Pellino-1 Positively Regulates Toll-like Receptor (TLR) 2 and TLR4 Signaling and Is Suppressed upon Induction of Endotoxin Tolerance
Pellino-1 Positively Regulates Toll-like Receptor (TLR) 2 and TLR4 Signaling and Is Suppressed upon Induction of Endotoxin Tolerance
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DOI：
10.1074/jbc.m115.640128
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发表时间：
2015-07-31
影响因子：
4.8
通讯作者：
Medvedev, Andrei E.
中科院分区：
文献类型：
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作者：
Murphy, Michael;Xiong, Yanbao;Medvedev, Andrei E.
内毒素耐受性通过减弱促炎细胞因子的诱导而重新编程Toll样受体（TLR）4介导的巨噬细胞应答，同时保留抗炎和抗菌介质的表达。我们先前证明了TLR 4缺陷诱导的IL-1受体相关激酶（IRAK）4、IRAK 1和TANK结合激酶（TBK）1的激活是内毒素耐受的关键标志，但机制仍不清楚。在这项研究中，我们研究了E3泛素连接酶Pellino-1在内毒素耐受性和TLR信号转导中的作用。LPS刺激增加Pellino-1的mRNA和蛋白质表达的巨噬细胞从小鼠注射生理盐水和介质预处理的人单核细胞，THP-1，和MonoMac-6细胞，而内毒素耐受废除LPS诱导的Pellino-1。Pellino-1在293/TLR 2和293/TLR 4/MD 2细胞中的过表达增强了TLR 2和TLR 4诱导的核因子κ B（NF-κ B）和IL-8 mRNA的表达，而Pellino-1敲低降低了这些反应。THP-1细胞中的Pellino-1消融损害了髓样分化初级应答蛋白（MyD 88）的诱导，以及含有Toll-IL-1 R结构域的衔接子诱导IFN-β（TRIF）依赖性细胞因子基因对TLR 4和TLR 2激动剂以及热灭活大肠杆菌和金黄色葡萄球菌的应答，而仅微弱地影响热灭活细菌的吞噬作用。共表达的Pellino-1增强了由转染的MyD 88、TRIF、IRAK 1、TBK 1、TGF-β激活的激酶（TAK）1和TNFR相关因子6驱动的NF-κ B B激活，而不影响p65诱导的应答。从机制上讲，Pellino-1增加了LPS驱动的K63连接的IRAK 1、TBK 1、TAK 1的多聚泛素化，以及TBK 1和IFN调节因子3的磷酸化。这些结果揭示了一种新的机制，通过这种机制，内毒素耐受性通过抑制Pellino-1来重新编程TLR 4信号传导，Pellino-1是MyD 88和TRIF依赖性信号传导的正调控因子，促进IRAK 1，TBK 1和TAK 1的K63连接的多聚泛素化。
Endotoxin tolerance reprograms Toll-like receptor (TLR) 4-mediated macrophage responses by attenuating induction of proinflammatory cytokines while retaining expression of anti-inflammatory and antimicrobial mediators. We previously demonstrated deficient TLR4-induced activation of IL-1 receptor-associated kinase (IRAK) 4, IRAK1, and TANK-binding kinase (TBK) 1 as critical hallmarks of endotoxin tolerance, but mechanisms remain unclear. In this study, we examined the role of the E3 ubiquitin ligase Pellino-1 in endotoxin tolerance and TLR signaling. LPS stimulation increased Pellino-1 mRNA and protein expression in macrophages from mice injected with saline and in medium-pretreated human monocytes, THP-1, and MonoMac-6 cells, whereas endotoxin tolerization abrogated LPS inducibility of Pellino-1. Overexpression of Pellino-1 in 293/TLR2 and 293/TLR4/MD2 cells enhanced TLR2- and TLR4-induced nuclear factor kappa B (NF-kappa B) and expression of IL-8 mRNA, whereas Pellino-1 knockdown reduced these responses. Pellino-1 ablation in THP-1 cells impaired induction of myeloid differentiation primary response protein (MyD88), and Toll-IL-1R domain-containing adapter inducing IFN-beta (TRIF)-dependent cytokine genes in response to TLR4 and TLR2 agonists and heat-killed Escherichia coli and Staphylococcus aureus, whereas only weakly affecting phagocytosis of heat-killed bacteria. Co-expressed Pellino-1 potentiated NF-kappa B activation driven by transfected MyD88, TRIF, IRAK1, TBK1, TGF-beta-activated kinase (TAK) 1, and TNFR-associated factor 6, whereas not affecting p65-induced responses. Mechanistically, Pellino-1 increased LPS-driven K63-linked polyubiquitination of IRAK1, TBK1, TAK1, and phosphorylation of TBK1 and IFN regulatory factor 3. These results reveal a novel mechanism by which endotoxin tolerance re-programs TLR4 signaling via suppression of Pellino-1, a positive regulator of MyD88- and TRIF-dependent signaling that promotes K63-linked polyubiquitination of IRAK1, TBK1, and TAK1.