Department of Biochemistry, Graduate School of Medicine, University of the Ryukyus, Okinawa 903-0215, Japan; Department of Anesthesiology, Graduate School of Medicine, University of the Ryukyus, Okinawa 903-0215, Japan.
Department of Biochemistry, Graduate School of Medicine, University of the Ryukyus, Okinawa 903-0215, Japan.
Pulm Pharmacol Ther. 2018 Feb;48:53-61. doi: 10.1016/j.pupt.2017.09.002. Epub 2017 Sep 9.
Accumulating evidence indicates that epidermal growth factor receptor (EGFR) is desensitized by phosphorylation of serine 1047 (Ser1047). We and other groups have reported that stimulation of a receptor of tumor-necrosis factor α (TNFα) and Toll-like receptor 5 (TLR5) induced the phosphorylation of Ser1047 through activation of p38 mitogen-activated protein kinase (p38 MAPK) in cultured lung alveolar epithelial A549 cells. However, phosphorylation of EGFR at Ser1047 by stimulation of any G-protein coupled receptors (GPCRs) has not been reported in any cultured cells. In the present study, we first confirmed that A549 cells expressed bradykinin (BK) B2 receptor, and then, we examined whether BK treatment of A549 cells activated MAPKs and induced the phosphorylation of EGFR at Ser1047. Immunoblotting analysis and reporter gene assays indicated that BK activated the pathways of extracellular signal-regulated kinase (ERK) and p38 MAPK. Inhibitor studies suggested that G was mainly involved in the activation of ERK and p38 MAPK. We found that stimulation of the BK B2 receptor, but not the BK B1 receptor, induced phosphorylation of EGFR at Ser1047. Pharmacological experiments indicated that both ERK and p38 MAPK were involved in the phosphorylation of EGFR. These results strongly suggested that BK regulates EGFR functions in lung alveolar epithelial cells. In addition, we found that BK treatment increased the mRNA level of dual specificity MAPK phosphatase 5 (DUSP5) in an ERK-dependent manner, which suggested that a negative feedback mechanism of ERK existed in the cells.
越来越多的证据表明，表皮生长因子受体 (EGFR) 通过丝氨酸 1047（Ser1047）的磷酸化而脱敏。我们和其他小组已经报告说，通过刺激肿瘤坏死因子 α（TNFα）和 Toll 样受体 5（TLR5）的受体，p38 丝裂原激活蛋白激酶（p38 MAPK）的激活诱导了 Ser1047 的磷酸化 在培养的肺肺泡上皮 A549 细胞中。然而，任何 G 蛋白偶联受体（GPCR）刺激 EGFR 在 Ser1047 上的磷酸化在任何培养细胞中都没有报道过。在本研究中，我们首先证实 A549 细胞表达缓激肽（BK）B2 受体，然后，我们检查了 BK 处理 A549 细胞是否激活了 MAPKs 并诱导了 EGFR 在 Ser1047 上的磷酸化。免疫印迹分析和报告基因分析表明，BK 激活了细胞外信号调节激酶（ERK）和 p38 MAPK 途径。抑制剂研究表明 G 主要参与 ERK 和 p38 MAPK 的激活。我们发现，刺激 BK B2 受体而不是 BK B1 受体，诱导 EGFR 在 Ser1047 上的磷酸化。药理学实验表明，ERK 和 p38 MAPK 均参与 EGFR 的磷酸化。这些结果强烈表明 BK 调节肺肺泡上皮细胞中的 EGFR 功能。此外，我们发现 BK 处理以 ERK 依赖性方式增加了双特异性 MAPK 磷酸酶 5（DUSP5）的 mRNA 水平，这表明细胞中存在 ERK 的负反馈机制。
