Genome-wide CRISPR-Cas9 viability screen reveals genes involved in TNF-α-induced apoptosis of human umbilical vein endothelial cells
Genome-wide CRISPR-Cas9 viability screen reveals genes involved in TNF-α-induced apoptosis of human umbilical vein endothelial cells
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全基因组 CRISPR-Cas9 活力筛选揭示了参与 TNF-α 诱导人脐静脉内皮细胞凋亡的基因
DOI：
10.1002/jcp.27595
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发表时间：
2019-06-01
影响因子：
5.6
通讯作者：
Zeng, Qiyi
中科院分区：
文献类型：
--
作者：
Cai, Meng;Li, Sitao;Zeng, Qiyi
肿瘤坏死因子α (tnf - α)是脓毒症中的关键细胞因子，通过促进炎症反应同时诱导血管内皮细胞凋亡来保护宿主免受病原体的侵害。不幸的是，靶向tnf - α信号通路某些成分以减少细胞凋亡的抑制剂未能转化为临床应用，部分原因是过度免疫抑制的不良影响。为了发现tnf - α信号通路中调节脓毒症发展过程中适度炎症和凋亡的潜在靶点，我们在人脐静脉内皮细胞（HUVECs）中进行了全基因组CRISPR/Cas9敲除筛选。肿瘤坏死因子受体超家族成员1A （TNFRSF1A）、b细胞淋巴瘤2 （BCL2）、BCL2相关死亡启动子（BAD）和NLR家族成员X1 （NLRX1）缺陷在候选调节因子列表中被确定为tnf - α细胞毒性的有效遗传抑制因子。crispr介导的NLRX1敲除可使细胞抵抗tnf - α的攻击，并且在tnf - α刺激后，NLRX1可被诱导与线粒体共定位。因此，我们的工作证明了用Cas9进行基因组级筛选的优势，并验证了NLRX1作为败血症期间tnf α诱导的血管内皮细胞凋亡的潜在调节剂。
Tumor necrosis factor alpha (TNF-alpha), a pivotal cytokine in sepsis, protects the host against pathogens by promoting an inflammatory response while simultaneously inducing apoptosis of the vascular endothelium. Unfortunately, inhibitors targeting certain components of the TNF-alpha signaling pathway to reduce cellular apoptosis have failed to translate into clinical applications, partly due to the adverse effects of excessive immunosuppression. In an attempt to discover potential targets in the TNF-alpha signaling pathway to modulate moderate inflammation and apoptosis during the development of sepsis, we performed a pooled genome-wide CRISPR/Cas9 knockout screen in human umbilical vein endothelial cells (HUVECs). Tumor necrosis factor receptor superfamily member 1A (TNFRSF1A), B-cell lymphoma 2 (BCL2), Bcl2-associated death promoter (BAD), and NLR family member X1 (NLRX1) deficiencies were identified as the effective genetic suppressors of TNF-alpha cytotoxicity on a list of candidate regulators. CRISPR-mediated NLRX1 knockout conferred cellular resistance to challenge with TNF-alpha, and NLRX1 could be induced to colocalize with mitochondria following TNF-alpha stimulation. Thus, our work demonstrates the advantage of genome-scale screening with Cas9 and validates NLRX1 as a potential modulator of TNF-alpha-induced vascular endothelial apoptosis during sepsis.