Ulivieri C, Peter A, Orsini E, Palmer E, Baldari C T
Department of Evolutionary Biology, University of Siena, Via Mattioli 4, 53100 Siena, Italy.
J Biol Chem. 2001 Feb 2;276(5):3574-80. doi: 10.1074/jbc.M008588200. Epub 2000 Oct 31.
A key role in the communication between the alphabetaTCR and the CD3/zeta complex is played by a specific motif within the connecting peptide domain of the TCR alpha chain (alpha-CPM). T cell hybridomas expressing an alpha-CPM-mutated TCR show a dramatic impairment in antigen-driven interleukin-2 production. This defect can be complemented by a calcium ionophore, indicating that activation of the calcium pathway is impaired. Several lines of evidence implicate Fyn in the regulation of calcium mobilization, at least in part through the activation of phospholipase Cgamma. Here we have investigated the potential involvement of Fyn in the TCR alpha-CPM signaling defect. Using T cell hybridomas expressing either a wild-type TCR or an alpha-CPM mutant, we show that Fyn fails to be activated by the mutant receptor following SEB binding and fails to generate tyrosine-phosphorylated Pyk2, a member of the focal adhesion kinase family. This defect correlated with an impairment in phospholipase Cgamma phosphorylation. Production of interlukin-2 and activation of the transcription factor NF-AT in response to triggering of the TCR alpha-CPM mutant with SEB were fully restored in the presence of constitutively active Fyn. Hence the signaling defect generated by the TCR alpha-CPM mutation results at least in part from an impaired coupling of the TCR.CD3 complex to Fyn activation.
TCRα链连接肽结构域（α-CPM）内的一个特定基序在αβTCR与CD3/ζ复合物之间的信号传导中发挥关键作用。表达α-CPM突变型TCR的T细胞杂交瘤在抗原驱动的白细胞介素-2产生方面表现出显著受损。这种缺陷可以通过钙离子载体来弥补，这表明钙信号通路的激活受损。几条证据表明Fyn参与钙动员的调节，至少部分是通过激活磷脂酶Cγ。在这里，我们研究了Fyn在TCRα-CPM信号缺陷中的潜在作用。使用表达野生型TCR或α-CPM突变体的T细胞杂交瘤，我们发现突变型受体在结合SEB后无法激活Fyn，也无法产生酪氨酸磷酸化的Pyk2（粘着斑激酶家族的一员）。这种缺陷与磷脂酶Cγ磷酸化受损相关。在组成型活性Fyn存在的情况下，用SEB触发TCRα-CPM突变体后，白细胞介素-2的产生和转录因子NF-AT的激活完全恢复。因此，TCRα-CPM突变产生的信号缺陷至少部分是由于TCR.CD3复合物与Fyn激活的偶联受损所致。
