title: Mechanism of Asp24 upregulation in Brucella abortus rough mutant with a disrupted O-antigen export system and effect of Asp24 in bacterial intracellular survival.
abstract: We previously showed that Brucella abortus rough mutant strain 2308 ΔATP (called the ΔrfbE mutant in this study) exhibits reduced intracellular survival in RAW264.7 cells and attenuated persistence in BALB/c mice. In this study, we performed microarray analysis to detect genes with differential expression between the ΔrfbE mutant and wild-type strain S2308. Interestingly, acid shock protein 24 gene (asp24) expression was significantly upregulated in the ΔrfbE mutant compared to S2308, as confirmed by quantitative reverse transcription-PCR (qRT-PCR) and Western blotting. Further studies using additional strains indicated that the upregulation of asp24 occurred only in rough mutants with disrupted O-antigen export system components, including the ATP-binding protein gene rfbE (bab1_0542) and the permease gene rfbD (bab1_0543), while the ΔwboA rough mutant (which lacks an O-antigen synthesis-related glycosyltransferase) and the RB51 strain (a vaccine strain with the rough phenotype) showed no significant changes in asp24 expression compared to S2308. In addition, abolishing the intracellular O-antigen synthesis of the ΔrfbE mutant by deleting the wboA gene (thereby creating the ΔrfbE ΔwboA double-knockout strain) recovered asp24 expression. These results indicated that asp24 upregulation is associated with intracellular O-antigen synthesis and accumulation but not with the bacterial rough phenotype. Further studies indicated that asp24 upregulation in the ΔrfbE mutant was associated neither with bacterial adherence and invasion nor with cellular necrosis on RAW264.7 macrophages. However, proper expression of the asp24 gene favors intracellular survival of Brucella in RAW264.7 cells and HeLa cells during an infection. This study reveals a novel mechanism for asp24 upregulation in B. abortus mutants.
authors.affiliation: Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences (CAAS), Shanghai, People's Republic of China.
authors.forename: Mingxing
authors.initials: M
authors.lastname: Tian
authors.affiliation: Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences (CAAS), Shanghai, People's Republic of China College of Veterinary Medicine, Yangzhou University, Yangzhou, People's Republic of China.
authors.forename: Jing
authors.initials: J
authors.lastname: Qu
authors.affiliation: Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences (CAAS), Shanghai, People's Republic of China.
authors.forename: Xiangan
authors.initials: X
authors.lastname: Han
authors.affiliation: Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences (CAAS), Shanghai, People's Republic of China.
authors.forename: Chan
authors.initials: C
authors.lastname: Ding
authors.affiliation: Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences (CAAS), Shanghai, People's Republic of China.
authors.forename: Shaohui
authors.initials: S
authors.lastname: Wang
authors.affiliation: College of Veterinary Medicine, Yangzhou University, Yangzhou, People's Republic of China.
authors.forename: Daxin
authors.initials: D
authors.lastname: Peng
authors.affiliation: Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences (CAAS), Shanghai, People's Republic of China yus@shvri.ac.cn.
authors.forename: Shengqing
authors.initials: S
authors.lastname: Yu
mesh_terms: D000818:Animals; D000942:Antigens, Bacterial / Q000235:genetics / Q000378:metabolism*; D001422:Bacterial Adhesion; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D002003:Brucella abortus / Q000235:genetics / Q000378:metabolism*; D002460:Cell Line; D015964:Gene Expression Regulation, Bacterial; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D009154:Mutation; D011817:Rabbits; D015854:Up-Regulation
chemical_list: D000942:Antigens, Bacterial; D001426:Bacterial Proteins
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