The Type III Secreted Protein BspR Regulates the Virulence Genes in Bordetella bronchiseptica

Abstract

Bordetella bronchiseptica is closely related with B. pertussis and B. parapertussis, the causative agents of whooping cough. These pathogenic species share a number of virulence genes, including the gene locus for the type III secretion system (T3SS) that delivers effector proteins. To identify unknown type III effectors in Bordetella, secreted proteins in the bacterial culture supernatants of wild-type B. bronchiseptica and an isogenic T3SS-deficient mutant were compared with iTRAQ-based, quantitative proteomic analysis method. BB1639, annotated as a hypothetical protein, was identified as a novel type III secreted protein and was designated BspR (Bordetella secreted protein regulator). The virulence of a BspR mutant (ΔbspR) in B. bronchiseptica was significantly attenuated in a mouse infection model. BspR was also highly conserved in B. pertussis and B. parapertussis, suggesting that BspR is an essential virulence factor in these three Bordetella species. Interestingly, the BspR-deficient strain showed hyper-secretion of T3SS-related proteins. Furthermore, T3SS-dependent host cell cytotoxicity and hemolytic activity were also enhanced in the absence of BspR. By contrast, the expression of filamentous hemagglutinin, pertactin, and adenylate cyclase toxin was completely abolished in the BspR-deficient strain. Finally, we demonstrated that BspR is involved in the iron-responsive regulation of T3SS. Thus, Bordetella virulence factors are coordinately but inversely controlled by BspR, which functions as a regulator in response to iron starvation.

Differential regulation of type III secretion and virulence genes inBordetella pertussisandBordetella bronchisepticaby a secreted anti-σ factor

“…In WT RB50, overexpression of btrA decreased expression of T3SS-associated genes but did not confer increased expression of toxin or adhesin loci, suggesting that BtrA levels in RB50 are sufficient for maximal expression under laboratory conditions. Together, these data confirm and extend previously published results (28) and show that BtrA differentially regulates three distinct modules of the BvgAS regulon: (i) Bvg + phase adhesin and toxin genes, which require BtrA for full level expression, (ii) Bvg + phase T3SS loci that require BtrS and are repressed by BtrA, and (iii) the Bvg phase flagellin locus, which is down-regulated by BtrA (Fig. 1G).…”

“…Recent studies implicate BtrA (also called BspR), which is encoded directly upstream of btrS, as an exported substrate of the Bsc T3SS, illustrated in Fig. 1B using B. bronchiseptica strain RB50 grown in vitro under conditions permissive for type III secretion (20,28). BtrA and Bsp22, a known T3SS substrate (26), are exported into supernatants in a manner dependent on the BscN ATPase.…”

“…Here, we characterize a regulatory node involving a T3SS-exported anti-σ factor, BtrA (20,28), and demonstrate its activity as a secreted BtrS antagonist that differentially controls expression of nearly 300 genes that define six distinct regulatory modules within the BvgAS virulence regulon. In B. bronchiseptica, the relative levels of BtrA and BtrS determine the magnitude of T3SS activity.…”

Glutamate Limitation, BvgAS Activation, and (p)ppGpp Regulate the Expression of the Bordetella pertussis Type 3 Secretion System

“…Its product, BspR, is one of the T3SS secreted proteins that is translocated into host cell nuclei and is also a negative transcriptional regulator of T3SS genes in the bacterial cytosol (26). To assess the effect of BspR on the expression of the T3SS, the levels of BspR were analyzed by immunoblotting.…”

Bordetella effector BopN is translocated into host cells via its N‐terminal residues

“…BteA, which triggers cytotoxicity in a wide variety of cultured cell lines , is translocated into the host cells via 48 aa residues at its N‐terminal. We reported in 2012 that BspR acts as a transcriptional regulator that positively and negatively regulates various virulence and non‐virulence genes . More recently, we observed that BspR is translocated into the nuclei of infected cells .…”