Morgan M M, Labno C M, Van Seventer G A, Denny M F, Straus D B, Burkhardt J K
Department of Pathology, University of Chicago, Chicago, IL 60637, USA.
J Immunol. 2001 Nov 15;167(10):5708-18. doi: 10.4049/jimmunol.167.10.5708.
The formation of a conjugate between a T cell and an APC requires the activation of integrins on the T cell surface and remodeling of cytoskeletal elements at the cell-cell contact site via inside-out signaling. The early events in this signaling pathway are not well understood, and may differ from the events involved in adhesion to immobilized ligands. We find that conjugate formation between Jurkat T cells and EBV-B cells presenting superantigen is mediated by LFA-1 and absolutely requires Lck. Mutations in the Lck kinase, Src homology 2 or 3 domains, or the myristoylation site all inhibit conjugation to background levels, and adhesion cannot be restored by the expression of Fyn. However, ZAP-70-deficient cells conjugate normally, indicating that Lck is required for LFA-1-dependent adhesion via other downstream pathways. Several drugs that inhibit T cell adhesion to ICAM-1 immobilized on plastic, including inhibitors of mitogen-activated protein/extracellular signal-related kinase kinase, phosphatidylinositol-3 kinase, and calpain, do not inhibit conjugation. Inhibitors of phospholipase C and protein kinase C block conjugation of both wild-type and ZAP-70-deficient cells, suggesting that a phospholipase C that does not depend on ZAP-70 for its activation is involved. These results are not restricted to Jurkat T cells; Ag-specific primary T cell blasts behave similarly. Although the way in which Lck signals to enhance LFA-1-dependent adhesion is not clear, we find that cells lacking functional Lck fail to recruit F-actin and LFA-1 to the T cell:APC contact site, whereas ZAP-70-deficient cells show a milder phenotype characterized by disorganized actin and LFA-1 at the contact site.
T细胞与抗原呈递细胞（APC）之间形成共轭物需要激活T细胞表面的整合素，并通过外向内信号传导重塑细胞-细胞接触部位的细胞骨架成分。该信号通路中的早期事件尚未完全了解，可能与粘附固定化配体所涉及的事件不同。我们发现，Jurkat T细胞与呈递超抗原的EBV-B细胞之间的共轭物形成是由淋巴细胞功能相关抗原-1（LFA-1）介导的，并且绝对需要Lck。Lck激酶、Src同源2或3结构域或肉豆蔻酰化位点的突变均将共轭作用抑制至背景水平，并且Fyn的表达无法恢复粘附作用。然而，缺乏ζ链相关蛋白激酶70（ZAP-70）的细胞能正常形成共轭物，这表明Lck是通过其他下游途径实现LFA-1依赖性粘附所必需的。几种抑制T细胞与固定在塑料上的细胞间粘附分子-1（ICAM-1）粘附的药物，包括丝裂原活化蛋白/细胞外信号调节激酶激酶抑制剂、磷脂酰肌醇-3激酶抑制剂和钙蛋白酶抑制剂，均不抑制共轭物形成。磷脂酶C和蛋白激酶C抑制剂可阻断野生型和ZAP-70缺陷型细胞的共轭作用，这表明存在一种不依赖ZAP-70激活的磷脂酶C参与其中。这些结果并不局限于Jurkat T细胞；抗原特异性原代T细胞母细胞表现出相似的行为。尽管Lck增强LFA-1依赖性粘附的信号传导方式尚不清楚，但我们发现缺乏功能性Lck的细胞无法将丝状肌动蛋白（F-actin）和LFA-1募集至T细胞与APC的接触部位，而缺乏ZAP-70的细胞表现出较温和的表型，其特征是接触部位的肌动蛋白和LFA-1排列紊乱。
