title: [Preparation of an autophagy-targeted antituberculosis DNA vaccine encoding LpqH fused with LC3].
abstract: OBJECTIVE
  To construct an autophagy-targeted vaccine harboring the genes encoding lipoprotein antigen precursor LpqH from Mycobacterium tuberculosis and microtubule-associated protein light chain-3(LC3), and to investigate its efficacy of inducing and targeting autophagy.
  
  
  METHODS
  The expressions of LC-3 and LC3-LpqH in RAW264.7 cells were detected by Western blotting after transfected with pCMV-LpqH and pCMV-LC3-LpqH plasmids respectively. The pCMV-LC3-LpqH or pCMV-LpqH plasmids were transfected into GFP-LC3-RAW264.7 cells to analyze the localization of LC3-LpqH by immunofluorescence staining.
  
  
  RESULTS
  After transfected with pCMV-LpqH DNA, RAW264.7 cells showed a significant increase of LC-3 amount. The LC3-LpqH fusion protein was also detected in RAW264.7 cells after pCMV-LC3-LpqH transfection and in a dose-dependent manner. Interestingly, LpqH was found to be transported to autophagosomes through the fusion protein, which was demonstrated by the co-localization of GFP-LC3 and LC3-LpqH on autophagosomes.
  
  
  CONCLUSIONS
  The recombinant plasmid encoding pCMV-LC3-LpqH could enhance the autophagy in vitro, and facilitate the localization of LpqH on autophagosomes. Our study provides a new practical strategy for the development of improved vaccines against Mycobacterium tuberculosis.
authors.affiliation: Institute of Infection and Immunology, Department of Immunology, Medical School, Anhui University of Science and Technology, Huainan 232001, China.
authors.forename: Dong
authors.initials: D
authors.lastname: Hu
authors.affiliation: Institute of Infection and Immunology, Department of Immunology, Medical School, Anhui University of Science and Technology, Huainan 232001, China.
authors.forename: Xiaokang
authors.initials: X
authors.lastname: Yang
authors.affiliation: Institute of Infection and Immunology, Department of Immunology, Medical School, Anhui University of Science and Technology, Huainan 232001, China.
authors.forename: Jing
authors.initials: J
authors.lastname: Wu
authors.affiliation: Deparment of Pathology, General Hospital of Oriental Group, Huainan 232001, China.
authors.forename: Ganxun
authors.initials: G
authors.lastname: Wang
authors.affiliation: Institute of Infection and Immunology, Department of Immunology, Medical School, Anhui University of Science and Technology, Huainan 232001, China.
authors.forename: Wenyang
authors.initials: W
authors.lastname: Wang
authors.affiliation: Deparment of Pathology, General Hospital of Oriental Group, Huainan 232001, China.
authors.forename: Zhenyu
authors.initials: Z
authors.lastname: Qi
authors.affiliation: Institute of Infection and Immunology, Department of Immunology, Medical School, Anhui University of Science and Technology, Huainan 232001, China.
authors.forename: Gong
authors.initials: G
authors.lastname: Chen
authors.affiliation: Institute of Infection and Immunology, Department of Immunology, Medical School, Anhui University of Science and Technology, Huainan 232001, China.
authors.forename: Rongbo
authors.initials: R
authors.lastname: Zhang
mesh_terms: D000818:Animals; D000942:Antigens, Bacterial / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D000995:Antitubercular Agents / Q000008:administration & dosage / Q000276:immunology; D001343:Autophagy / Q000276:immunology*; D015153:Blotting, Western; D002460:Cell Line; D005455:Fluorescent Antibody Technique; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D008074:Lipoproteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D008264:Macrophages / Q000166:cytology / Q000276:immunology / Q000378:metabolism; D051379:Mice; D008807:Mice, Inbred BALB C; D008869:Microtubule-Associated Proteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D009169:Mycobacterium tuberculosis / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D010588:Phagosomes / Q000276:immunology / Q000378:metabolism; D010957:Plasmids / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D014162:Transfection; D014376:Tuberculosis / Q000276:immunology / Q000382:microbiology / Q000517:prevention & control; D032581:Tuberculosis Vaccines / Q000008:administration & dosage / Q000235:genetics / Q000276:immunology*; D019444:Vaccines, DNA / Q000008:administration & dosage / Q000235:genetics / Q000276:immunology
chemical_list: D000942:Antigens, Bacterial; D000995:Antitubercular Agents; D008074:Lipoproteins; C480860:Map1lc3b protein, mouse; D008869:Microtubule-Associated Proteins; D011993:Recombinant Fusion Proteins; D032581:Tuberculosis Vaccines; D019444:Vaccines, DNA; D049452:Green Fluorescent Proteins