Brucella abortus Lon functions as a generalized stress response protease and is required for wild-type virulence in BALB/c mice

The gene encoding a Lon protease homologue has been cloned from Brucella abortus. The putative Brucella abortus Lon shares > 60% amino acid identity with its Escherichia coli counterpart and the recombinant form of this protein restores the capacity of an Escherichia coli lon mutant to resist kil...

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Bibliographic Details
Published in:Molecular microbiology 2000-02, Vol.35 (3), p.577-588
Main Authors: Robertson, G.T, Kovach, M.E, Allen, C.A, Ficht, T.A, Roop, R.M. II
Format: Article
Language:English
Subjects:
Adaptation, Physiological
Adenosine Triphosphate - metabolism
Animals
ATP-Dependent Proteases
Brucella abortus
Brucella abortus - enzymology
Brucella abortus - pathogenicity
Brucella abortus - physiology
Brucellosis - microbiology
Cloning, Molecular
Escherichia coli Proteins
Gene Expression Regulation, Bacterial
Heat-Shock Proteins - genetics
Heat-Shock Proteins - metabolism
Heat-Shock Response
Lon proteinase
Macrophages, Peritoneal - microbiology
Mice
Mice, Inbred BALB C
Molecular Sequence Data
Mutation
Protease La
proteinases
Recombinant Proteins - genetics
Recombinant Proteins - metabolism
Serine Endopeptidases - genetics
Serine Endopeptidases - metabolism
Stress, Physiological
Transcription, Genetic
Virulence - physiology
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Summary:The gene encoding a Lon protease homologue has been cloned from Brucella abortus. The putative Brucella abortus Lon shares > 60% amino acid identity with its Escherichia coli counterpart and the recombinant form of this protein restores the capacity of an Escherichia coli lon mutant to resist killing by ultraviolet irradiation and regulate the expression of a cpsB::lacZ fusion to wild‐type levels. A σ32 type promoter was identified upstream of the predicted lon coding region and Northern analysis revealed that transcription of the native Brucella abortus lon increases in response to heat shock and other environmental stresses. ATP‐dependent proteolytic activity was also demonstrated for purified recombinant Lon. To evaluate the capacity of the Brucella abortus Lon homologue to function as a stress response protease, the majority of the lon coding region was removed from virulent strain Brucella abortus 2308 via allelic exchange. In contrast to the parent strain, the Brucella abortus lon mutant, designated GR106, was impaired in its capacity to form isolated colonies on solid medium at 41°C and displayed an increased sensitivity to killing by puromycin and H2O2. GR106 also displayed reduced survival in cultured murine macrophages and significant attenuation in BALB/c mice at 1 week post infection compared with the virulent parental strain. Beginning at 2 weeks and continuing for 6 weeks post infection, however, GR106 and 2308 displayed equivalent spleen and liver colonization levels in mice. These findings suggest that the Brucella abortus Lon homologue functions as a stress response protease that is required for wild‐type virulence during the initial stages of infection in the mouse model, but is not essential for the establishment and maintenance of chronic infection in this host.
ISSN:0950-382X
1365-2958
DOI:10.1046/j.1365-2958.2000.01726.x