升高。这些数据支持这样一种模型，即 Lck 的靶向作用，而不是其激酶活性，被 PLC-γ1 改变，可能通过 Lck Tyr 磷酸化和激酶与 SLP-76 和 TSAd 等蛋白支架的增加结合来实现。
