Regulation of ezrin tension by S-nitrosylation mediates non-small cell lung cancer invasion and metastasis
Regulation of ezrin tension by S-nitrosylation mediates non-small cell lung cancer invasion and metastasis
复制标题
S-亚硝基化调节ezrin张力介导非小细胞肺癌侵袭和转移
DOI：
10.7150/thno.32479
复制
发表时间：
2019-01-01
期刊：
影响因子：
12.4
通讯作者：
Guo, Jun
中科院分区：
文献类型：
--
作者：
Zhang, Xiaolong;Li, Guangming;Guo, Jun
癌症的侵袭和转移依赖于化学和机械活动的准确和快速的调节。膜细胞骨架交联蛋白ezrin的S-亚硝基化（SNO）可能以张力依赖性方式调节恶性过程。方法：采用生物素开关法检测非小细胞肺癌（NSCLC）组织和A549细胞株中亚硝基埃兹蛋白的表达水平。用ezrin的几个半胱氨酸突变质粒来鉴定SNO的活性位点。新设计的ezrin或突变ezrin张力探针的基础上Förster共振能量转移（FRET）理论，应用视觉观察实时张力变化。通过细胞骨架解聚和运动分子抑制实验，揭示了SNO作用后ezrin力学性能的变化。使用Transwell测定和异种移植小鼠模型来评估不同组中A549细胞的侵袭性。荧光染色也用于检查细胞的位置和结构。结果如下：高水平的诱导型一氧化氮合酶（iNOS）可诱导ezrin-SNO，从而促进NSCLC细胞在体内外的恶性行为。Cys 117被确定为ezrin-SNO的唯一活性位点。同时，在iNOS诱导的SNO后观察到ezrin张力水平增加。ezrin张力增强与NSCLC的侵袭性呈正相关。此外，微丝（MF）的力量，而不是微管（MT）的力量在调节ezrin张力，特别是在ezrin亚硝基化后起主导作用。结论：本研究揭示了一个SNO相关机制的机械张力ezrin。Ezrin-SNO通过促进从细胞骨架到细胞膜的机械转导来促进NSCLC细胞的侵袭和转移。这些研究暗示了靶向ezrin在抑制NSCLC侵袭和转移方面的治疗潜力。
Cancer invasion and metastasis depend on accurate and rapid modulation of both chemical and mechanical activities. The S-nitrosylation (SNO) of membrane cytoskeletal cross-linker protein ezrin may regulate the malignant process in a tension-dependent manner. Methods: The level of nitrosylated ezrin in non-small cell lung cancer (NSCLC) tissues and A549 cell line were evaluated by biotin-switch assay. A few cysteine mutated plasmids of ezrin were used to identify active site for SNO. Newly designed ezrin or mutated-ezrin tension probes based on Förster resonance energy transfer (FRET) theory were applied to visually observe real-time tension changes. Cytoskeleton depolymerizing and motor molecular inhibiting experiments were performed to reveal the alternation of the mechanical property of ezrin after SNO. Transwell assays and xenograft mouse model were used to assess aggressiveness of A549 cells in different groups. Fluorescent staining was also applied to examine cellular location and structures. Results: High inducible nitric oxide synthase (iNOS) levels were observed to induce ezrin-SNO, and then promote malignant behaviors of NSCLC cells both in vitro and in vivo. Cys117 was identified as the only active site for ezrin-SNO. Meanwhile, an increased level of ezrin tension was observed after iNOS-induced SNO. Enhanced ezrin tension was positively correlated with aggressiveness of NSCLC. Moreover, Microfilament (MF) forces instead of microtubule (MT) forces played dominant roles in modulating ezrin tension, especially after ezrin nitrosylation. Conclusion: This study revealed a SNO-associated mechanism underlying the mechanical tension of ezrin. Ezrin-SNO promotes NSCLC cells invasion and metastasis through facilitating mechanical transduction from the cytoskeleton to the membrane. These studies implicate the therapeutic potential by targeting ezrin in the inhibition NSCLC invasion and metastasis.