School of Pharmacy, The University of Queensland, Brisbane, QLD, Australia.
Mater Research Institute, The University of Queensland, Brisbane, QLD, Australia.
Lab Invest. 2020 Feb;100(2):224-233. doi: 10.1038/s41374-019-0280-3. Epub 2019 Jun 26.
The Ca signal is essential in both hypoxia- and epidermal growth factor (EGF)-mediated epithelial to mesenchymal transition (EMT) in MDA-MB-468 breast cancer cells. This finding suggests that Ca-permeable ion channels participate in the induction of expression of some mesenchymal markers such as vimentin. However, the ion channels involved in vimentin expression induction have not been fully characterized. This work sought to define how differential modulation of the calcium signal effects the induction of vimentin and the Ca influx pathways involved. We identified that the intracellular Ca chelator EGTA-AM, cytochalasin D (a modulator of cytoskeletal dynamics and cell morphology), and the sarco/endoplasmic reticulum ATPase inhibitor thapsigargin are all inducers of vimentin in MDA-MB-468 breast cancer cells. EGTA-AM- and thapsigargin-mediated induction of vimentin expression in MDA-MB-468 cells involves store-operated Ca entry, as evidenced by sensitivity to silencing of the molecular components of this pathway, STIM1 and ORAI1. In stark contrast, cytochalasin D-mediated vimentin induction was insensitive to silencing of ORAI1, despite sensitivity to silencing of its canonical activator the endoplasmic reticulum Ca sensor STIM1. Cytochalasin D-mediated vimentin induction was, however, sensitive to silencing of another reported STIM1 target, TRPC1. Subsequent studies identified that EGTA-AM-induced vimentin expression also partially involved a TRPC1-dependent pathway. These studies define a complex interplay between vimentin expression in this model and the specific Ca-permeable ion channels involved. The complexity in the engagement of different Ca influx pathways that regulate vimentin induction are opportunities but also potential challenges in targeting Ca signaling to block EMT in cancer cells. Our findings further highlight the need to identify potential indispensable ion channels that can regulate induction of specific mesenchymal markers via different stimuli.
钙信号在 MDA-MB-468 乳腺癌细胞的低氧和表皮生长因子（EGF）介导的上皮间质转化（EMT）中是必不可少的。这一发现表明，钙通透性离子通道参与了一些间充质标志物如波形蛋白表达的诱导。然而，参与波形蛋白表达诱导的离子通道尚未得到充分表征。本工作旨在确定钙信号的差异调节如何影响波形蛋白的诱导以及所涉及的钙内流途径。我们确定，细胞内钙螯合剂 EGTA-AM、细胞松弛素 D（细胞骨架动力学和细胞形态的调节剂）和肌浆/内质网 ATP 酶抑制剂 thapsigargin 都是 MDA-MB-468 乳腺癌细胞中波形蛋白的诱导剂。EGTA-AM 和 thapsigargin 介导的 MDA-MB-468 细胞中波形蛋白表达的诱导涉及储存操纵的钙内流，这一点可以通过对该途径的分子成分 STIM1 和 ORAI1 的沉默敏感性来证明。与此形成鲜明对比的是，细胞松弛素 D 介导的波形蛋白诱导对 ORAI1 的沉默不敏感，尽管对其经典激活剂内质网钙传感器 STIM1 的沉默敏感。然而，细胞松弛素 D 介导的波形蛋白诱导对另一种报道的 STIM1 靶标 TRPC1 的沉默敏感。随后的研究表明，EGTA-AM 诱导的波形蛋白表达也部分涉及 TRPC1 依赖性途径。这些研究定义了该模型中波形蛋白表达与所涉及的特定钙通透性离子通道之间的复杂相互作用。调节波形蛋白诱导的不同钙内流途径的参与的复杂性是靶向钙信号以阻断癌细胞 EMT 的机会，但也是潜在的挑战。我们的研究结果进一步强调了需要识别潜在的不可或缺的离子通道，这些通道可以通过不同的刺激来调节特定的间充质标志物的诱导。
