Abstract
ABSTRACTShigella spp. are highly adapted pathogens that cause bacillary dysentery in human and nonhuman primates. An unusual feature of Shigella pathogenesis is that this organism invades the colonic epithelia from the basolateral pole. Therefore, it has evolved the ability to disrupt the intestinal epithelial barrier to reach the basolateral surface. We have shown previously that the secreted serine protease A (SepA), which belongs to the family of serine protease autotransporters of Enterobacteriaceae, is responsible for the initial destabilization of the intestinal epithelial barrier that facilitates Shigella invasion. However, the mechanisms used by SepA to regulate this process remain unknown. To investigate the protein targets cleaved by SepA in the intestinal epithelium, we incubated a sample of homogenized human colon with purified SepA or with a catalytically inactive mutant of this protease. We discovered that SepA targets an array of 18 different proteins, including alpha-1 antitrypsin (AAT), a major circulating serine proteinase inhibitor in humans. In contrast to other serine proteases, SepA cleaved AAT without forming an inhibiting complex, which resulted in the generation of a neutrophil chemoattractant. We demonstrated that the products of the AAT-SepA reaction induce a mild but significant increase in neutrophil transepithelial migration in vitro. Moreover, the presence of AAT during Shigella infection stimulated neutrophil migration and dramatically enhanced the number of bacteria invading the intestinal epithelium in a SepA-dependent manner. We conclude that by cleaving AAT, SepA releases a chemoattractant that promotes neutrophil migration, which in turn disrupts the intestinal epithelial barrier to enable Shigella invasion.
Highlights
Shigella spp. are highly adapted pathogens that cause bacillary dysentery in human and nonhuman primates
We have shown previously that serine protease A (SepA) facilitates the bacterial transit of S. flexneri from the apical to the basolateral surface of the epithelium by disrupting the intestinal epithelial barrier [12]
To investigate the protein targets cleaved by SepA in the intestinal epithelium, we incubated a sample of homogenized human colon with purified wild-type (WT) SepA or with a catalytically inactive mutant of this protease, serine 211-to-alanine point mutation (S211A) SepA
Summary
Shigella spp. are highly adapted pathogens that cause bacillary dysentery in human and nonhuman primates. We conclude that by cleaving AAT, SepA releases a chemoattractant that promotes neutrophil migration, which in turn disrupts the intestinal epithelial barrier to enable Shigella invasion. The second mechanism posits that Shigella creates a breach in the colonic epithelium to facilitate paracellular transit into the submucosa [10,11,12] Shigella generates this breach directly by disrupting the intercellular junctions [10] or destabilizing the intestinal epithelial barrier [12] or indirectly by stimulating the transepithelial migration of neutrophils from the submucosa to the intestinal lumen, which opens up intercellular spaces that allow Shigella paracellular transit [11]
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