The Role Of Mycobacterium Tuberculosis Rv3904c(EsxE) In Stress And Immune Response

Tuberculosis(TB) is a global pandemic, resulted by M.tuberculosis, causing multiorgans damage, which is given priority to pulmonary tuberculosis. In recent years, drug resistance and co-infection with HIV make the treatments of TB more difficult, especially for MDR-TB, it is estimated that about 480000 new MDR-TB infections, but only 26% were diagnosis and treatment. Although effective prevention have made great achievements, TB is still the most serious public health threat, it is extremely urgent to better diagnosis treatment and development of new drugs.MTB can damage the antimicrobial pathway to macrophages, prevent from the lysosomes, mediated virulence by ESX secretion system, resulting in cell innate immune response. ESX-1 secretion system exists in a variety of actinomycetes, secreting esat-6 and cfp-10 protein, is an important MTB virulence factors. MTB genome can encod five potential gene cluster of type VII secretion system, which has similar sequences and different function.Type VII secretion system plays an important function in MTB host-pathogen interaction process, which could provide the better strategies to TB for prevention, diagnosis and treatment. While, esat- 6 is considered an important target antigen as early diagnosis strategy development.Esat-6(early secretary antigen target 6), a small secretary protein, is very important to virulence and protective immunity in MTB, the main advantage of antigen mediated by MTB early infection and can produce high level IFN-γ. Esat-6 belongs to the esat-6 family, which is crucial to early immune response in MTB infection. Esat-6 is a small protein, about 100 amino acid residues, is a member of esat-6 family. Esat-6 family has 23 proteins(Esx A-W), all of genes exist in pairs, except for Esx Q, some of highly similar, and has a similar sequence motif WXG(Trp-Xaa-Gly), which functionis unknown. This study is focused on Rv3904c(Esx E), one of the esat-6 / WXG100 family protein, which play roles in pressure and the immune response. We clone Rv3904 c gene from the MTB H37 Rvgenome, construct recombinant strain. Ms-Rv3904 c was cultured in 7H9 liquid medium, and induced by acetamide, and thendetect expression Rv3904 c protein by Western blotting; compared with Ms-Vec, we detectthe growth of Ms-Rv3904 cin vitro, biofilm differences; sentitive to antibiotics(Norfloxacin, soniazid, Vancomycin, Ofloxacin) and surface active material SDS;moreover, we check the suRvival of Ms-Rv3904 c and expression of inflammatory cytokines, further understand intracellular immune response of macrophages infected with Ms-Rv3904 c.Preliminary study found that overexpression of EsxE enhanced suRvival of recombinant Ms-Rv3904 c under various extracellular and intracellular stress. Antibiotics detection found that Ms-Rv3904 c had weak susceptibility to vancomycin and ofloxacin, while had obvious influence on norfloxacin and isoniazid, especially is highly sensitive to norfloxacin. After infected macrophage THP-1, we determined the expression of cytokines at transcription level, found that EsxE obviously increased production of IL-1β, IL-18 and IL-10, and other cytokines, IL-12 p40, TNF-α, IL-6, IL-18 and caspase-1 also had different up-regulation. All these results contribute to understand the function of esat-6 family proteins.