The extracellular loops of Salmonella Typhimurium outer membrane protein A (OmpA) maintain the stability of Salmonella containing vacuole (SCV) in murine macrophages and protect the bacteria from autophagy-dependent lysosomal degradation
After entering the host cells, Salmonella Typhimurium (STM) stays inside a modified acidic compartment called Salmonella containing vacuole (SCV). The biogenesis and stability maintenance of SCV are critical but inadequately understood areas. Our research has provided a novel mechanistic view on the role of a bacterial porin OmpA in maintaining the stability of SCV. We found that the deletion of OmpA forces the bacteria to escape from the SCV during the immediate early stage of infection. Within murine and human monocyte-derived macrophages, in the absence of OmpA, the bacteria were inefficient in retaining the LAMP-1, a standard SCV marker around itself. The cytosolic population of the mutant bacteria already left the SCV can activate the host autophagy machinery. On the contrary to the wild type bacteria confined within the SCV, STM ΔompA colocalizes more with autophagy markers syntaxin 17 and LC3B. Inhibition of autophagy pathway using bafilomycin A1 restored the intracellular proliferation of STM ΔompA. The activation of the autophagy pathway further targeted the autophagosomes carrying STM ΔompA to the lysosomes for degradation. We further showed that the four extracellular loops of OmpA exposed outside played a crucial role in retaining the LAMP-1 pool around the SCV. We have altered the extracellular loop sequences of OmpA by site-directed mutagenesis and observed that the bacteria could not hold the LAMP-1 pool around the SCV and slowly started losing the SCV membrane, which finally led to the release of the loop mutants into the cytosol of the host macrophages. We ultimately proved that the cytosolic population of STM having mutations in the OmpA extracellular loops does not activate the lysosomal degradation pathway like STM ΔompA, which helps them to survive within the murine macrophages.