The 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 ab ortus. The putative Brucella abortus Lon shares >60% amino acid identity wi th its Escherichia coli counterpart and the recombinant form of this protei n restores the capacity of an Escherichia coli ion mutant to resist killing by ultraviolet irradiation and regulate the expression of a cpsB::lacZ fus ion to wild-type levels. A sigma(32) type promoter was identified upstream of the predicted ion coding region and Northern analysis revealed that tran scription of the native Brucella abortus ion increases in response to heat shock and other environmental stresses. ATP-dependent proteolytic activity was also demonstrated for purified recombinant Lon, To evaluate the capacit y of the Brucella abortus Lon homologue to function as a stress response pr otease, the majority of the ion coding region was removed from virulent str ain Brucella abortus 2308 via allelic exchange. In contrast to the parent s train, the Brucella abortus ion mutant, designated GR106, was impaired in i ts capacity to form isolated colonies on solid medium at 41 degrees C and d isplayed an increased sensitivity to killing by puromycin and H2O2 GR106 al so displayed reduced survival in cultured murine macrophages and significan t attenuation in BALB/c mice at 1 week post infection compared with the vir ulent parental strain. Beginning at 2 weeks and continuing for 6 weeks post infection, however, GR106 and 2308 displayed equivalent spleen and liver c olonization levels in mice. These findings suggest that the Brucella abortu s Lon homologue functions as a stress response protease that is required fo r wild-type virulence during the initial stages of infection in the mouse m odel, but is not essential for the establishment and maintenance of chronic infection in this host.