The heat-shock protein ClpB of Francisella tularensis is involved in stress tolerance and is required for multiplication in target organs of infected mice.
Intracellular bacterial pathogens generally express chaperones such as Hsp100s during multiplication in host cells, allowing them to survive potentially hostile conditions. Francisella tularensis is a highly infectious bacterium causing the zoonotic disease tularaemia. The ability of F. tularensis to multiply and survive in macrophages is considered essential for its virulence. Although previous mutant screens in Francisella have identified the Hsp100 chaperone ClpB as important for intracellular survival, no detailed study has been performed. We demonstrate here that ClpB of F. tularensis live vaccine strain (LVS) is important for resistance to cellular stress. Promoter analysis shows that the transcriptional start is preceded by a sigma32-like promoter sequence and we demonstrate that expression of clpB is induced by heat shock. This indicates that expression of clpB is dependent on the heat-shock response mediated by sigma32, the only alternative sigma-factor present in Francisella. Our studies demonstrate that ClpB contributes to intracellular multiplication in vitro, but is not essential. However, ClpB is absolutely required for Francisella to replicate in target organs and induce disease in mice. Proteomic analysis of membrane-enriched fractions shows that five proteins are recovered at lower levels in the mutant strain. The crucial role of ClpB for in vivo persistence of Francisella may be linked to its assumed function in reactivation of aggregated proteins under in vivo stress conditions.
Meibom
Karin L
KL
Université Paris Descartes, Faculté de Médecine René Descartes, Paris F-75015, France. kmeibom@necker.fr
Dubail
Iharilalao
I
Dupuis
Marion
M
Barel
Monique
M
Lenco
Juraj
J
Stulik
Jiri
J
Golovliov
Igor
I
Sjöstedt
Anders
A
Charbit
Alain
A
D000222:Adaptation, Physiological / Q000235:genetics / Q000502:physiology; D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D015180:Electrophoresis, Gel, Two-Dimensional; D018274:Electroporation; D005260:Female; D005604:Francisella tularensis / Q000235:genetics / Q000378:metabolism* / Q000648:ultrastructure; D006360:Heat-Shock Proteins / Q000235:genetics / Q000378:metabolism*; D006358:Hot Temperature; D008264:Macrophages / Q000382:microbiology*; D051379:Mice; D008807:Mice, Inbred BALB C; D050296:Microbial Viability; D046529:Microscopy, Electron, Transmission; D011401:Promoter Regions, Genetic / Q000235:genetics; D020543:Proteome / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012808:Sigma Factor / Q000378:metabolism; D024363:Transcription Initiation Site
D001426:Bacterial Proteins; D006360:Heat-Shock Proteins; D020543:Proteome; D012808:Sigma Factor