Role Of DNA Sensor IFI204 And AIM2 In IFN-? Secrection And Autophagy Induced By Mycobacterium Bovis

Tuberculosis(TB)is one of the largest infectious disease caused human death in the world.TB in human is mainly caused by Mycobacterium tuberculosis(Mtb),but 4.2%to 10.6%of people are caused by Mycobacterium bovis(M.bovis)which is a big threat to human health and cattle industry.Similar to Mtb,alveolar macrophages are considered to be the main host cells in early stage of M.bovis infection,which making the first defense line of the host.Intracellular double-stranded DNA recognition receptors are activated by mycobacteria when they are phagocytosed by macrophages,which caused type ?interferon expression and autophagy activation.IFN-? is a cytokine produced by the host which is proved to promote Mtb infection,whereas host cells can reduce the survival of intracellular mycobacteria by selective autophagy.Whether intracellular double-stranded DNA receptors are involved during M.bovis infection and how to regulate IFN-? expression and autophagy is not yet clear.In this study,the effects of DNA recognition receptors IFI204 and AIM2 on the induction of IFN-?secretion and autophagy in macrophages infected by M.bovis were studied,and the specific mechanisms of these two recognition receptors in immune response during M.bovis infection were clarified,which enhance our understanding of the interaction between mycobacteria and host cells.1.Mouse macrophage J774A.1 and mouse bone marrow-derived macrophages(BMDMs)were infected with M.bovis in MOI of 10,and the total proteins were extracted to detect phosphorylation of IRF3,and the translocation of IRF3 from cytoplasm to nucleus was detected in cytoplasmic/nuclear proteins which confirmed that M.bovis can induce phosphorylation of IRF3 and IFN-? expression.BMDMs were infected with M.bovis in MOI of 10,and the autophagy-related proteins LC3 and p62 were detected at different infection times and the expression of autophagy-related protein LC3 and p62 was detected in different MOI at the same time.The results showed that Mycobacterium can enhance autophagy in macrophages.The location of the bacteria in macropgaes was observed by transmission electron microscopy.The results showed that the M.bovis in macrophages could release into the cytoplasm.2.In order to study the effect of IFI204 and AIM2 on the expression of IFN-? and autophagy during M.bovis infection,we used siRNA to silence the expression of IFI204 and AIM2,respectively,and then detect the mRNA expression of IFN-?,IL-1?,TNF-? and IL-10 and protein expreesion of LC3,TBK-1 and IRF3.The results showed that IFN-? secretion and autophagy induced by M.bovis were dependent on IFI204 and its downstream STING-related signaling pathway,and in this process,IFI204 was required to be acetylated and transferred to the cytoplasm.The IFN-? secretion and autophagy were inhibited by AIM2 through competitive inhibition of STING downstream signaling pathways.3.We investigated the effects of IFI204 and AIM2 on the survival of M.bovis in infected macrophages.We used siRNA to silence the expression of IFI204 and AIM2 in mouse macrophage J774A.1 and mouse BMDMs,respectively.The results showed that IFI204 did not affect the survival of intracellular mycobacteria,but AIM2 inhibited the survival of M.bovis in macrophages.