Analysis of new binding proteins on N-terminal lesion of MEKK2,3,4

MEK激酶(Mekk)家族成员受多种细胞外刺激的调控，从生长因子到DNA损伤刺激，因此对细胞感知各种环境刺激非常重要。MEKK1参与了细胞迁移的调控，而粘着斑激酶(FAK)在细胞迁移和整合素信号转导中起重要作用。我们发现MEKK1和FAK是从小鼠成纤维细胞中免疫共沉淀出来的。此外，MEKK1和FAK之间的联系似乎在生理上是相关的，因为它被表皮生长因子(EGF)治疗而增强。将FAK定位于膜上也增强了其与MEKK1的结合，表明MEKK1定位于膜相关的亚细胞结构域，可能是局灶性粘连。有趣的是，在MEKK1缺陷的成纤维细胞中，胰岛素受体底物-I(IRS-1)的表达减少，这与早期在FAK缺陷的成纤维细胞中的发现相似。在MEKK1基因缺陷的细胞中，胰岛素样生长因子1(IGF-I)诱导的ERK激活也被抑制，但PI3K/Akt激活不受影响。虽然据报道整合素通过FAK介导的JNK激活来调节IRS-1基因的转录，但在MEKK1缺陷的细胞中，没有观察到纤维连接蛋白刺激的FAK、ERK或JNK的激活。MEKK1表达的重建恢复了IRS-1的表达和IGF-1诱导的ERK激活。综上所述，这些发现表明MEKK1在局灶性粘连中与FAK相互作用，并调节IRS-1的表达。

The MEK Kinase (MEKK) family members are regulated by a diverse array of extracellular stimuli ranging from growth factors to DNA damaging stimuli and so are important for the cell to sense exposure to various environmental stimuli. MEKK1 has been shown to contribute to the regulation of cell migration, while focal adhesion kinase (FAK) is a major player involved in both cell migration and integrin signaling. We show that MEKK1 and FAK are co-immunoprecipitated from mouse fibroblasts. Moreover, the association between MEKK1 and FAK appears to be physiologically relevant, as it is enhanced by treatment with epidermal growth factor (EGF). Targeting FAK to the membrane also enhanced its association with MEKK1, indicating MEKK1 to be localized to a membrane-related subcellular domain, perhaps focal adhesions. Interestingly, expression of insulin receptor substrate-I (IRS-1) was diminished in MEKK1-deficient fibroblasts, which is similar to an earlier finding in FAK-deficient fibroblasts. Insulin-like growth factor 1 (IGF-I)-induced ERK activation was also diminished in MEKK1-deficient cells, but PI3K/Akt activation was not. Although integrin reportedly regulates the transcription of the IRS-1 gene via FAK-mediated JNK activation, no impairment of fibronectin-stimulated activation of FAK, ERK or JNK was observed in MEKK1 -deficient cells. Reconstitution of MEKK1 expression restored IRS-1 expression as well as IGF-1-induced ERK activation. Taken together, these findings indicate that MEKK1 interacts with FAK in focal adhesions and regulates IRS-1 expression.