Project description:We recently reported that the in vitro and in vivo survival of Rickettsia australis are Atg5-dependent, in association with an inhibited level of anti-rickettsial cytokine, IL-1β. In the present study, we sought to investigate how R. australis interacts with host innate immunity via an Atg5-dependent autophagic response. We found that the serum levels of IFN-γ and G-CSF in R. australis-infected Atg5flox/floxLyz-Cre mice were significantly less compared to Atg5flox/flox mice, accompanied by significantly lower rickettsial loads in tissues with inflammatory cellular infiltrations including neutrophils. R. australis infection differentially regulated a significant number of genes in bone marrow-derived macrophages (BMMs) in an Atg5-depdent fashion as determined by RNA sequencing and Ingenuity Pathway Analysis, including genes in the molecular networks of IL-1 family cytokines and PI3K-Akt-mTOR. The secretion levels of inflammatory cytokines, such as IL-1α, IL-18, TNF-α, and IL-6, by R. australis-infected Atg5flox/floxLyz-Cre BMMs were significantly greater compared to infected Atg5flox/flox BMMs. Interestingly, R. australis significantly increased the levels of phosphorylated mTOR and P70S6K at a time when the autophagic response is induced. Rapamycin treatment nearly abolished the phosphorylated mTOR and P70S6K but did not promote significant autophagic flux during R. australis infection. These results highlight that R. australis modulates an Atg5-dependent autophagic response, which is not sensitive to regulation by mTORC1 signaling in macrophages. Overall, we demonstrate that R. australis counteracts host innate immunity including IL-1β-dependent inflammatory response to support the bacterial survival via an mTORC1-resistant autophagic response in macrophages.
Project description:Secretion systems are used as weapons by a variety of Gram-negative bacteria. Among them the Type VI Secretion System (T6SS) gained more interest throughout the last years. The system functions as a molecular nano-weapon: it is used in inter-kingdom competition by various bacteria to deliver toxic effectors in target cells. Here we describe the role of the T6SS in Photorhabdus laumondii subsp. laumondii strain DJC, an entomopathogenic biocontrol agent able to live in different environmental niches, such as in symbiosis with nematodes and in the rhizosphere on plant roots. Using bioinformatic and protein motif analyses we identified four T6SS gene clusters (T6SS-1, T6SS-2, T6SS-3 and T6SS-4) and multiple orphan T6SS related genes in the genome of P. laumondii. Furthermore, we highlighted 11 T6SS effector-immunity pairs, including three undescribed membrane disrupting effectors, each with putatively different antibacterial activities. By label-free mass spectrometry of P. laumondii wild type cells and respective T6SS-deficient strains, we could point out a cross-link between T6SS and other Photorhabdus’ virulence related mechanisms such as PVCs, T3SS and pyocins. Furthermore, a change in motility as well as in the secondary metabolism was observed upon T6SS-deficiency. Here, we shed light on the T6SS in P. laumondii DJC and suggesting a cross-link of various virulence mechanisms, which could help to gain knowledge on T6SS and better figure out the Photorhabdus ability to live in polymicrobial environments.