Investigation of the replication and the role of (p)ppGpp in the pathogen Brucella abortus

Abstract

Brucella spp. are facultative intracellular bacteria responsible for Brucellosis, a
worldwide anthropozoonosis. This neglected disease is found in a variety of mammals and humans are considered as accidental hosts. The genome of B. abortus is divided into two chromosomes, and several tools were developed to monitor the state of chromosomes replication throughout the cell cycle at the single cell level. The study of chromosomes replication showed first that the chromosome I starts its replication before the chromosome II, indicating the existence of coordinated mechanisms regulating the initiation of replication of both chromosomes. Secondly, these investigations revealed that B. abortus presents a biphasic infection process, with a first non-proliferation phase characterized by a G1 arrest, and a second phase in which bacteria restart their cell cycle, and actively proliferate. Moreover, G1 bacteria were shown to be preferentially internalized inside host cells, and thus represented the infectious form of the pathogen. These results demonstrated that the cell cycle of B. abortus is intimately linked to its infection strategy.
Here, we investigated mechanisms potentially involved in the control of the cell cycle, especially in chromosomes replication regulation, focusing our researches on conserved actors already known to regulate DNA replication in Escherichia coli and Caulobacter crescentus. We notably found that the alarmone (p)ppGpp plays a role in this control. Indeed, the overproduction of (p)ppGpp in B. abortus led to growth and replication defects, reflecting a general cell cycle delay. The impact observed on replication could be explained, in part, by the alteration of the level of the replication initiator DnaA upon (p)ppGpp overproduction.
Finally, we showed that (p)ppGpp is important for the establishment of a successful infection process, since mutants which cannot produce (p)ppGpp, or which are depleted for this alarmone were strongly attenuated during the infection.

Date of Award	15 Nov 2019
Original language	English
Awarding Institution	University of Namur
Sponsors	Fund for Research Training in Industry and Agriculture (FRIA)
Supervisor	Xavier De Bolle (Supervisor), Regis Hallez (Jury), Stephan Köhler (Jury), Laurence Van Melderen (Jury) & Justine Collier (Jury)