Besson Benoit, Sonthonnax Florian, Duchateau Magalie, Ben Khalifa Youcef, Larrous Florence, Eun Hyeju, Hourdel Véronique, Matondo Mariette, Chamot-Rooke Julia, Grailhe Regis, Bourhy Hervé
Unité Dynamique des Lyssavirus et Adaptation à l'Hôte, Paris, France.
Université Paris Diderot, Sorbonne Paris Cité, Cellule Pasteur, Paris, France.
PLoS Pathog. 2017 Oct 30;13(10):e1006697. doi: 10.1371/journal.ppat.1006697. eCollection 2017 Oct.
At the crossroad between the NF-κB and the MAPK pathways, the ternary complex composed of p105, ABIN2 and TPL2 is essential for the host cell response to pathogens. The matrix protein (M) of field isolates of rabies virus was previously shown to disturb the signaling induced by RelAp43, a NF-κB protein close to RelA/p65. Here, we investigated how the M protein disturbs the NF-κB pathway in a RelAp43-dependant manner and the potential involvement of the ternary complex in this mechanism. Using a tandem affinity purification coupled with mass spectrometry approach, we show that RelAp43 interacts with the p105-ABIN2-TPL2 complex and we observe a strong perturbation of this complex in presence of M protein. M protein interaction with RelAp43 is associated with a wide disturbance of NF-κB signaling, involving a modulation of IκBα-, IκBβ-, and IκBε-RelAp43 interaction and a favored interaction of RelAp43 with the non-canonical pathway (RelB and p100/p52). Monitoring the interactions between host and viral proteins using protein-fragment complementation assay and bioluminescent resonance energy transfer, we further show that RelAp43 is associated to the p105-ABIN2-TPL2 complex as RelAp43-p105 interaction stabilizes the formation of a complex with ABIN2 and TPL2. Interestingly, the M protein interacts not only with RelAp43 but also with TPL2 and ABIN2. Upon interaction with this complex, M protein promotes the release of ABIN2, which ultimately favors the production of RelAp43-p50 NF-κB dimers. The use of recombinant rabies viruses further indicates that this mechanism leads to the control of IFNβ, TNF and CXCL2 expression during the infection and a high pathogenicity profile in rabies virus infected mice. All together, our results demonstrate the important role of RelAp43 and M protein in the regulation of NF-κB signaling.
在核因子κB（NF-κB）和丝裂原活化蛋白激酶（MAPK）信号通路的交叉点上，由p105、ABIN2和TPL2组成的三元复合物对于宿主细胞对病原体的反应至关重要。狂犬病病毒野毒株的基质蛋白（M）先前已被证明会干扰由RelAp43介导的信号传导，RelAp43是一种与RelA/p65相近的NF-κB蛋白。在此，我们研究了M蛋白如何以依赖RelAp43的方式干扰NF-κB信号通路，以及三元复合物在该机制中的潜在作用。通过串联亲和纯化结合质谱分析方法，我们发现RelAp43与p105-ABIN2-TPL2复合物相互作用，并且在存在M蛋白的情况下观察到该复合物受到强烈干扰。M蛋白与RelAp43的相互作用与NF-κB信号的广泛紊乱相关，涉及IκBα、IκBβ和IκBε与RelAp43相互作用的调节，以及RelAp43与非经典途径（RelB和p100/p52）的优先相互作用。使用蛋白质片段互补分析和生物发光共振能量转移监测宿主蛋白与病毒蛋白之间的相互作用，我们进一步表明RelAp43与p105-ABIN2-TPL2复合物相关，因为RelAp43-p105相互作用稳定了与ABIN2和TPL2形成的复合物。有趣的是，M蛋白不仅与RelAp43相互作用，还与TPL2和ABIN2相互作用。与该复合物相互作用后，M蛋白促进ABIN2的释放，最终有利于RelAp43-p50 NF-κB二聚体的产生。使用重组狂犬病病毒进一步表明，该机制导致感染期间干扰素β（IFNβ）、肿瘤坏死因子（TNF）和CXC趋化因子配体2（CXCL2）表达的调控，并使狂犬病病毒感染的小鼠呈现高致病性特征。总之，我们的结果证明了RelAp43和M蛋白在NF-κB信号调节中的重要作用。
