The light-activated signal transduction pathway in Brucella

PARIS GASTON; TONG-SENG TSENG; MARCUS FREDERICKSON; WINSLOW R. BRIGGS; ROBERTO A. BOGOMOLNI; FERNANDO A. GOLDBAUM

Ventura

Conferencia; Gordon Conference on Signal Transduction in Microorganisms; 2010

Gordon Research Conference

p { margin-bottom: 0.21cm; } Brucella is responsible of one of the world's most widespread zoonotic diseases, causing miscarriage and infertility in animals and a febrile disease, known as Malta Fever or Undulant fever in humans. Brucella can be considered a furtive pathogen that is devoid of classical virulence factors. Instead, the true virulence of Brucella resides in its ability to adapt to environmental conditions encountered in its replicative niche. Therefore, the control mechanisms that regulate metabolic adaptation are very important players in the infection process. We have shown that Brucella genome encodes for sensory histidine kinase that contains a LOV sensor domain. LOV domains are light sensory modules that bind flavins and undergo a self-contained photocycle that is dependent on the presence of a conserved cys residue. Upon illumination the cys forms a covalent bond between the sulfur and C4a carbon of FMN. We have demostrated that the LOV-histidine kinase (LOV-HK) from Brucella functions as the photoreceptor in a blue light-activated signalling pathway that up-regulates virulence in a macrophage infection model. Thus, LOV-HK autophosphorylation is increased upon illumination and the mutation C69A which cannot form the covalent adduct shown no increase of phosphorylation, confirming that the covalent state is the signalling form of the protein. Deletion of the LOV-HK gene in Brucella shown an attenuated phenotype in cell infection assays and this effect could be complemented by the wild type LOV-HK but not by LOV-HK C69A, suggesting that LOV-HK works as light-activated kinase in Brucella infection. Using bacterial two-hybrid we have identified a single domain response regulator (RR) protein from Brucella as the partner of LOV-HK. The RR protein identified contains a single receiver domain with an organization similar to CheY proteins from others bacterias. Because this similarity we named it as CheY-L. Fast phosphotransfer between LOV-HK and CheY-L confirmed that the observed interaction. Bacterial two-hybrid assays also shown that CheY-L interacts with cytoplasmatic components of the flagella, FliM and FliG. We have identified CheY-L and flagellar proteins FliM and FliG as components of the signal transduction pathway initiated by light that regulates the virulence of Brucella.