Jin J-K, Tien P-C, Cheng C-J, Song J H, Huang C, Lin S-H, Gallick G E
1] Department of Genitourinary Medical Oncology, Unit 18-4, David H. Koch Center for Applied Research of Genitourinary Cancers, The University of Texas M. D. Anderson Cancer Center, Houston, TX, USA [2] The Program in Cancer Metastasis, The University of Texas Graduate School of Biomedical Sciences at Houston, Houston, TX, USA.
Department of Genitourinary Medical Oncology, Unit 18-4, David H. Koch Center for Applied Research of Genitourinary Cancers, The University of Texas M. D. Anderson Cancer Center, Houston, TX, USA.
Oncogene. 2015 Apr 2;34(14):1811-21. doi: 10.1038/onc.2014.116. Epub 2014 May 5.
Talins are adaptor proteins that regulate focal adhesion signaling by conjugating integrins to the cytoskeleton. Talins directly bind integrins and are essential for integrin activation. We previously showed that β1 integrins are activated in metastatic prostate cancer (PCa) cells, increasing PCa metastasis to lymph nodes and bone. However, how β1 integrins are activated in PCa cells is unknown. In this study, we identified a novel mechanism of β1 integrin activation. Using knockdown experiments, we first demonstrated that talin1, but not talin2, is important in β1 integrin activation. We next showed that talin1 S425 phosphorylation, but not total talin1 expression, correlates with metastatic potential of PCa cells. Expressing a non-phosphorylatable mutant, talin1(S425A), in talin1-silenced PC3-MM2 and C4-2B4 PCa cells, decreased activation of β1 integrins, integrin-mediated adhesion, motility and increased the sensitivity of the cells to anoikis. In contrast, reexpression of the phosphorylation-mimicking mutant talin1(S425D) led to increased β1 integrin activation and generated biologic effects opposite to talin1(S425A) expression. In the highly metastatic PC3-MM2 cells, expression of a non-phosphorylatable mutant, talin1(S425A), in talin1-silenced PC3-MM2 cells, abolished their ability to colonize in the bone following intracardiac injection, while reexpression of phosphorylation-mimicking mutant talin1(S425D) restored their ability to metastasize to bone. Immunohistochemical staining demonstrated that talin S425 phosphorylation is significantly increased in human bone metastases when compared with normal tissues, primary tumors or lymph node metastases. We further showed that p35 expression, an activator of Cdk5, and Cdk5 activity were increased in metastatic tumor cells, and that Cdk5 kinase activity is responsible for talin1 phosphorylation and subsequent β1 integrin activation. Together, our study reveals Cdk5-mediated phosphorylation of talin1 leading to β1 integrin activation is a novel mechanism that increases metastatic potential of PCa cells.
踝蛋白是一种衔接蛋白,通过将整合素与细胞骨架结合来调节粘着斑信号传导。踝蛋白直接结合整合素,对整合素激活至关重要。我们之前表明,β1整合素在转移性前列腺癌细胞中被激活,增加了前列腺癌向淋巴结和骨的转移。然而,β1整合素在前列腺癌细胞中如何被激活尚不清楚。在本研究中,我们确定了一种β1整合素激活的新机制。通过敲低实验,我们首先证明踝蛋白1而非踝蛋白2在β1整合素激活中起重要作用。接下来我们表明,踝蛋白1的S425磷酸化而非总踝蛋白1表达与前列腺癌细胞的转移潜能相关。在踝蛋白1沉默的PC3-MM2和C4-2B4前列腺癌细胞中表达不可磷酸化的突变体踝蛋白1(S425A),可降低β1整合素的激活、整合素介导的粘附和迁移,并增加细胞对失巢凋亡的敏感性。相反,磷酸化模拟突变体踝蛋白1(S425D)的重新表达导致β1整合素激活增加,并产生与踝蛋白1(S425A)表达相反的生物学效应。在高转移性PC3-MM2细胞中,在踝蛋白1沉默的PC3-MM2细胞中表达不可磷酸化的突变体踝蛋白1(S425A),消除了它们经心内注射后在骨中定植的能力,而磷酸化模拟突变体踝蛋白1(S425D)的重新表达恢复了它们转移至骨的能力。免疫组织化学染色表明,与正常组织、原发性肿瘤或淋巴结转移相比,人骨转移灶中踝蛋白S425磷酸化显著增加。我们进一步表明,细胞周期蛋白依赖性激酶5(Cdk5)的激活剂p35表达和Cdk5活性在转移性肿瘤细胞中增加,并且Cdk5激酶活性负责踝蛋白1的磷酸化及随后的β1整合素激活。总之,我们的研究揭示Cdk5介导的踝蛋白1磷酸化导致β1整合素激活是一种增加前列腺癌细胞转移潜能的新机制。