Department of Biochemistry and Molecular Biology, Center for Digestive Diseases, Rochester, MN, USA.
Oncogene. 2012 Mar 8;31(10):1228-41. doi: 10.1038/onc.2011.329. Epub 2011 Aug 15.
Pancreatic ductal tumors invade local parenchyma and metastasize to distant organs. Src-mediated tyrosine kinase signaling pathways promote pancreatic ductal adenocarcinoma (PDAC) metastasis, though the molecular mechanisms supporting this invasive process are poorly understood and represent important and novel therapeutic targets. The large GTPase Dynamin 2 (Dyn2), a Src-kinase substrate, regulates membrane-cytoskeletal dynamics although it is yet to be defined if it contributes to tumor cell migration and invasion. Therefore, the goal of this study was to test if Dyn2 is upregulated in human pancreatic tumors and to define its role in cell migration and metastatic invasion using in vitro assays and nude mouse models. Histological analysis showed that 81% of 85 patients had elevated Dyn2 in PDAC. To test if Dyn2 overexpression alters metastatic properties of human pancreatic tumor cells, stable clones of BxPC-3 cells overexpressing either wild-type Dyn2 or a phosphorylation-deficient mutant Dyn2Y(231/597)F known to attenuate Dyn2 function, were generated and analyzed. Importantly, tumor cells overexpressing Dyn2 protruded lamellipodia at twice the rate, migrated faster (180%) and farther (2.5-fold greater distance) on glass and through transwell chambers (2-3-fold more cells through the filter) compared with cells expressing Dyn2Y(231/597)F or vector alone. Further, depletion of Dyn2 and dynamin inhibitors Myristyl trimethyl ammonium bromides and Dynasore significantly reduced cell migration, wound healing and invasion in transwell assays compared with controls. To test the metastatic potential conferred by increased Dyn2 expression, the BxPC-3 cell lines were implanted orthotopically into the pancreas of nude mice. Cells expressing Dyn2-green fluorescent protein exhibited a threefold increase in large distal tumors compared with cells expressing Dyn2Y(231/597)F or vector alone. Finally, histological analysis revealed that Dyn2 is upregulated in 60% of human metastatic pancreatic tumors. These findings are the first to implicate dynamin in any neoplastic condition and to directly demonstrate a role for this mechanoenzyme in invasive cell migration.
胰腺导管肿瘤侵犯局部实质并转移至远处器官。Src 介导的酪氨酸激酶信号通路促进胰腺导管腺癌 (PDAC) 的转移，尽管支持这种侵袭过程的分子机制尚未被充分理解，但却是重要的新治疗靶点。大型 GTP 酶 Dynamin 2 (Dyn2) 是 Src 激酶的底物，调节膜-细胞骨架动力学，尽管尚未确定它是否有助于肿瘤细胞迁移和侵袭。因此，本研究的目的是检测 Dyn2 是否在人类胰腺肿瘤中上调，并使用体外测定和裸鼠模型来定义其在细胞迁移和转移性侵袭中的作用。组织学分析显示，85 例患者中有 81%的 PDAC 中 Dyn2 升高。为了检测 Dyn2 过表达是否改变人胰腺肿瘤细胞的转移特性，生成并分析了稳定克隆的 BxPC-3 细胞，这些细胞过表达野生型 Dyn2 或磷酸化缺陷突变体 Dyn2Y(231/597)F，已知该突变体可减弱 Dyn2 功能。重要的是，与表达 Dyn2Y(231/597)F 或空载的细胞相比，过表达 Dyn2 的肿瘤细胞的片状伪足伸出速度快两倍，在玻璃上迁移速度更快 (快 180%)、更远 (距离增加 2.5 倍)，穿过 Transwell 室的细胞也多 (滤膜上多 2-3 倍的细胞)。此外，与对照相比，Dyn2 的耗竭和 dynamin 抑制剂 Myristyl trimethyl ammonium bromides 和 Dynasore 显著减少了 Transwell 测定中的细胞迁移、伤口愈合和侵袭。为了测试 Dyn2 表达增加赋予的转移潜能，将 BxPC-3 细胞系原位植入裸鼠胰腺中。与表达 Dyn2Y(231/597)F 或空载的细胞相比，表达 Dyn2-绿色荧光蛋白的细胞在大的远端肿瘤中增加了三倍。这些发现是首次将 dynamin 牵连到任何肿瘤疾病中，并直接证明了这种机械酶在侵袭性细胞迁移中的作用。
