Diverse Genetic Regulon of the Virulence-Associated Transcriptional Regulator MucR in Brucella abortus 2308

Abstract

The Ros-type regulator MucR is one of the few transcriptional regulators that have been linked to virulence in Brucella. Here, we show that a Brucella abortus in-frame mucR deletion strain exhibits a pronounced growth defect during in vitro cultivation and, more importantly, that the mucR mutant is attenuated in cultured macrophages and in mice. The genetic basis for the attenuation of Brucella mucR mutants has not been defined previously, but in the present study the genes regulated by MucR in B. abortus have been elucidated using microarray analysis and real-time reverse transcription-PCR (RT-PCR). In B. abortus 2308, MucR regulates a wide variety of genes whose products may function in establishing and maintaining cell envelope integrity, polysaccharide biosynthesis, iron homeostasis, genome plasticity, and transcriptional regulation. Particularly notable among the MucRregulated genes identified is arsR6 (nolR), which encodes a transcriptional regulator previously linked to virulence in Brucella melitensis 16 M. Importantly, electrophoretic mobility shift assays (EMSAs) determined that a recombinant MucR protein binds directly to the promoter regions of several genes repressed by MucR (including arsR6 [nolR]), and in Brucella, as in other alphaproteobacteria, MucR binds to its own promoter to repress expression of the gene that encodes it. Overall, these studies have uncovered the diverse genetic regulon of MucR in Brucella, and in doing so this work has begun to define the MucR-controlled genetic circuitry whose misregulation contributes to the virulence defect of Brucella mucR mutants.

RegA Plays a Key Role in Oxygen-Dependent Establishment of Persistence and in Isocitrate Lyase Activity, a Critical Determinant of In vivo Brucella suis Pathogenicity

“…Induction of the transcriptional regulator MucR represents an outstanding example of such a major role for RegA, since MucR acts mainly as a repressor of Brucella genes involved in very diverse functions (Caswell et al, 2013; Mirabella et al, 2013). In the late exponential growth phase, mucR expression was shown to be under negative control of the regulator VjbR (Mirabella et al, 2013).…”

Global Rsh-dependent transcription profile of Brucella suisduring stringent response unravels adaptation to nutrient starvation and cross-talk with other stress responses

“…GntR5 has been identified as HutC, a transcriptional regulator that exerts two different roles, as it acts as a co-activator of transcription of the virB operon, and represses the hut genes implicated in the histidine utilization pathway [34]. The regulator MucR has been described as being involved in virulence of Brucella melitensis and B. abortus in macrophage and murine models of infection [35,36], in lipid A-core and cyclic-β-glucan synthesis in B. melitensis [37], and in the successful establishment of symbiosis in S. meliloti , including control of exopolysaccharide biosynthesis, necessary for biofilm formation [38,39]. …”

Coordinated Zinc Homeostasis Is Essential for the Wild-Type Virulence of Brucella abortus

“…Nonetheless, several zinc-containing proteins and enzymes are important for Brucella, including the superoxide dismutase C (SodC), which is required for Brucella virulence (10,11), the carbonic anhydrases I and II, and the histidinol dehydrogenase (12). Additionally, MucR is a zinc finger transcriptional regulatory protein that controls the expression of numerous virulence-associated genetic systems, and MucR is required for Brucella pathogenesis (13,14,39,40). Therefore, the attenuation of the ⌬zntR mutant strain may result from a combination of deficiencies related to the lack of sufficient zinc, including the failure of the bacteria to mount an appropriate oxidative stress response, multiple metabolic aberrations, and the inability to coordinate gene expression required for survival within the host.…”

“…It has been estimated that Ͼ5% of bacterial proteins bind Zn (9), and several Zn-containing proteins that are important for the basic physiology and virulence of Brucella strains have been identified (10,11,12,13,14,15). Moreover, the Zn uptake system protein ZnuA is required for Brucella virulence (16, 17), and Bru- …”

“…These metal ions have extremely high binding affinities for generic divalent cation binding sites in proteins, and the binding of these cations to inappropriate sites, such as sites requiring Fe or Mn for proper protein function, can inactivate the proteins, leading to toxicity and cell death (6). Thus, it is not surprising that organisms from single-celled bacteria to multicellular mammals have evolved cellular mechanisms to stringently control the uptake, export, utilization, and storage of metal ions.It has been estimated that Ͼ5% of bacterial proteins bind Zn (9), and several Zn-containing proteins that are important for the basic physiology and virulence of Brucella strains have been identified (10,11,12,13,14,15). Moreover, the Zn uptake system protein ZnuA is required for Brucella virulence (16, 17), and Bru- …”