Project description:We have previously reported that Rickettsia conorii and Rickettsia montanensis have distinct intracellular fates within THP-1 macrophages, suggesting that the ability to proliferate within macrophages may be a distinguishable factor between pathogenic and non-pathogenic Spotted fever group (SFG) members. To start unraveling the molecular mechanisms underlying the capacity (or not) of SFG Rickettsia to establish their replicative niche in macrophages, we have herein profiled the host proteomic alterations resulted by the infection of THP-1 macrophages with R. conorii and R. montanensis using a high throughput quantitative proteomics approach (SWATH-MS). Our results revealed that these two members of SFG Rickettsia with distinct pathogenicity attributes for humans, trigger differential proteomic signatures in macrophage-like cells. Although infection by both rickettsial species resulted in a lower abundance of enzymes of glycolysis and pentose phosphate pathway, the pathogenic R. conorii specifically induced the accumulation of several enzymes of the tricarboxylic acid cycle, oxidative phosphorylation, fatty acid -oxidation and glutaminolysis, as well as of several inner and outer membrane mitochondrial transporters. These results suggest a profound metabolic rewriting of macrophages by R. conorii towards a metabolic signature of an M2-like (anti-inflammatory) activation program. Moreover, our results revealed that several subunits forming the proteasome and immunoproteasome are found in lower abundance upon infection with both rickettsial species, which may help bacteria to escape immune surveillance. Remarkably, R. conorii-infection specifically induced the accumulation of several host proteins implicated in protein processing and quality control in ER, suggesting that this pathogenic Rickettsia may be able to compensate the accumulation of misfolded proteins by increasing the ER protein folding capacity and subsequently restore host cell homeostasis. This work reveals novel aspects of macrophage-Rickettsia interactions, expanding our knowledge of how pathogenic rickettsiae explore host cells to their advantage.

Project description:Rickettsia conorii, the causative agent of the MSF is transmitted to humans by the bite of infected ticks Rhipicephalus sanguineus. The skin thus constitutes an important barrier for the entry and propagation of R. conorii. Given this, analysis of the survival strategies used by the bacterium within infected skin is critical for our understanding of rickettsiosis. Here, we report the first genome-wide analysis of R. conorii gene expression from infected human skin biopsies. Our data showed that R. conorii exhibit a striking transcript signature that was remarkably conserved across patients, regardless of genotype. The expression profiles obtained using custom Agilent microarrays were validated by quantitative RT-PCR. Within eschars, approximately 15% (n = 211) of the total predicted R. conorii ORFs appeared differentially expressed. These genes are mostly down-regulated and encode proteins essential for bacterial replication. Some of the strategies displayed by rickettsiae to overcome the host defense barriers, thus avoiding killing, were also pointed out. The observed up-regulation of rickettsial genes associated with DNA repair is likely to correspond to a DNA-damaging agent enriched environment generated by the host cells to eradicate the pathogens. Survival of R. conorii within eschars also involves adaptation to osmotic stress, changes in cell surface proteins and up-regulation of some virulence factors. Interestingly, in contrast to down-regulated transcripts, we noticed that up-regulated ones rather exhibit a small nucleotide size, most of them being exclusive for the spotted fever group rickettsiae. On the overall, this work provides some insights on how R. conorii counteracts the host cell attack at the site of inoculation which corresponds to the front line of human host defense against rickettsia diffusion and that resembles to battlefield. 9 R.conorii clinical isolates from eschars co-hybridized with reference strain R.conorii Strain Malish 7

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