title: Burkholderia mallei tssM encodes a putative deubiquitinase that is secreted and expressed inside infected RAW 264.7 murine macrophages.
abstract: Burkholderia mallei, a category B biothreat agent, is a facultative intracellular pathogen that causes the zoonotic disease glanders. The B. mallei VirAG two-component regulatory system activates the transcription of approximately 60 genes, including a large virulence gene cluster encoding a type VI secretion system (T6SS). The B. mallei tssM gene encodes a putative ubiquitin-specific protease that is physically linked to, and transcriptionally coregulated with, the T6SS gene cluster. Mass spectrometry and immunoblot analysis demonstrated that TssM was secreted in a virAG-dependent manner in vitro. Surprisingly, the T6SS was found to be dispensable for the secretion of TssM. The C-terminal half of TssM, which contains Cys and His box motifs conserved in eukaryotic deubiquitinases, was purified and biochemically characterized. Recombinant TssM hydrolyzed multiple ubiquitinated substrates and the cysteine at position 102 was critical for enzymatic activity. The tssM gene was expressed within 1 h after uptake of B. mallei into RAW 264.7 murine macrophages, suggesting that the TssM deubiquitinase is produced in this intracellular niche. Although the physiological substrate(s) is currently unknown, the TssM deubiquitinase may provide B. mallei a selective advantage in the intracellular environment during infection.
authors.affiliation: Department of Biochemistry, Emory University School of Medicine, Atlanta, Georgia 30322, USA.
authors.forename: John
authors.initials: J
authors.lastname: Shanks
authors.forename: Mary N
authors.initials: MN
authors.lastname: Burtnick
authors.forename: Paul J
authors.initials: PJ
authors.lastname: Brett
authors.forename: David M
authors.initials: DM
authors.lastname: Waag
authors.forename: Kevin B
authors.initials: KB
authors.lastname: Spurgers
authors.forename: Wilson J
authors.initials: WJ
authors.lastname: Ribot
authors.forename: Mark A
authors.initials: MA
authors.lastname: Schell
authors.forename: Rekha G
authors.initials: RG
authors.lastname: Panchal
authors.forename: Frank C
authors.initials: FC
authors.lastname: Gherardini
authors.forename: Keith D
authors.initials: KD
authors.lastname: Wilkinson
authors.forename: David
authors.initials: D
authors.lastname: Deshazer
mesh_terms: D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D042726:Burkholderia mallei / Q000201:enzymology* / Q000235:genetics / Q000472:pathogenicity*; D002460:Cell Line; D006224:Cricetinae; D010450:Endopeptidases / Q000235:genetics / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D005896:Glanders / Q000382:microbiology / Q000401:mortality; D054884:Host-Pathogen Interactions; D008264:Macrophages / Q000201:enzymology / Q000382:microbiology*; D008647:Mesocricetus / Q000382:microbiology; D051379:Mice; D064570:Ubiquitin-Specific Proteases
chemical_list: D001426:Bacterial Proteins; D010450:Endopeptidases; D064570:Ubiquitin-Specific Proteases