Bisel Blaine, Calamai Martino, Vanzi Francesco, Pavone Francesco Saverio
European Laboratory for Non-linear Spectroscopy (LENS), University of Florence, Sesto Fiorentino, Italy.
PLoS One. 2013 Dec 2;8(12):e80446. doi: 10.1371/journal.pone.0080446. eCollection 2013.
Cell polarization is a process of coordinated cellular rearrangements that prepare the cell for migration. GM1 is synthesized in the Golgi apparatus and localized in membrane microdomains that appear at the leading edge of polarized cells, but the mechanism by which GM1 accumulates asymmetrically is unknown. The Golgi apparatus itself becomes oriented toward the leading edge during cell polarization, which is thought to contribute to plasma membrane asymmetry. Using quantitative image analysis techniques, we measure the extent of polarization of the Golgi apparatus and GM1 in the plasma membrane simultaneously in individual cells subject to a wound assay. We find that GM1 polarization starts just 10 min after stimulation with growth factors, while Golgi apparatus polarization takes 30 min. Drugs that block Golgi polarization or function have no effect on GM1 polarization, and, conversely, inhibiting GM1 polarization does not affect Golgi apparatus polarization. Evaluation of Golgi apparatus and GM1 polarization in single cells reveals no correlation between the two events. Our results indicate that Golgi apparatus and GM1 polarization are controlled by distinct intracellular cascades involving the Ras/Raf/MEK/ERK and the PI3K/Akt/mTOR pathways, respectively. Analysis of cell migration and invasion suggest that MEK/ERK activation is crucial for two dimensional migration, while PI3K activation drives three dimensional invasion, and no cumulative effect is observed from blocking both simultaneously. The independent biochemical control of GM1 polarity by PI3K and Golgi apparatus polarity by MEK/ERK may act synergistically to regulate and reinforce directional selection in cell migration.
细胞极化是一个协调细胞重排的过程，为细胞迁移做好准备。GM1在高尔基体中合成，并定位于极化细胞前缘出现的膜微区，但GM1不对称积累的机制尚不清楚。在细胞极化过程中，高尔基体自身会朝向细胞前缘定向，这被认为有助于质膜不对称性的形成。我们使用定量图像分析技术，在进行伤口试验的单个细胞中同时测量高尔基体的极化程度和质膜中GM1的极化程度。我们发现，GM1极化在生长因子刺激后仅10分钟就开始了，而高尔基体极化则需要30分钟。阻断高尔基体极化或功能的药物对GM1极化没有影响，反之，抑制GM1极化也不影响高尔基体极化。对单个细胞中高尔基体和GM1极化的评估显示，这两个事件之间没有相关性。我们的结果表明，高尔基体和GM1极化分别由涉及Ras/Raf/MEK/ERK和PI3K/Akt/mTOR途径的不同细胞内级联反应控制。对细胞迁移和侵袭的分析表明，MEK/ERK激活对于二维迁移至关重要，而PI3K激活驱动三维侵袭，同时阻断两者未观察到累积效应。PI3K对GM1极性的独立生化控制以及MEK/ERK对高尔基体极性的独立生化控制可能协同作用，以调节和加强细胞迁移中的方向选择。
