The E3 ubiquitin ligase NEDD4 mediates cell migration signaling of EGFR in lung cancer cells.
The E3 ubiquitin ligase NEDD4 mediates cell migration signaling of EGFR in lung cancer cells.
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E3 泛素连接酶 NEDD4 介导肺癌细胞中 EGFR 的细胞迁移信号。
DOI：
10.1186/s12943-018-0784-2
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发表时间：
2018-02-19
期刊：
影响因子：
37.3
通讯作者：
Lin Q
中科院分区：
文献类型：
--
作者：
Shao G;Wang R;Sun A;Wei J;Peng K;Dai Q;Yang W;Lin Q
EGFR依赖性细胞迁移在肺癌进展中起重要作用。我们前期的研究发现HECT E3泛素连接酶NEDD 4与胃癌细胞的转移密切相关，并参与EGFR在胃癌细胞中的迁移和侵袭信号转导。然而，NEDD 4如何促进EGFR依赖性肺癌细胞迁移尚不清楚。本研究旨在阐明NEDD 4介导EGFR肺癌迁移信号的机制。慢病毒载体负载的NEDD 4 shRNA用于耗尽肺癌细胞系中的内源性NEDD 4。使用伤口愈合和transwell测定来确定NEDD 4敲低对EGFR依赖性或非依赖性肺癌细胞迁移的影响。通过免疫共沉淀法测定NEDD 4与活化的EGFR的结合。采用免疫组化（IHC）法检测63例肺腺癌组织中NEDD 4与EGFR或PTEN的共表达。分别采用GST-Uba pulldown试验、免疫印迹、免疫荧光染色和人组织蛋白酶B ELISA试验检测NEDD 4异位表达或敲低对PTEN泛素化和下调、AKT活化和溶酶体分泌的影响。组织蛋白酶B特异性抑制剂CA-074 Me用于评估组织蛋白酶B在肺癌细胞迁移中的作用。NEDD 4的敲除显著降低了非小细胞肺癌（NSCLC）细胞中EGF刺激的细胞迁移。免疫共沉淀实验发现，在EGF刺激下，NEDD 4与EGFR复合物结合，免疫组化染色显示NEDD 4与EGFR在肺腺癌肿瘤组织中共表达，提示NEDD 4可能通过与EGFR信号复合物相互作用介导肺癌细胞迁移。有趣的是，NEDD 4促进EGF诱导的组织蛋白酶B分泌，可能是通过溶酶体胞吐作用，因为NEDD 4的连接酶死亡突变体的过表达阻碍了溶酶体分泌，并且NEDD 4的敲低显著降低了EGF诱导的组织蛋白酶B的细胞外量。与NEDD 4的作用一致，组织蛋白酶B对于基础和EGF刺激的肺癌细胞迁移都是关键的。我们的研究提出了一种新的机制，EGFR促进肺癌细胞迁移，是由NEDD 4通过调节组织蛋白酶B分泌介导的。NEDD 4通过促进溶酶体分泌组织蛋白酶B介导EGFR肺癌细胞迁移信号。
EGFR-dependent cell migration plays an important role in lung cancer progression. Our previous study observed that the HECT E3 ubiquitin ligase NEDD4 is significantly correlated with tumor metastasis and required for migration and invasion signaling of EGFR in gastric cancer cells. However, how NEDD4 promotes the EGFR-dependent lung cancer cell migration is unknown. This study is to elucidate the mechanism by which NEDD4 mediates the EGFR lung cancer migration signaling. Lentiviral vector-loaded NEDD4 shRNA was used to deplete endogenous NEDD4 in lung cancer cell lines. Effects of the NEDD4 knockdown on the EGFR-dependent or independent lung cancer cell migration were determined using the wound-healing and transwell assays. Association of NEDD4 with activated EGFR was assayed by co-immunoprecipitation. Co-expression of NEDD4 with EGFR or PTEN was determined by immunohistochemical (IHC) staining in 63 lung adenocarcinoma tissue samples. Effects of NEDD4 ectopic expression or knockdown on PTEN ubiquitination and down-regulation, AKT activation and lysosomal secretion were examined using the GST-Uba pulldown assay, immunoblotting, immunofluorescent staining and a human cathepsin B ELISA assay respectively. The specific cathepsin B inhibitor CA-074Me was used for assessing the role of cathepsin B in lung cancer cell migration. Knockdown of NEDD4 significantly reduced EGF-stimulated cell migration in non-small cell lung carcinoma (NSCLC) cells. Co-immunoprecipitation assay found that NEDD4 is associated with EGFR complex upon EGF stimulation, and IHC staining indicates that NEDD4 is co-expressed with EGFR in lung adenocarcinoma tumor tissues, suggesting that NEDD4 might mediate lung cancer cell migration by interaction with the EGFR signaling complex. Interestingly, NEDD4 promotes the EGF-induced cathepsin B secretion, possibly through lysosomal exocytosis, as overexpression of the ligase-dead mutant of NEDD4 impedes lysosomal secretion, and knockdown of NEDD4 significantly reduced extracellular amount of cathepsin B induced by EGF. Consistent with the role of NEDD4, cathepsin B is pivotal for both basal and the EGF-stimulated lung cancer cell migration. Our studies propose a novel mechanism underlying the EGFR-promoted lung cancer cell migration that is mediated by NEDD4 through regulation of cathepsin B secretion. NEDD4 mediates the EGFR lung cancer cell migration signaling through promoting lysosomal secretion of cathepsin B.
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影响因子：
158.5
作者：
Lynch, TJ;Bell, DW;Haber, DA
通讯作者：
Haber, DA
影响因子：
3.3
作者：
Aoh, Quyen L.;Castle, Anna M.;Castle, J. David
通讯作者：
Castle, J. David
HECT E3 泛素连接酶 NEDD4 与 SQSTM1 相互作用并泛素化，以实现包涵体自噬。
影响因子：
4
作者：
Lin, Qiong;Dai, Qian;Yang, Wannian
通讯作者：
Yang, Wannian
影响因子：
4.5
作者：
Katz, M;Shtiegman, K;Yarden, Y
通讯作者：
Yarden, Y
DOI：
10.1073/pnas.0803233105
发表时间：
2008-06-24
影响因子：
11.1
作者：
Fouladkou, Fatemeh;Landry, Tamara;Rotin, Daniela
通讯作者：
Rotin, Daniela