Sahu Sanjaya Kumar, Kumar Manish, Chakraborty Sohini, Banerjee Srijon Kaushik, Kumar Ranjeet, Gupta Pushpa, Jana Kuladip, Gupta Umesh D, Ghosh Zhumur, Kundu Manikuntala, Basu Joyoti
Department of Chemistry, Bose Institute, Kolkata, India.
Bioinformatics Centre, Bose Institute, Kolkata, India.
PLoS Pathog. 2017 May 30;13(5):e1006410. doi: 10.1371/journal.ppat.1006410. eCollection 2017 May.
For efficient clearance of Mycobacterium tuberculosis (Mtb), macrophages tilt towards M1 polarization leading to the activation of transcription factors associated with the production of antibacterial effector molecules such as nitric oxide (NO) and proinflammatory cytokines such as interleukin 1 β (IL-1β) and tumor necrosis factor α (TNF-α). At the same time, resolution of inflammation is associated with M2 polarization with increased production of arginase and cytokines such as IL-10. The transcriptional and post-transcriptional mechanisms that govern the balance between M1 and M2 polarization, and bacteria-containing processes such as autophagy and trafficking of Mtb to lysosomes, are incompletely understood. Here we report for the first time, that the transcription factor KLF4 is targeted by microRNA-26a (miR-26a). During Mtb infection, downregulation of miR-26a (observed both ex vivo and in vivo) facilitates upregulation of KLF4 which in turn favors increased arginase and decreased iNOS activity. We further demonstrate that KLF4 prevents trafficking of Mtb to lysosomes. The CREB-C/EBPβ signaling axis also favors M2 polarization. Downregulation of miR-26a and upregulation of C/ebpbeta were observed both in infected macrophages as well as in infected mice. Knockdown of C/ebpbeta repressed the expression of selected M2 markers such as Il10 and Irf4 in infected macrophages. The importance of these pathways is substantiated by observations that expression of miR-26a mimic or knockdown of Klf4 or Creb or C/ebpbeta, attenuated the survival of Mtb in macrophages. Taken together, our results attribute crucial roles for the miR-26a/KLF4 and CREB-C/EBPβsignaling pathways in regulating the survival of Mtb in macrophages. These studies expand our understanding of how Mtb hijacks host signaling pathways to survive in macrophages, and open up new exploratory avenues for host-targeted interventions.
为了有效清除结核分枝杆菌（Mtb），巨噬细胞倾向于M1极化，从而导致与抗菌效应分子（如一氧化氮（NO））以及促炎细胞因子（如白细胞介素1β（IL-1β）和肿瘤坏死因子α（TNF-α））产生相关的转录因子激活。同时，炎症的消退与M2极化相关，精氨酸酶和细胞因子（如IL-10）的产生增加。调控M1和M2极化之间平衡以及含细菌过程（如自噬和Mtb向溶酶体的转运）的转录和转录后机制尚未完全明确。在此，我们首次报道转录因子KLF4是微小RNA-26a（miR-26a）的作用靶点。在Mtb感染期间，miR-26a的下调（在体外和体内均观察到）促进了KLF4的上调，这反过来又有利于精氨酸酶增加和诱导型一氧化氮合酶（iNOS）活性降低。我们进一步证明KLF4可阻止Mtb向溶酶体的转运。CREB-C/EBPβ信号轴也有利于M2极化。在感染的巨噬细胞以及感染的小鼠中均观察到miR-26a的下调和C/ebpbeta的上调。敲低C/ebpbeta可抑制感染巨噬细胞中选定的M2标志物（如Il10和Irf4）的表达。miR-26a模拟物的表达或Klf4、Creb或C/ebpbeta的敲低可减弱Mtb在巨噬细胞中的存活，这些观察结果证实了这些途径的重要性。综上所述，我们的结果表明miR-26a/KLF4和CREB-C/EBPβ信号通路在调节Mtb在巨噬细胞中的存活中起关键作用。这些研究扩展了我们对Mtb如何劫持宿主信号通路以在巨噬细胞中存活的理解，并为以宿主为靶点的干预开辟了新的探索途径。
