Brucella abortus Cell Cycle and Infection Are Coordinated

Abstract

Brucellae are facultative intracellular pathogens. The recent development of methods and genetically engineered strains allowed the description of cell-cycle progression of Brucella abortus, including unipolar growth and the ordered initiation of chromosomal replication. B. abortus cell-cycle progression is coordinated with intracellular trafficking in the endosomal compartments. Bacteria are first blocked at the G1 stage, growth and chromosome replication being resumed shortly before reaching the intracellular proliferation compartment. The control mechanisms of cell cycle are similar to those reported for the bacterium Caulobacter crescentus, and they are crucial for survival in the host cell. The development of single-cell analyses could also be applied to other bacterial pathogens to investigate their cell-cycle progression during infection.

Keywords: Brucella; cell cycle; growth; intracellular infection; replication.

Figure 1.. Cell Cycle of Brucella abortus and Caulobacter crescentus.

(A) Cell cycle of B. abortus. Cell division generates two daughter cells. The B. abortus generation time is approximately 210 min during exponential phase in rich medium, on average these bacteria duplicate and segregate oriI ~50 min after division. This corresponds to the G1/S transition of the cell cycle. The oriII is duplicated and segregated from ~105 min post-division, while a constriction is detected ~35 min before cell division. These constricting bacteria are also called predivisional. The red dot represents the oriI, and the green dot represents oriII. The proposed timing is an average generated from the proportion of cell types in a mixed population, all attempts to synchronize B. abortus cell cycle having failed to this point. The question mark indicates that a differentiation event is probably occurring at this step of the cell cycle [51]. (B) The cell cycle of C. crescentus [76]. Cell division generates two specialized daughter cells: a stalked cell able to rapidly reinitiate chromosome replication, and a swarmer cell going through a G1 phase before differentiating into a stalked cell. The duration of the cell cycle is given for a culture of C. crescentus in rich medium. The cell cycle time is about 105 min for swarmer cells, while it is about 85 min for stalked cells.

Figure 2.. Coordination of the Cell Cycle of Brucella with Intracellular Trafficking.

From a mixed population in culture, the G1 bacteria have a disproportionately high probability of entry into host cells (HeLa cells or RAW264.7 macrophages). Once inside the host cells, these bacteria remain at the G1 stage of the cell cycle for several hours, depending on the host cell type. When they are still in endosomes (endosomal Brucella-containing vacuoles, eBCVs), these bacteria resume growth and replication of chromosomes. Shortly after DNA replication initiation they are found in their replicative niche (relicative Brucella-containing vacuoles, rBCVs), that is, in the endoplasmic reticulum (ER). The asymmetric nature of Brucells abortus division is not shown here since the type of daughter (small or large) cell internalized was not determined.

Figure I.. A Conserved Cell-Cycle Regulatory Network in Alphaproteobacteria.

Protein–protein interactions are shown as black lines, pathways of phosphoryl transfer in blue, and transcriptional control in green. Cyclic di-GMP (cdG) is produced by PleD and hydrolyzed by PdeA, and it controls CckA activity (dashed black lines). The corresponding genes conserved in Brucella abortus are shown with a red frame, and partial heterocomplementation of Caulobacter crescentus divK and pleC mutants by homologous B. abortus genes [51] is shown in yellow. The yellow frame around the CckA-ChpT-CpdR-CtrA phosphorelay indicates that it is functional in B. abortus [49].