The Function Of Lipoprotein Rv0847 Gene Of Mycobacterium Tuberculosis

Tuberculosis(Tuberculosis,TB)is a zoonotic disease caused by Mycobacterium tuberculosis(M.tb)complex infection.It seriously threatens global public health security and caused huge economic losses to the breeding industry.M.tb is an intracellular bacteria that could retain long term persistence in macrophages and dendritic cells.However,the mechanism by which M.tb escapes host immune clearance is still unclear,which restricts the development of anti-tuberculosis vaccines and drug development.Lipoproteins of M.tb have strong immunogenicity and can directly interact with host,and could be potential target in the diagnosis,treatment and vaccine development to tuberculosis.Previous study from our lab conducted the screening of autophagy-related M.tb genes,and found that Rv0847 can interfere with host cell autophagy.However,the molecular mechanism of this process still remains elusive.This research has achieved the following findings:1.Rv0847 of M.tb has no effect on the growth rate of M.tb in vitro but can significantly promote the intracellular survival of M.tbIn this study,knockout strains(H37Ra-ΔRv0847),complementary strains(H37Ra-ΔRv0847::Rv0847)and overexpression strains(H37Ra-Rv0847-OE)of M.tb were constructed respectively.Then the in vitro growth curves of different strains were measured,and the results showed that Rv0847 significantly promoted the intracellular survival of M.tb without affecting the growth rate in vitro.2.Knockout of Rv0847 attenuates the pathogenicity of Mtb.In order to reveal the effect of Rv0847 expression on the pathogenicity of M.tb.Mouse tail vein injection infection model to assess the pathogenicity of H37Ra-WT,H37Ra-ΔRv0847,H37Ra-ΔRv0847::Rv0847.The result showed that the pathogenicity of H37Ra-ΔRv0847 is weakened comparing to H37Ra-WT.3.Rv0847 of M.tb inhibits host cell autophagyIn this study,GML-RAW264.7-Rv0847 cell line and RAW264.7-Rv0847 cell line were constructed.GML-RAW264.7-Rv0847 and GML-RAW264.7-vector cell lines were treated with rapamycin to induce autophagy,and the autophagy was observed under a laser confocal microscope after slice preparation.The experimental results showed that Rv0847 can inhibit host cell autophagy.4.Potential mechanism of Rv0847 of M.tb inhibits host cell autophagyIn this study,the yeast two-hybrid method was used to find out the protein interacted with Rv0847.Bait library was constructed,and were then hybridized with prey library.The results showed that Rv0847 can interact with VAMP3 and VAMP7.In summary: this study found that Rv0847 significantly promotes intracellular survival of M.tb by inhibiting host cell autophagy;and knocking out Rv0847 attenuates the pathogenicity of M.tb.The mechanism by which Rv0847 inhibits host cell autophagy may be that Rv0847 interacts with VAMP3 and VAMP7.On the basis of this study,the molecular mechanism of Rv0847 inhibiting host autophagy could be further determined,which is expected to obtain new targets for the design of anti-tuberculosis drugs and promoted the prevention and control of tuberculosis.