Advances in the study of intracellular transport of brucela abortus 2308 (virulent strain) in macrophages

Abstract

In recent years, this research team has progressed in the study of intracellular transport of the virulent strain (2308) of Brucella abortus in macrophage cell lines (J-774 and Raw) through multiple microscopic approaches. We demonstrated the transit of B. abortus 2308 through compartments of the endocytic pathway, marking early endosomes and lysosomes with gold (20 and 60 nm, respectively). We have shown that B. abortus 2308 occupies two different types of compartments: phagolysosomes and modified phagosomes, significantly reducing their fusion to endosomes. On the other hand, macrophages transfected with GFP-Rabs were used to evaluate the location of Rab 5 and 11 proteins, involved in the vesicular transport of the cell. Confocal microscopy showed that B. abortus 2308 (stained with Rhodamine) recruits Rab11 to the phagosome membrane that contains them, in the form of discrete patches. In addition, B. abortus 2308 co-locates with vesicles that overexpress Rab 5 in a large proportion. The permanence of Rab 5 and 11 associated with phagosomes containing B. abortus 2308 suggests that the bacteria actively retain these Rabs to avoid maturation of the phagosome that contains them. Subsequently, the effect of kinase inhibitors (AKTi) on the multiplication and intracellular survival of B. abortus 2308 at different times post macrophage infection was studied by confocal microscopy. These results allowed confirming that B. abortus 2308 uses the AKT/AS160 pathway to activate Rabs involved in the transport of nutrients necessary for its replication and the generation of a safe intracellular site. The compound used as an AKT inhibitor could constitute a new pharmacological approach for the treatment of brucellosis.