Department of Obstetrics and Gynecology, St Joseph's Hospital and Medical Center, Phoenix, AZ 85004, USA.
Oncogene. 2010 Sep 2;29(35):4947-58. doi: 10.1038/onc.2010.240. Epub 2010 Jun 21.
Upregulation of epidermal growth factor receptor (EGFR) and subsequent increases in extracellular-regulated kinase (ERK) and Akt signaling are implicated in prostate cancer progression. Impaired endocytic downregulation of EGFR also contributes to oncogenic phenotypes such as metastasis. Thus, understanding the roles of divergent signaling pathways in the regulation of EGFR trafficking and EGFR-driven invasive migration may enable the development of more effective therapies. In this study, we use the human prostate cancer cell lines, DU145 and PC3, to investigate the effects of both the ERK and Akt pathways on epidermal growth factor (EGF)-mediated EGFR signaling, trafficking and cell motility. We show that DU145 and PC3 cells overexpress EGFR and migrate in a ligand (EGF)-dependent manner. Next, we show that pharmacological inhibition of ERK (but not Akt) signaling enhances EGF-induced EGFR activation, ubiquitination and downregulation, and may lead to enhanced receptor turnover. These findings negatively correlate with ERK-mediated threonine phosphorylation of EGFR, implicating it as a possible mechanism. Further, we uncover that EGF promotes disassembly of cell-cell junctions, downregulation of E-cadherin and upregulation of the transcriptional repressor, Snail, typical characteristics of epithelial-mesenchymal transition (EMT). These effects are dependent on activation of Akt, as inhibition of Akt signaling abolishes EGF/EGFR-driven cell migration and EMT. Knockdown of endogenous Snail also prevents EGFR-mediated downregulation of E-cadherin, EMT and cell migration. Surprisingly, inhibition of the ERK pathway augments EGFR-dependent motility, occurring concomitantly with elevation of EGF-induced Akt activity. Collectively, our results suggest that EGF-triggered ERK activation has profound feedback on EGFR signaling and trafficking by EGFR threonine phosphorylation, and Akt has a pivotal role in EGFR-mediated cell migration by activating EMT. More important, our results also suggest that therapeutic targeting of ERK signaling may have undesirable outcomes (for example, augmenting EGFR-driven motility).
表皮生长因子受体 (EGFR) 的上调及其随后细胞外调节激酶 (ERK) 和 Akt 信号的增加与前列腺癌的进展有关。EGFR 的内吞下调受损也有助于肿瘤发生的表型，如转移。因此，了解不同信号通路在调节 EGFR 转运和 EGFR 驱动的侵袭性迁移中的作用，可能能够开发更有效的治疗方法。在这项研究中，我们使用人前列腺癌细胞系 DU145 和 PC3 来研究 ERK 和 Akt 通路对表皮生长因子 (EGF) 介导的 EGFR 信号转导、转运和细胞迁移的影响。我们发现 DU145 和 PC3 细胞过表达 EGFR，并以配体 (EGF) 依赖的方式迁移。接下来，我们发现 ERK（而非 Akt）信号通路的药理学抑制增强了 EGF 诱导的 EGFR 激活、泛素化和下调，并且可能导致受体周转率增加。这些发现与 ERK 介导的 EGFR 苏氨酸磷酸化呈负相关，暗示其可能是一种机制。此外，我们发现 EGF 促进细胞-细胞连接的解体、E-钙黏蛋白的下调和转录抑制因子 Snail 的上调，这些都是上皮-间充质转化 (EMT) 的典型特征。这些效应依赖于 Akt 的激活，因为 Akt 信号通路的抑制消除了 EGF/EGFR 驱动的细胞迁移和 EMT。内源性 Snail 的敲低也阻止了 EGFR 介导的 E-钙黏蛋白下调、EMT 和细胞迁移。令人惊讶的是，ERK 通路的抑制增强了 EGFR 依赖性运动，同时伴随着 EGF 诱导的 Akt 活性的升高。总的来说，我们的结果表明，EGF 触发的 ERK 激活通过 EGFR 苏氨酸磷酸化对 EGFR 信号转导和转运产生深远的反馈，而 Akt 通过激活 EMT 在 EGFR 介导的细胞迁移中发挥关键作用。更重要的是，我们的结果还表明，ERK 信号通路的治疗靶向可能会产生不良后果（例如，增强 EGFR 驱动的运动）。
