Deep Sequencing-based Expression Transcirptional Profiling Changes Of Mouse Macrophages During Brucella Melitensis16M Infection

Brucellosis,which is caused by Brucella, is a worldwide zoonotic infectiousdisease and it has been one hundred years. Brucella lack classic pathogenic factors,such as toxins, capsule, pilus, flagellum, cytolysin, extracellular enzyme and so on.But Brucella, which is intracellular, can survival in host cell and escape the defensemechanism of host. It indicated that Brucella has the special pathogenic mechanism.In current study, the transcriptional profile of RAW264.7cells infected with theBrucella melitensis strain16M was investigated using a technology based on deepsequencing for the first time. A suitable infection conditions was establishedaccording to the intracellular infection assay, macrophage cytotoxicity assay andindirect immunofluorescent assay when MOI is1000. Extracting RNA of infectioncell samples we found that there are approximately58%genes expressedup-regulation in which3576significantly changed genes4h post infection andapproximately45%genes expressed up-regulation in which3962significantlychanged genes24h post infection with the technology based on deep sequencing. Andwe verified the genes expressed significance difference with the real time RT-PCR.The passways significantly enriched postinfection include lysosome pathway, Fcgamma R-mediated phagocytosis and protein processing in the endoplasmicreticulum,which play an important role in phagocytosis, colonization and maturity.Apotosis pathway and p53signaling pathway which are related to apotosis. NOD-likereceptor signaling pathway, B-cell receptor and toll-like receptor signaling pathwayswhich are related to immunity.In the study, Acp5of lysosome was changed significally postinfection. Acp5whichis one of candicate enzyme transporting Man-6-P plays an important role intransporting specific mark of lysosome protein Man-6-P. If the hydrolysis enzyme inthelysosome want to work, the marker protein of Man-6-P must be posttranslationalmodification. Overexpression of Acp5can cause the phosphorylation efficiency oflysosome protein significantly increase and expression of Man-6-P glycoprotein ismore increased if the expressiμμμon of Acp5is inhibited. So we inhibited theexpression of Acp5utilized the technology of RNAi to evaluation the effect with the infection of Brucella.In current study, we established a database, which is based on significant differentexpression genes of mouse macrophage infected Brucella, which would be providethe basis for evaluating the role of host during the infection.