KronmillerBrentCGRBEssentialRoleResponse.pdf
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Abstract
Successful host cell colonization by the Q fever pathogen, Coxiella burnetii, requires translocation of effector proteins into
the host cytosol by a Dot/Icm type 4B secretion system (T4BSS). In Legionella pneumophila, the two-component system (TCS)
PmrAB regulates the Dot/Icm T4BSS and several additional physiological processes associated with pathogenesis. Because PmrA
consensus regulatory elements are associated with some dot/icm and substrate genes, a similar role for PmrA in regulation of the
C. burnetii T4BSS has been proposed. Here, we constructed a C. burnetii pmrA deletion mutant to directly probe PmrA-mediated
gene regulation. Compared to wild-type bacteria, C. burnetii ΔpmrA exhibited severe intracellular growth defects that coincided
with failed secretion of effector proteins. Luciferase gene reporter assays demonstrated PmrA-dependent expression of 5 of
7 dot/icm operons and 9 of 11 effector-encoding genes with a predicted upstream PmrA regulatory element. Mutational analysis
verified consensus sequence nucleotides required for PmrA-directed transcription. RNA sequencing and whole bacterial cell
mass spectrometry of wild-type C. burnetii and the ΔpmrA mutant uncovered new components of the PmrA regulon, including
several genes lacking PmrA motifs that encoded Dot/Icm substrates. Collectively, our results indicate that the PmrAB TCS is a
critical virulence factor that regulates C. burnetii Dot/Icm secretion. The presence of PmrA-responsive genes lacking PmrA regulatory
elements also suggests that the PmrAB TCS controls expression of regulatory systems associated with the production of
additional C. burnetii proteins involved in host cell parasitism