Institute of Biomedical Research Cadiz (INIBICA), 11009 Cádiz, Spain.
Rheumatology Section, Unit of Orthopedic Surgery, Traumatology and Rheumatology, HUPM, 11009 Cádiz, Spain.
Cells. 2021 Feb 6;10(2):343. doi: 10.3390/cells10020343.
Intracellular signaling through the T cell receptor (TCR) is essential for T cell development and function. Proper TCR signaling requires the sequential activities of Lck and ZAP-70 kinases, which result in the phosphorylation of tyrosine residues located in the CD3 ITAMs and the LAT adaptor, respectively. LAT, linker for the activation of T cells, is a transmembrane adaptor protein that acts as a scaffold coupling the early signals coming from the TCR with downstream signaling pathways leading to cellular responses. The leukemic T cell line Jurkat and its derivative mutants J.CaM1.6 (Lck deficient) and J.CaM2 (LAT deficient) have been widely used to study the first signaling events upon TCR triggering. In this work, we describe the loss of LAT adaptor expression found in a subline of J.CaM1.6 cells and analyze cis-elements responsible for the LAT expression defect. This new cell subline, which we have called J.CaM1.7, can re-express LAT adaptor after Protein Kinase C (PKC) activation, which suggests that activation-induced LAT expression is not affected in this new cell subline. Contrary to J.CaM1.6 cells, re-expression of Lck in J.CaM1.7 cells was not sufficient to recover TCR-associated signals, and both LAT and Lck had to be introduced to recover activatory intracellular signals triggered after CD3 crosslinking. Overall, our work shows that the new LAT negative J.CaM1.7 cell subline could represent a new model to study the functions of the tyrosine kinase Lck and the LAT adaptor in TCR signaling, and their mutual interaction, which seems to constitute an essential early signaling event associated with the TCR/CD3 complex.
细胞内信号通过 T 细胞受体 (TCR) 对于 T 细胞的发育和功能至关重要。适当的 TCR 信号需要 Lck 和 ZAP-70 激酶的顺序活性，这分别导致位于 CD3 ITAMs 和 LAT 衔接子上的酪氨酸残基的磷酸化。LAT，T 细胞激活的链接器，是一种跨膜衔接蛋白，作为支架将来自 TCR 的早期信号与导致细胞反应的下游信号通路耦联。白血病 T 细胞系 Jurkat 及其衍生的突变体 J.CaM1.6（Lck 缺陷）和 J.CaM2（LAT 缺陷）已被广泛用于研究 TCR 触发后的第一个信号事件。在这项工作中，我们描述了在 J.CaM1.6 细胞的一个亚系中发现的 LAT 衔接子表达的丧失，并分析了负责 LAT 表达缺陷的顺式元件。我们将这个新的细胞亚系称为 J.CaM1.7，它可以在蛋白激酶 C（PKC）激活后重新表达 LAT 衔接子，这表明在这个新的细胞亚系中，激活诱导的 LAT 表达不受影响。与 J.CaM1.6 细胞相反，在 J.CaM1.7 细胞中重新表达 Lck 不足以恢复 TCR 相关信号，并且必须引入 LAT 和 Lck 以恢复 CD3 交联后触发的激活性细胞内信号。总体而言，我们的工作表明，新的 LAT 阴性 J.CaM1.7 细胞亚系可能代表一种新的模型，用于研究酪氨酸激酶 Lck 和 LAT 衔接子在 TCR 信号转导中的功能及其相互作用，这似乎构成了与 TCR/CD3 复合物相关的一个基本早期信号事件。
