Stimulation of autophagy suppresses the intracellular survival ofBurkholderia pseudomalleiin mammalian cell lines

Abstract

Burkholderia pseudomallei is the causative agent of melioidosis, a tropical infection of humans and other animals. The bacterium is an intracellular pathogen that can escape from endosomes into the host cytoplasm, where it replicates and infects adjacent cells. We investigated the role played by autophagy in the intracellular survival of B. pseudomallei in phagocytic and non-phagocytic cell lines. Autophagy was induced in response to B. pseudomallei invasion of murine macrophage (RAW 264.7) cells and a proportion of the bacteria co-localized with the autophagy effector protein LC3, a marker for autophagosome formation. Pharmacological stimulation of autophagy in RAW 264.7 and murine embryonic fibroblast (MEF) cell lines resulted in increased co-localization of B. pseudomallei with LC3 while basal levels of co-localization could be abrogated using inhibitors of the autophagic pathway. Furthermore, induction of autophagy decreased the intracellular survival of B. pseudomallei in these cell lines, but bacterial survival was not affected in MEF cell lines deficient in autophagy. Treatment of infected macrophages with chloramphenicol increased the proportion of bacteria within autophagosomes indicating that autophagic evasion is an active process relying on bacterial protein synthesis. Consistent with this hypothesis, we identified a B. pseudomallei type III secreted protein, BopA, which plays a role in mediating bacterial evasion of autophagy. We conclude that the autophagic pathway is a component of the innate defense system against invading B. pseudomallei, but which the bacteria can actively evade. However, when autophagy is pharmacologically induced using rapamycin, bacteria are actively sequestered in autophagosomes, ultimately decreasing their survival.

Burkholderia pseudomallei survival in lung epithelial cells benefits from miRNA-mediated suppression of ATG10

“…19 Consistent with the previous study, 12 there was an increase at 2 h and 4 h p.i. in the ratio of MAP1LC3B-II to ACTB and a degradation of SQSTM1 with B. pseudomallei infection as compared to uninfected controls in A549 cells ( Fig.…”

“…Previous studies have reported that autophagic evasion of B. pseudomallei is an active process relying on TTSS effector, BopA, which shows 23% amino acid identity to the Shigella TTSS effector, IcsB. 12,22 In Shigella, IcsB competitively binds to its surface protein VirG to inhibit the autophagic process; ATG5 recognizes VirG. 8 Therefore, in a manner analogous to IcsB, it appears unlikely that BopA is the key factor to interfere with ATG10 expression upon B. pseudomallei infection.…”

“…12 Thus, the detailed mechanism by which B. pseudomallei-mediates subversion of autophagy should be further clarified.…”

CASP4/caspase-11 promotes autophagosome formation in response to bacterial infection

“…Accordingly, inflammatory CASP1 has been described to coordinate autophagy in response to L. pneumophila [24]. However, this is the first report of CASP4 deficiency negatively affecting autophagy in macrophages, which may explain the ability of other autophagy-interacting bacteria to be cleared in the presence of CASP4 [2,8,16]. In contrast, clearance of nonpathogenic E. coli does not depend on CASP1 or CASP4 [16,25].…”

“…This bacteria-specific autophagic process, designated xenophagy, promotes bacterial clearance and prevents cellular escape. Bacteria such as Listeria monocytogenes [6,7] or Burkholderia pseudomallei [8], although recognized by members of the autophagic pathway, have developed mechanisms to avoid lysosomal degradation and xenophagy.…”