Laboratory Animal Medicine, College of Veterinary Medicine, Chonnam National University, Gwangju 61186, Republic of Korea.
Department of Microbiology, Institute for Immunology and Immunological Diseases, Brain Korea 21 PLUS Project for Medical Science, Yonsei University College of Medicine, Seoul 03722, Republic of Korea.
Cytokine. 2018 Apr;104:104-109. doi: 10.1016/j.cyto.2017.10.006. Epub 2017 Oct 16.
Mycobacterium tuberculosis is a highly virulent bacterium that causes tuberculosis. It infects about one third of the world's population. Type I interferons (IFNs) play a detrimental role in host defense against M. tuberculosis infection. Proteins secreted by M. tuberculosis through ESX-1 secretion system contribute to type I IFNs production. However, the precise mechanism by which 6-kDa early secretory antigen target (ESAT6), one of ESX-1-mediated secretory proteins, induces type I IFNs production in host cells is currently unclear. Therefore, the objective of the present study was to determine the underlying molecular mechanism regulating ESAT6-mediated gene expression of IFN-β in macrophages. Recombinant ESAT6 produced from E. coli expression system induced IFN-β gene expression in various types of macrophages such as mouse bone marrow-derived macrophages (BMDMs), peritoneal macrophages, and MH-S cells (murine alveolar macrophage cell line). Deficiency of TLR4 and TRIF absolutely abrogated ESAT6-induced IFN-β gene expression. TLR2 and MyD88 were partially involved in IFN-β gene expression in response to low dose of ESAT6. Another recombinant ESAT6 produced from baculovirus system also upregulated IFN-β gene expression via TLR4-dependent pathway. Polymyxin B (PMB) treatment impaired LPS-induced IFN-β expression. However, IFN-β expression induced by ESAT6 was not influenced by PMB. This suggests that ESAT6-mediated IFN-β expression is not due to LPS contamination. Treatment with ESAT6 resulted in activation of TBK1 and IRF3 in macrophages. Such activation was abolished in TLR4- and TRIF-deficient cells. Moreover, inhibition of IRF3 and TBK1 suppressed IFN-β gene expression in response to ESAT6. Our results suggest that ESAT6 might contribute to virulence of M. tuberculosis by regulating type I IFNs production through TLR4-TRIF signaling pathway.
结核分枝杆菌是一种高度毒力的细菌，可引起结核病。它感染了世界上约三分之一的人口。I 型干扰素（IFNs）在宿主防御结核分枝杆菌感染中起有害作用。结核分枝杆菌通过 ESX-1 分泌系统分泌的蛋白质有助于 I 型 IFNs 的产生。然而，ESX-1 介导的分泌蛋白之一 6-kDa 早期分泌抗原靶标（ESAT6）如何诱导宿主细胞产生 I 型 IFNs 的精确机制尚不清楚。因此，本研究旨在确定调节 ESAT6 介导的巨噬细胞 IFN-β基因表达的潜在分子机制。来自大肠杆菌表达系统的重组 ESAT6 诱导各种类型的巨噬细胞（如小鼠骨髓来源的巨噬细胞（BMDMs）、腹腔巨噬细胞和 MH-S 细胞（鼠肺泡巨噬细胞系））中 IFN-β基因表达。TLR4 和 TRIF 的缺失绝对阻断了 ESAT6 诱导的 IFN-β基因表达。TLR2 和 MyD88 部分参与了低剂量 ESAT6 诱导的 IFN-β基因表达。来自杆状病毒系统的另一种重组 ESAT6 也通过 TLR4 依赖性途径上调 IFN-β基因表达。多粘菌素 B（PMB）处理可损害 LPS 诱导的 IFN-β表达。然而，ESAT6 诱导的 IFN-β表达不受 PMB 影响。这表明 ESAT6 介导的 IFN-β表达不是由于 LPS 污染。用 ESAT6 处理导致巨噬细胞中 TBK1 和 IRF3 的激活。在 TLR4 和 TRIF 缺陷细胞中，这种激活被消除。此外，IRF3 和 TBK1 的抑制抑制了 ESAT6 应答中的 IFN-β基因表达。我们的结果表明，ESAT6 可能通过 TLR4-TRIF 信号通路调节 I 型 IFNs 的产生来促进结核分枝杆菌的毒力。
