RalA exerts an inhibitory effect on IL-1β/IL-18 secretion by blocking NLRP3 inflammasome activation in levornidazole-treated human THP-1 macrophages
RalA exerts an inhibitory effect on IL-1β/IL-18 secretion by blocking NLRP3 inflammasome activation in levornidazole-treated human THP-1 macrophages
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RalA 通过阻断左奥硝唑处理的人 THP-1 巨噬细胞中 NLRP3 炎性体的激活，对 IL-1 beta/IL-18 分泌产生抑制作用
DOI：
10.1016/j.intimp.2020.106898
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发表时间：
2020
影响因子：
5.6
通讯作者：
Ruiqin Yao
中科院分区：
文献类型：
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作者：
Xingqi Wang;Lingshan Gou;Yuzhi Gao;Yuqing Huang;Rui Kuai;Yu Li;Yujing Wang;Yanhong Chen;Jun Li;Chao Cheng;Zhaojun Feng;Xuefeng Wu;Ruiqin Yao
NLRP 3炎性体是炎症反应的重要介质，其调控是一个活跃的研究领域。RalA是一种Ras样的GTdR，在细胞生物学中起着关键作用。到目前为止，关于RalA调节炎症反应的研究很少。生物信息学分析预测RalA可能参与NLRP 3炎性体的调控网络，这在THP-1巨噬细胞中得到证实。通过对化合物的虚拟筛选，发现从我们的虚拟小分子化合物库中筛选出的左旋咪唑具有与RalA结合的潜力。值得注意的是，RalA/左奥硝唑的相互作用通过表面等离子体共振-Biacore T200、LC/MS分析和蛋白质印迹分析来验证。分子动力学模拟结果表明，RalA的构象变化可能受到左奥硝唑的调控。此外，RalA依赖性地抑制ATP + LPS刺激的THP-1源性巨噬细胞分泌IL-1β/IL-18，提示RalA可能对NLRP 3炎性小体活化具有抑制作用。免疫共沉淀和RalA耗竭实验结果表明，左奥硝唑可诱导RalA阻断NLRP 3/ASC/pro-caspase-1复合物的组装，从而降低裂解型caspase-1和IL-1β/IL-18的分泌水平。我们的研究表明RalA具有抗炎作用，并确定了其靶向化合物。总之，本研究阐明了RalA/左奥硝唑抑制NLRP 3炎性小体活化和IL-1β/IL-18分泌的新药理学机制。
The NLRP3 inflammasome is an important mediator of inflammatory responses and its regulation is an active area of research. RalA is a Ras-like GTPase, which play pivotal roles in the biology of cells. So far, there have been very few studies on RalA regulating inflammatory responses. Bioinformatics analysis predicted that RalA might participate in the regulatory network of NLRP3 inflammasome, which has been confirmed in THP-1 macrophages. After virtual screening of compounds, it was found that levonidazole selected from our virtual small molecule compound library has the potential to bind to RalA. Of note, the interaction of RalA/levornidazole was verified by Surface Plasmon Resonance-Biacore T200, LC/MS analysis and Western blotting analysis. Molecular dynamics simulations revealed that the conformational changes of RalA might be regulated by levornidazole. Additionally, IL-1β/IL-18 secretion from ATP + LPS stimulated THP-1-derived macrophages was RalA-dependently suppressed by levornidazole, suggesting that RalA might have an inhibitory effect on NLRP3 inflammasome activation. The results of co-immunoprecipitation and RalA depletion experiments showed that levornidazole could induce RalA to block the assembly of NLRP3/ASC/pro-caspase-1 complex, thereby reducing the levels of cleaved-caspase-1 and IL-1β/IL-18 secretion. Our study has suggested an anti-inflammatory function of RalA and identified its targeting chemical compound. Overall, this study clarifies a novel pharmacological mechanism by which RalA/levornidazole inhibits NLRP3 inflammasome activation and IL-1β/IL-18 secretion.