Abstract
Coxiella burnetii is the causative agent of the zoonotic disease Q fever. To date, the lipopolysaccharide (LPS) is the only defined and characterized virulence determinant of C. burnetii. In this study, proteome profiles of C. burnetii Nine Mile phase I (RSA 493, NMI) and its isogenic Nine Mile phase II (RSA 439 NMII) isolate with a deep rough LPS were compared on L-929 mouse fibroblasts and in complex (ACCM-2), and defined (ACCM-D) media. Whole proteome extracts were analyzed using a label-free quantification approach. Between 659 and 1,046 C. burnetii proteins of the 2,132 annotated coding sequences (CDS) were identified in any particular experiment. Proteome profiles clustered according to the cultivation conditions used, indicating different regulation patterns. NMI proteome profiles compared to NMII in ACCM-D indicate transition from an exponential to a stationary phase. The levels of regulatory proteins such as RpoS, CsrA2, UspA1, and UspA2 were increased. Comparison of the oxidative stress response of NMI and NMII indicated that ACCM-2 represents a high oxidative stress environment. Expression of peroxidases, superoxide dismutases, as well as thioredoxins was increased for NMI. In contrast, in ACCM-D, only osmoregulation seems to be necessary. Proteome profiles of NMII do not differ and indicate that both axenic media represent similar oxidative stress environments. Deep rough LPS causes changes of the outer membrane stability and fluidity. This might be one reason for the observed differences. Proteins associated with the T4SS and Sec translocon as well as several effector proteins were detectable under all three conditions. Interestingly, none of these putatively secreted proteins are upregulated in ACCM-2 compared to ACCM-D, and L-929 mouse fibroblasts. Curiously, a higher similarity of proteomic patterns (overlapping up- and downregulated proteins) of ACCM-D and bacteria grown in cell culture was observed. Particularly, the proteins involved in a better adaptation or homeostasis in response to the harsh environment of the parasitophorous vacuole were demonstrated for NMI. This semi-quantitative proteomic analysis of C. burnetii compared axenically grown bacteria to those propagated in cell culture.
Highlights
Coxiella burnetii, a Gram-negative and obligate intracellular bacterium, is the etiological agent of query (Q) fever
Using the labelfree quantification (LFQ) approach described in this study, between 659 and 1,046 C. burnetii proteins of 2,132 annotated CDS were identified in each sample (Table S1)
The observed quantities of the proteins are depicted as log2 transformation of LFQ intensities and include the values imputed by Perseus-type value imputation (Cox et al, 2011, 2014)
Summary
A Gram-negative and obligate intracellular bacterium, is the etiological agent of query (Q) fever. This almost worldwide-distributed zoonotic disease manifests as an acute, flu-like illness or as a potentially lifethreatening chronic disease in humans (Maurin and Raoult, 1999). More than 100 effectors have been described, but only few of them are known to subvert the host cell-signaling pathways, including vesicle trafficking, apoptosis, autophagy, and immune responses, to ensure intracellular survival of C. burnetii (Larson et al, 2013; Moffatt et al, 2015; Mansilla Pareja et al, 2017; Schäfer et al, 2017; Weber et al, 2018). C. burnetii is highly adapted to this unique niche and employs several antioxidative stress strategies, such as expression of DNA repair genes and detoxification of reactive oxygen species (ROS) and reactive nitrogen species (RNS) released by the host (Briggs et al, 2008; Mertens et al, 2008; Siemsen et al, 2009; Hill and Samuel, 2011; Brennan et al, 2015)
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