The Brucella Cell Envelope

Abstract

The cell envelope is a multilayered structure that insulates the interior of bacterial cells from an often chaotic outside world. Common features define the envelope across the bacterial kingdom, but the molecular mechanisms by which cells build and regulate this critical barrier are diverse and reflect the evolutionary histories of bacterial lineages. Intracellular pathogens of the genus Brucella exhibit marked differences in cell envelope structure, regulation, and biogenesis when compared to more commonly studied gram-negative bacteria and therefore provide an excellent comparative model for study of the gram-negative envelope. We review distinct features of the Brucella envelope, highlighting a conserved regulatory system that links cell cycle progression to envelope biogenesis and cell division. We further discuss recently discovered structural features of the Brucella envelope that ensure envelope integrity and that facilitate cell survival in the face of host immune stressors.

Keywords: Alphaproteobacteria; gram negative; intracellular pathogen; zoonosis.

Figure 1

(a) Brucella spp., like other Rhizobiales, exhibit unipolar growth from the new cell pole. As the cell cycle progresses, cell growth shifts from being exclusively polar to being at both the pole and the nascent division site (growth sites colored orange). The Brucella developmental regulators, PdhS and DivK, exhibit dynamic polar localization to the inner membrane as a function of cell cycle. The new (N) pole and old (O) cell pole are marked. (b) Model of the CckA-ChpT-CtrA phosphorelay. The histidine kinase CckA autophosphorylates on a conserved histidine residue and transfers a phosphoryl group to a conserved aspartic acid residue on its C-terminal receiver domain. CckA~P transfers a phosphoryl group to the ChpT phosphotransferase, which can subsequently transfer this phosphoryl group to the receiver domain of CtrA. CtrA~P is a DNA-binding response regulator that modulates transcription of genes controlling cell polarity, division, and intracellular survival.

Figure 2

(a) Phylogenetic distribution and synteny of the envelope integrity protein B (eipB) genomic region in Proteobacteria; gene neighborhood is anchored on bamA. Brucella eipB is part of a highly conserved cell envelope gene cluster in the Proteobacteria that includes genes involved in outer membrane protein assembly (bamA), undecaprenyl phosphate biosynthesis, phospholipid synthesis (cdsA), lipopolysaccharide synthesis (lpxDAIB), and translation (ef-ts). eipB (pink) is conserved in the Rhizobiales group of the Alphaproteobacteria (phylogenetic classifications on the right). Skp is present outside the Rhizobiales. Phylogenetic tree is based on BamA protein sequence (left). (b) Crystal structures of Brucella EipB (PDB: 6NTR), Escherichia coli LolB (PDB: 1IWM), E. coli LolA (PDB: 1IWL), and E. coli BamA (PDB: 5OR1). Abbreviation: PDB, Protein Data Bank.

Figure 3

(a) Overview of the envelope layers of Brucella. Notably, a subset of Brucella outer membrane proteins are covalently linked to the peptidoglycan cell wall via a conserved sequence at the protein N terminus. (b) Chemical structure of Escherichia coli lipid A (100). (c) Chemical structure of Brucella spp. lipid A showing VLCFA tails and pyrophosphorylethanolamine modification of diaminoglucose backbone (outlined with dashed-line box). Structure adapted from model and data presented in References and . (d) Brucella smooth LPS structure showing lipid A, core oligosaccharide, and O-polysaccharide (O-antigen), based on models and data presented in References , , and . Abbreviations: Kdo, 3-deoxy-d-manno-2-octulosonic acid; LPS, lipopolysaccharide; OMP, outer membrane protein; PG, peptidoglycan; VLCFA, very-long-chain fatty acid.

Figure 4

(a) Phylogenetic distribution and synteny of the envelope integrity protein A (eipA) genomic region in Proteobacteria; gene neighborhood is anchored on eipA. In Brucella and other Rhizobiales, eipA is part of a genetic locus that encodes the essential cell cycle regulatory proteins CtrA and ChpT. (b) Crystal structures of Brucella EipA (PDB 5UC0) and a Burkholderia SYLF domain protein (PDB 7OFN). Abbreviation: PDB, Protein Data Bank.

Figure 5

(a) The GSR signaling pathway in Alphaproteobacteria is activated by environmental signals and integrates features of two-component and sigma factor–dependent regulation of gene expression. Studies of this pathway in Brucella abortus support a model in which phosphorylation of PhyR is controlled by two HWE-family sensor kinases: LovhK and Bab1_1673. PhyR phosphorylation promotes its binding to NepR, which releases σ^E1 to control transcription of select cell envelope genes that influence in vitro stress survival and host infection. The LOV and PAS sensory domains of the cytoplasmic GSR-activating kinase, LovhK, are labeled. The periplasmic CHASE family sensor domain of the transmembrane GSR-repressive kinase, Bab1_1673, is labeled. Solid lines indicate a direct interaction; dashed lines indicate interactions that may be direct or indirect. (b) Brucella spp. BvrR/BvrS is a conserved, archetypal two-component system that is a critical regulator of infection. In response to detection of intracellular signals during infection, the histidine kinase BvrS phosphorylates BvrR, which then regulates transcription of a suite of cell envelope and metabolic genes important for intracellular survival and replication. Abbreviations: DBD, DNA-binding domain; GSR, general stress response; REC, REC, receiver domain; RNAP, RNA polymerase; OMP, outer membrane protein.