BACKGROUND Cytokine-dependent activation of signalling pathways is tightly orchestrated. The spatiotemporal activation of signalling pathways dictates the specific physiological responses to cytokines. Dysregulated signalling accounts for neoplastic, developmental, and inflammatory diseases. Grb2-associated binder (Gab) family proteins are multi-site docking proteins, which expand cytokine-induced signal transduction in a spatial- and time-dependent manner by coordinating the recruitment of proteins involved in mitogen activated protein kinase (MAPK)/extracellular-signal regulated kinase (ERK) and phosphatidyl-inositol-3-kinase (PI3K) signalling. Interaction of Gab family proteins with these signalling proteins determines strength, duration and localization of active signalling cascades. However, the underlying molecular mechanisms of signal orchestration by Gab family proteins in IL-6-induced signalling are only scarcely understood. METHODS We performed kinetic analyses of interleukin-6 (IL-6)-induced MAPK activation and analysed downstream responses. We compared signalling in wild-type cells, Gab1 knock-out cells, those reconstituted to express Gab1 mutants, and cells expressing gp130 receptors or receptor mutants. RESULTS Interleukin-6-induced MAPK pathway activation can be sub-divided into an early Gab1-independent and a subsequent Gab1-dependent phase. Early Gab1-independent MAPK activation is critical for the subsequent initiation of Gab1-dependent amplification of MAPK pathway activation and requires binding of SH2 domain-containing phosphatase 2 (SHP2) to the interleukin-6 receptor complex. Subsequent and coordinated recruitment of Grb2 and SHP2 to Gab1 is essential for Gab1-dependent amplification of IL-6-induced late MAPK pathway activation and subsequent gene expression. CONCLUSIONS Overall, we elaborated the molecular requirements for Gab1-dependent, spatiotemporal orchestration of interleukin-6-dependent MAPK signalling. We discriminated IL-6-induced Gab1-independent, early activation of MAPK signalling and Gab1-dependent, sustained activation of MAPK signalling.

中文翻译：

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多站点对接蛋白Grb2相关的粘合剂1（Gab1）以SHP2，Grb2和时间依赖性方式增强白介素6诱导的MAPK途径激活。

背景技术紧密地精心设计信号传导途径的细胞因子依赖性激活。信号通路的时空激活决定了对细胞因子的特定生理反应。信号失调解释了肿瘤性，发育性和炎性疾病。Grb2相关联的结合蛋白（Gab）家族蛋白是多部位对接蛋白，通过协调募集参与有丝分裂原活化蛋白激酶（MAPK）/细胞外蛋白的蛋白的募集，以空间和时间依赖的方式扩展细胞因子诱导的信号转导。信号调节激酶（ERK）和磷脂酰肌醇3激酶（PI3K）信号传导。Gab家族蛋白与这些信号蛋白的相互作用决定了活性信号级联的强度，持续时间和位置。然而，很少了解Gab家族蛋白在IL-6诱导的信号转导过程中进行信号编排的潜在分子机制。方法我们对白介素6（IL-6）诱导的MAPK活化进行了动力学分析，并分析了下游反应。我们比较了野生型细胞，Gab1敲除细胞，重组表达Gab1突变体的细胞和表达gp130受体或受体突变体的细胞中的信号传导。结果白介素6诱导的MAPK途径激活可细分为早期Gab1无关和随后的Gab1依赖性。早期的不依赖Gab1的MAPK激活对于随后开始的不依赖Gab1的MAPK途径激活的扩增至关重要，并且需要将包含SH2域的磷酸酶2（SHP2）与白介素6受体复合物结合。随后和协调募集Grb2和SHP2到Gab1对于依赖Gab1的IL-6诱导的晚期MAPK途径激活和后续基因表达的扩增至关重要。结论总体而言，我们阐述了白介素6依赖性MAPK信号转导的Gab1依赖性，时空编排的分子要求。我们区分了IL-6诱导的不依赖Gab1的MAPK信号的早期激活和不依赖Gab1的MAPK信号的持续激活。