Institute of Physiological Chemistry, University of Ulm, Albert-Einstein-Allee 11, 89081 Ulm, Germany.
J Biol Chem. 2011 Mar 4;286(9):7522-34. doi: 10.1074/jbc.M110.155895. Epub 2011 Jan 3.
T cell receptor (TCR) ligation induces increased diacylglycerol and Ca(2+) levels in T cells, and both secondary messengers are crucial for TCR-induced nuclear factor of activated T cells (NF-AT) and NF-κB signaling pathways. One prominent calcium-dependent enzyme involved in the regulation of NF-AT and NF-κB signaling pathways is the protein phosphatase calcineurin. However, in contrast to NF-AT, which is directly dephosphorylated by calcineurin, the molecular basis of the calcium-calcineurin dependence of the TCR-induced NF-κB activity remains largely unknown. Here, we demonstrate that calcineurin regulates TCR-induced NF-κB activity by controlling the formation of a protein complex composed of Carma1, Bcl10, and Malt1 (CBM complex). For instance, increased calcium levels induced by ionomycin or thapsigargin augmented the phorbol 12-myristate 13-acetate-induced formation of the CBM complex and activation of NF-κB, whereas removal of calcium by the calcium chelator EGTA-acetoxymethyl ester (AM) attenuated both processes. Furthermore, inhibition of the calcium-dependent phosphatase calcineurin with the immunosuppressive agent cyclosporin A (CsA) or FK506 as well as siRNA-mediated knockdown of calcineurin A strongly affected the PMA + ionomycin- or anti-CD3 + CD28-induced CBM complex assembly. Mechanistically, the positive effect of calcineurin on the CBM complex formation seems to be linked to a dephosphorylation of Bcl10. For instance, Bcl10 was found to be hyperphosphorylated in Jurkat T cells upon treatment with CsA or EGTA-AM, and calcineurin dephosphorylated Bcl10 in vivo and in vitro. Furthermore, we show here that calcineurin A interacts with the CBM complex. In summary, the evidence provided here argues for a previously unanticipated role of calcineurin in CBM complex formation as a molecular basis of the inhibitory function of CsA or FK506 on TCR-induced NF-κB activity.
T 细胞受体 (TCR) 交联诱导 T 细胞中二酰甘油和 Ca(2+)水平升高，这两种第二信使对于 TCR 诱导的核因子活化 T 细胞 (NF-AT) 和 NF-κB 信号通路至关重要。参与调节 NF-AT 和 NF-κB 信号通路的一种突出的钙依赖性酶是蛋白磷酸酶钙调神经磷酸酶。然而，与直接被钙调神经磷酸酶去磷酸化的 NF-AT 不同，TCR 诱导的 NF-κB 活性的钙-钙调神经磷酸酶依赖性的分子基础在很大程度上仍然未知。在这里，我们证明钙调神经磷酸酶通过控制由 Carma1、Bcl10 和 Malt1 组成的蛋白质复合物（CBM 复合物）的形成来调节 TCR 诱导的 NF-κB 活性。例如，离子霉素或 thapsigargin 诱导的 Ca(2+)水平增加增强了佛波醇 12-肉豆蔻酸 13-醋酸酯诱导的 CBM 复合物形成和 NF-κB 的激活，而 EGTA-乙氧羰基甲酯 (AM) 去除 Ca(2+)则减弱了这两个过程。此外，免疫抑制剂环孢素 A (CsA) 或 FK506 抑制钙依赖性磷酸酶钙调神经磷酸酶，以及 siRNA 介导的钙调神经磷酸酶 A 敲低强烈影响 PMA +离子霉素或抗 CD3 + CD28 诱导的 CBM 复合物组装。从机制上讲，钙调神经磷酸酶对 CBM 复合物形成的正效应似乎与 Bcl10 的去磷酸化有关。例如，在 CsA 或 EGTA-AM 处理后，Jurkat T 细胞中的 Bcl10 被发现过度磷酸化，并且钙调神经磷酸酶在体内和体外使 Bcl10 去磷酸化。此外，我们在这里表明钙调神经磷酸酶 A 与 CBM 复合物相互作用。总之，这里提供的证据表明钙调神经磷酸酶在 CBM 复合物形成中具有以前未预料到的作用，是 CsA 或 FK506 对 TCR 诱导的 NF-κB 活性的抑制功能的分子基础。
