Refining the Plasmid-Encoded Type IV Secretion System Substrate Repertoire of Coxiella burnetii

Abstract

The intracellular bacterial agent of Q fever, Coxiella burnetii, translocates effector proteins into its host cell cytosol via a Dot/ Icm type IV secretion system (T4SS). The T4SS is essential for parasitophorous vacuole formation, intracellular replication, and inhibition of host cell death, but the effectors mediating these events remain largely undefined. Six Dot/Icm substrate-encoding genes were recently discovered on the C. burnetii cryptic QpH1 plasmid, three of which are conserved among all C. burnetii isolates, suggesting that they are critical for conserved pathogen functions. However, the remaining hypothetical proteins encoded by plasmid genes have not been assessed for their potential as T4SS substrates. In the current study, we further defined the T4SS effector repertoire encoded by the C. burnetii QpH1, QpRS, and QpDG plasmids that were originally isolated from acute-disease, chronic-disease, and severely attenuated isolates, respectively. Hypothetical proteins, including those specific to QpRS or QpDG, were screened for translocation using the well-established Legionella pneumophila T4SS secretion model. In total, six novel plasmid-encoded proteins were translocated into macrophage-like cells by the Dot/Icm T4SS. Four newly identified effectors are encoded by genes present only on the QpDG plasmid from severely attenuated Dugway isolates, suggesting that the presence of specific effectors correlates with decreased virulence. These results further support the idea of a critical role for extrachromosomal elements in C. burnetii pathogenesis.

Right on Q: Genetics begin to Unravel Coxiella Burnetii host cell Interactions

“…Four of ten dot/icm genes tested complemented the corresponding mutant in L. pneumophila, suggesting functional similarity between the two systems [85,86]. Consequently, and owing to the lack of genetic tools for Coxiella, L. pneumophila was extensively used as a surrogate host to identify Coxiella T4BSS substrates using both CyaA and BlaM cytosolic translocation reporter assays [87][88][89][90][91][92][93][94][95][96]. Indeed, of the 143 currently recognized Coxiella Dot/Icm effectors, 125 were initially identified in L. pneumophila.…”

“…However, detection of native effectors is exceedingly difficult; thus, ectopic expression of effector-fluorescent protein chimeras has been extensively used for trafficking studies, despite known artifacts of this procedure [120]. Effectors traffic to mitochondria [87,92,94,121], endoplasmic reticulum [91,94,96,122], Golgi apparatus [87], lysosomes [87,92,107,122], autophagosomes [91,93], endocytic vesicles [73,107], nucleus [87,94,[121][122][123], microtubules [92,122] and ubiquitinated proteins [91,93]. An interesting subset of five effectors designated Cvp (Coxiella vacuolar protein) also traffic to the CCV [73,107].…”

“…Three effector genes are conserved between all plasmids and the integrated plasmid sequences [93], while three and four are uniquely encoded by QpH1 (carried by Nine Mile and Henzerling strains) and QpDG (carried by the Dugway strain), respectively [93]. The strict maintenance of plasmid sequences suggests they serve critical functions in Coxiella biology, with conserved effector genes predicted to play important roles in host cell manipulation [93]. By analyzing genome sequences from 10 Coxiella strains, Pearson et al [11] recently defined the Coxiella pan genome as consisting of 2148 genes, with 341 genes not conserved among all strains.…”

Identification of Novel Coxiella burnetii Icm/Dot Effectors and Genetic Analysis of Their Involvement in Modulating a Mitogen-Activated Protein Kinase Pathway

“…To date, about 100 effector proteins have been identified in C. burnetii, which constitute about 5% of its proteome. The C. burnetii effector proteins were identified thus far based on different criteria, including (i) existence of eukaryotic motifs such as ankyrin domains (18,19), (ii) presence of a PmrA binding site in their upstream regulatory region (20,21), (iii) identification of their C-terminal translocation signal (17), and (iv) location on plasmids (22,23). Most intriguingly, most of the effectors identified are unique proteins not present in any other sequenced organism (24,25).…”

Coxiella burnetii, agent de la fièvre Q

“…Ce qui fournit une explication plausible à la maintenance absolue des gènes plasmidiques chez ce pathogène . Les effecteurs du T4SS sont encodés par les plasmides Q pH1, Q pRS et Q pDG qui ont été respectivement isolés de fièvres Q aiguës, de fièvres Q chroniques et d'isolats sévèrement atténués (Maturana et al 2013). …”