A genome‐wide screen identifies a Bordetella type III secretion effector and candidate effectors in other species

Abstract

SummaryBordetella bronchiseptica utilizes a type III secretion system (TTSS) for induction of non-apoptotic cytotoxicity in host cells and modulation of host immunity. The identity of Bordetella TTSS effectors, however, has remained elusive. Here we report a genome-wide screen for TTSS effectors based on shared biophysical and functional characteristics of class I chaperones and their frequent colocalization with TTSS effectors. When applied to B. bronchiseptica , the screen identified the first TTSS chaperone-effector locus, btcA-bteA , and we experimentally confirmed its function. Expression of bteA is co-ordinated with expression of TTSS apparatus genes, BteA is secreted through the TTSS of B. bronchiseptica , it is required for cytotoxicity towards mammalian cells, and it is highly conserved in the human-adapted subspecies B. pertussis and B. parapertussis . Transfection of bteA into epithlieal cells results in rapid cell death, indicating that BteA alone is sufficient to induce potent cytotoxicity. Finally, an in vitro interaction between BteA and BtcA was demonstrated. The search for TTSS chaperones and effectors was then expanded to other bacterial genomes, including mammalian and insect pathogens, where we identified a large number of novel candidate chaperones and effectors. Although the majority of putative effectors are proteins of unknown function, several have similarities to eukaryotic protein domains or previously identified effectors from other species.

Transcriptional Profiling of the Iron Starvation Response in Bordetella pertussis Provides New Insights into Siderophore Utilization and Virulence Gene Expression

“…Analysis of the B. pertussis Tohama I derivative BP953 (81), carrying chromosomal fhaB-lacZ and ptx-phoA transcriptional fusions, revealed only modest elevation of fhaB and ptxA expression under iron-depleted growth conditions as compared with iron-replete conditions (46% and 14% increases in reporter activity after 48 h of culturing, respectively) (data not shown). Also among the B. pertussis genes showing elevated transcript abundance under iron starvation conditions were Bvg-dependent genes of the T3SS, which have been extensively analyzed in B. bronchiseptica (26,37,47,48,53,54,64,65,92,93). We examined the influence of iron starvation on Bordetella T3SS gene expression and protein production in greater detail (see below).…”

“…69 kDa in monomeric form). BteA is also thought to form large homomultimeric protein complexes or aggregates of greater than 200-kDa molecular mass (37,47,65). The extracellular protein (band 6) produced by the ⌬bvg mutant strain RB54 under iron-replete growth conditions was identified as flagellin (FlaA, 40.4 kDa; BB2539).…”

Differential regulation of type III secretion and virulence genes in Bordetella pertussis and Bordetella bronchiseptica by a secreted anti-σ factor

“…Remarkably, only a single effector, BteA, has been definitively identified as a translocated substrate (15,16). In B. bronchiseptica, BteA is necessary and sufficient for cytotoxicity in vitro, and mutations in bteA recapitulate phenotypes associated with deleting the T3SS ATPase gene, bscN (14)(15)(16).…”

“…In vitro, T3SS Bsc is a particularly conspicuous virulence determinant owing to its ability to induce rapid cytotoxicity in a diverse array of eukaryotic cells, ranging from mammalian to yeast (15,20). In vivo studies with B. bronchiseptica show that T3SS Bsc plays an immunomodulatory role that facilitates persistence in the lower respiratory tract (21,22).…”

“…BvgAS establishes a spectrum of phenotypic phases in response to environmental cues, functioning as a rheostat that controls the infectious cycle (7,8). The Bvg + phase is characterized by high level BvgAS activity, expression of virulence factors [including pertussis toxin (9), adenylate cyclase toxin (10,11), filamentous hemagglutinin (12), fimbriae (13), the Bordetella secretion complex (Bsc) type III secretion system (T3SS Bsc ) (14)(15)(16)], and the ability to survive and replicate on respiratory epithelia (17). In the Bvg phase, the BvgAS phosphorelay is quiescent, Bvg + phase genes are silent, and "virulence-repressed genes" are maximally transcribed (4,17,18).…”