Reynoso Eduardo, Liu Hua, Li Lin, Yuan Anthony L, Chen She, Wang Zhigao
From the Department of Molecular Biology, University of Texas Southwestern Medical Center, Dallas, Texas 75390.
School of Pharmacy, Jiangxi University of Traditional Chinese Medicine, Nanchang, Jiangxi 330006, China, and.
J Biol Chem. 2017 Oct 20;292(42):17514-17524. doi: 10.1074/jbc.M117.799353. Epub 2017 Sep 6.
Necroptosis is an immunogenic cell death program that is associated with a host of human diseases, including inflammation, infections, and cancer. Receptor-interacting protein kinase 3 (RIPK3) and its substrate mixed lineage kinase domain-like protein (MLKL) are required for necroptosis activation. Specifically, RIPK3-dependent MLKL phosphorylation promotes the assembly of disulfide bond-dependent MLKL polymers that drive the execution of necroptosis. However, how MLKL disulfide bond formation is regulated is not clear. In this study we discovered that the MLKL-modifying compound necrosulfonamide cross-links cysteine 86 of human MLKL to cysteine 32 of the thiol oxidoreductase thioredoxin-1 (Trx1). Recombinant Trx1 preferentially binds to monomeric MLKL and blocks MLKL disulfide bond formation and polymerization Inhibition of MLKL polymer formation requires the reducing activity of Trx1. Importantly, shRNA-mediated knockdown of Trx1 promotes MLKL polymerization and sensitizes cells to necroptosis. Furthermore, pharmacological inhibition of Trx1 with compound PX-12 induces necroptosis in multiple cancer cell lines. Altogether, these findings demonstrate that Trx1 is a critical regulator of necroptosis that suppresses cell death by maintaining MLKL in a reduced inactive state. Our results further suggest new directions for targeted cancer therapy in which thioredoxin inhibitors like PX-12 could potentially be used to specifically target cancers expressing high levels of MLKL or MLKL short isoforms.
坏死性凋亡是一种免疫原性细胞死亡程序，与许多人类疾病相关，包括炎症、感染和癌症。受体相互作用蛋白激酶3（RIPK3）及其底物混合谱系激酶结构域样蛋白（MLKL）是坏死性凋亡激活所必需的。具体而言，RIPK3依赖的MLKL磷酸化促进了二硫键依赖的MLKL聚合物的组装，从而驱动坏死性凋亡的执行。然而，MLKL二硫键形成是如何被调控的尚不清楚。在本研究中，我们发现MLKL修饰化合物坏死磺酰胺将人MLKL的半胱氨酸86与硫醇氧化还原酶硫氧还蛋白-1（Trx1）的半胱氨酸32交联。重组Trx1优先结合单体MLKL，并阻断MLKL二硫键的形成和聚合。抑制MLKL聚合物形成需要Trx1的还原活性。重要的是，shRNA介导的Trx1敲低促进MLKL聚合，并使细胞对坏死性凋亡敏感。此外，用化合物PX-12对Trx1进行药理学抑制可在多种癌细胞系中诱导坏死性凋亡。总之，这些发现表明Trx1是坏死性凋亡的关键调节因子，通过将MLKL维持在还原的非活性状态来抑制细胞死亡。我们的结果进一步为靶向癌症治疗提出了新方向，其中像PX-12这样的硫氧还蛋白抑制剂可能潜在地用于特异性靶向表达高水平MLKL或MLKL短异构体的癌症。
