Sahuquillo A G, Roumier A, Teixeiro E, Bragado R, Alarcón B
Department of Immunology, Fundación Jiménez-Díaz, Avenida Reyes Católicos 2, 28040 Madrid, Spain.
J Exp Med. 1998 Apr 20;187(8):1179-92. doi: 10.1084/jem.187.8.1179.
We have previously shown that a tyrosine to leucine replacement in the transmembrane region of T cell receptor (TCR)-beta results in a deficient induction of CD95-L and apoptosis upon TCR triggering in a transfected T cell line. By contrast, interleukin (IL)-2 production and the expression of CD25 and CD69 were normally induced. Since the mutation in TCR-beta also resulted in impaired association of CD3-zeta, it was proposed that this chain is specifically required for the induction of apoptosis. We now show that the deficient induction of CD95-L and apoptosis does not derive from a general lower production of second messengers, since intracellular Ca2+ fluxes and tyrosine phosphorylation of total proteins were elicited at wild-type levels. Unlike in T cell clones stimulated with partial agonists, both p21 and p18 forms of tyrosine-phosphorylated CD3-zeta were detected, although the overall level of tyrosine-phosphorylated CD3-zeta was low. More strikingly, inducible association of ZAP70 to CD3-zeta was strongly inhibited, despite a normal induction of ZAP70 tyrosine phosphorylation. Finally, ZAP70 was not concentrated near the plasma membrane in the apoptosis-deficient cells. These results suggest that CD3-zeta is necessary for engagement of a specific signaling pathway leading to CD95-L expression that also needs the recruitment of ZAP70.
我们之前已经表明，在转染的T细胞系中，T细胞受体（TCR）-β跨膜区的酪氨酸替换为亮氨酸会导致TCR触发时CD95-L诱导不足和细胞凋亡。相比之下，白细胞介素（IL）-2的产生以及CD25和CD69的表达则正常诱导。由于TCR-β中的突变也导致CD3-ζ的缔合受损，因此有人提出该链是诱导细胞凋亡所特需的。我们现在表明，CD95-L和细胞凋亡诱导不足并非源于第二信使的普遍较低产生，因为细胞内Ca2+通量和总蛋白的酪氨酸磷酸化是在野生型水平引发的。与用部分激动剂刺激的T细胞克隆不同，虽然酪氨酸磷酸化的CD3-ζ的总体水平较低，但仍检测到了酪氨酸磷酸化的CD3-ζ的p21和p18形式。更引人注目的是，尽管ZAP70酪氨酸磷酸化正常诱导，但ZAP70与CD3-ζ的可诱导缔合却受到强烈抑制。最后，在缺乏细胞凋亡的细胞中，ZAP70没有集中在质膜附近。这些结果表明，CD3-ζ对于导致CD95-L表达的特定信号通路的参与是必需的，该信号通路还需要募集ZAP70。
