Nguyen Chi Huu, Stadler Serena, Brenner Stefan, Huttary Nicole, Krieger Sigurd, Jäger Walter, Dolznig Helmut, Krupitza Georg
Department for Clinical Pharmacy and Diagnostics, Faculty of Life Sciences, University of Vienna, Althanstrasse 14, A-1090 Vienna, Austria.
Clinical Institute of Pathology, Medical University of Vienna, Waehringer Guertel 18-20, A-1090 Vienna, Austria.
Br J Cancer. 2016 Jul 26;115(3):364-70. doi: 10.1038/bjc.2016.201. Epub 2016 Jun 30.
The arachidonic acid metabolite 12(S)-HETE is suspected to enhance metastatic spread by inducing cancer cell- and lymph endothelial cell (LEC) motility. However, the molecular mechanisms leading to 12(S)-HETE-triggered cell migration are still elusive.
To delineate the signalling pathways involved in 12(S)-HETE-mediated migration, inhibitors against RHO and ROCK, and specific siRNAs downregulating 12(S)-HETE receptor (12-HETER) and myosin light chain 2 (MLC2) were used. The breaching of the endothelial barrier was investigated by an assay measuring tumour spheroid-triggered 'circular chemorepellent-induced defects' (CCIDs), and respective signal transduction was elucidated by western blotting.
We provide evidence that 12(S)-HETE phosphorylated (and activated) MLC2, which regulates actin/myosin-based contraction. MLC2 activation was found to be essential for LEC retraction and CCID formation. Furthermore, we show that 12(S)-HETE activated a 12-HETER-RHO-ROCK-MYPT signalling cascade to induce MLC2 function.
Signalling via this pathway is described for this metabolite for the first time. This may provide potential targets for the intervention of metastatic colonisation.
花生四烯酸代谢产物12(S)-羟基二十碳四烯酸（12(S)-HETE）被怀疑通过诱导癌细胞和淋巴管内皮细胞（LEC）的运动性来增强转移扩散。然而，导致12(S)-HETE触发细胞迁移的分子机制仍不清楚。
为了阐明参与12(S)-HETE介导迁移的信号通路，使用了针对RHO和ROCK的抑制剂，以及下调12(S)-HETE受体（12-HETER）和肌球蛋白轻链2（MLC2）的特异性小干扰RNA（siRNA）。通过测量肿瘤球状体触发的“圆形化学排斥诱导缺陷”（CCID）的试验研究内皮屏障的破坏，并通过蛋白质印迹法阐明各自的信号转导。
我们提供的证据表明，12(S)-HETE使调节基于肌动蛋白/肌球蛋白收缩的MLC2磷酸化（并激活）。发现MLC2激活对于LEC收缩和CCID形成至关重要。此外，我们表明12(S)-HETE激活了12-HETER-RHO-ROCK-MYPT信号级联反应以诱导MLC2功能。
首次描述了该代谢产物通过该途径的信号传导。这可能为转移性定植的干预提供潜在靶点。
