A Zinc-dependent metalloproteinase in the intracellular adaptation of Brucella abortus in macrophages

Brucella abortus is a pathogen that survives in macrophages. Several virulence factors participate in this process, including the open reading frame (ORF) BAB1_0270 codifying of a Zinc-dependent metalloproteinase. Here, its contribution in the process of intracellular adaptation was analyzed by infecting RAW264.7 macrophages with the mutant B. abortus Δ270 strain. Results showed that this Zinc-dependent metalloproteinase is a cytoplasmic protein that conforms an operon with a transcriptional regulator, which may constitute a type II toxin-antitoxin system. Functionally, this Zinc-dependent metalloproteinase participated neither in the adherence nor the initial intracellular traffic of B. abortus in macrophages. Nevertheless, its deletion significantly increased the co-localization of B. abortus Δ270 with phagolysosomal cathepsin D, reducing both its co-localization with calnexin, present in endoplasmic reticulum derived vesicles, and its intracellular replication within macrophages. Besides, B. abortus Δ270-infected macrophages produced significantly higher levels of TNF-α, IL-6, CD80 and CD86 than B. abortus 2308, even when several genes involved in virulence ( vjbR , hutC , bvrR , virB1 ) were up-regulated in this mutant. Finally, its deletion significantly reduced the capacity of B. abortus Δ270 to adapt, grow and express several virulence factors under acidic conditions. Based on these results, we discuss the role of this Zinc-dependent metalloproteinase in the regulation of the virulence of this pathogen, concluding that it contributes significantly to the intracellular adaptation of B. abortus 2308 during the infection of macrophages.