Department of Neurology and Neurosurgery, Montreal Neurological Institute, McGill University, Montreal, Quebec, Canada.
PLoS One. 2011;6(9):e25408. doi: 10.1371/journal.pone.0025408. Epub 2011 Sep 28.
Deregulation of mechanisms that control cell motility plays a key role in tumor progression by promoting tumor cell dissemination. Secreted netrins and their receptors, Deleted in Colorectal Cancer (DCC), neogenin, and the UNC5 homologues, regulate cell and axon migration, cell adhesion, and tissue morphogenesis. Netrin and netrin receptor expression have previously been shown to be disrupted in invasive tumors, including glioblastoma. We determined that the human glioblastoma cell lines U87, U343, and U373 all express neogenin, UNC5 homologues, and netrin-1 or netrin-3, but only U87 cells express DCC. Using transfilter migration assays, we demonstrate DCC-dependent chemoattractant migration of U87 cells up a gradient of netrin-1. In contrast, U343 and U373 cells, which do not express DCC, were neither attracted nor repelled. Ectopic expression of DCC by U343 and U373 cells resulted in these cells becoming competent to respond to a gradient of netrin-1 as a chemoattractant, and also slowed their rate of spontaneous migration. Here, in addition to netrins' well-characterized chemotropic activity, we demonstrate an autocrine function for netrin-1 and netrin-3 in U87 and U373 cells that slows migration. We provide evidence that netrins promote the maturation of focal complexes, structures associated with cell movement, into focal adhesions. Consistent with this, netrin, DCC, and UNC5 homologues were associated with focal adhesions, but not focal complexes. Disrupting netrin or DCC function did not alter cell proliferation or survival. Our findings provide evidence that DCC can slow cell migration, and that neogenin and UNC5 homologues are not sufficient to substitute for DCC function in these cells. Furthermore, we identify a role for netrins as autocrine inhibitors of cell motility that promote focal adhesion formation. These findings suggest that disruption of netrin signalling may disable a mechanism that normally restrains inappropriate cell migration.
细胞运动调控机制的失调在肿瘤进展中起着关键作用，促进肿瘤细胞的扩散。分泌的 netrins 和它们的受体、Deleted in Colorectal Cancer (DCC)、neogenin 和 UNC5 同源物调节细胞和轴突迁移、细胞黏附和组织形态发生。netrin 和 netrin 受体的表达先前已被证明在侵袭性肿瘤中被破坏，包括神经胶质瘤。我们确定人神经胶质瘤细胞系 U87、U343 和 U373 均表达 neogenin、UNC5 同源物和 netrin-1 或 netrin-3，但只有 U87 细胞表达 DCC。使用跨膜迁移测定，我们证明 DCC 依赖的 U87 细胞沿 netrin-1 梯度的化学趋化性迁移。相比之下，不表达 DCC 的 U343 和 U373 细胞既不受吸引也不受排斥。U343 和 U373 细胞异位表达 DCC 导致这些细胞能够对 netrin-1 梯度作为化学引诱物做出反应，并减缓其自发迁移的速度。在这里，除了 netrins 的特征性化学趋性活性外，我们还证明了 U87 和 U373 细胞中 netrin-1 和 netrin-3 的自分泌功能可减缓迁移速度。我们提供的证据表明 netrins 促进了与细胞运动相关的焦点复合物结构向焦点黏附的成熟。与此一致的是，netrin、DCC 和 UNC5 同源物与焦点黏附物相关，而与焦点复合物无关。破坏 netrin 或 DCC 功能不会改变细胞增殖或存活。我们的研究结果提供了证据表明 DCC 可以减缓细胞迁移，并且 neogenin 和 UNC5 同源物不足以替代这些细胞中的 DCC 功能。此外，我们确定了 netrins 作为细胞运动的自分泌抑制剂的作用，促进了焦点黏附的形成。这些发现表明，netrin 信号的破坏可能会使一种通常限制不适当细胞迁移的机制失效。
