Abstract
The whooping cough agent, Bordetella pertussis, secretes an adenylate cyclase toxin–hemolysin (CyaA, ACT, or AC-Hly) that catalyzes the conversion of intracellular ATP to cAMP and through its signaling annihilates the bactericidal activities of host sentinel phagocytes. In parallel, CyaA permeabilizes host cells by the formation of cation-selective membrane pores that account for the hemolytic activity of CyaA. The pore-forming activity contributes to the overall cytotoxic effect of CyaA in vitro, and it has previously been proposed to synergize with the cAMP-elevating activity in conferring full virulence on B. pertussis in the mouse model of pneumonic infection. CyaA primarily targets myeloid phagocytes through binding of their complement receptor 3 (CR3, integrin αMβ2, or CD11b/CD18). However, with a reduced efficacy, the toxin can promiscuously penetrate and permeabilize the cell membrane of a variety of non-myeloid cells that lack CR3 on the cell surface, including airway epithelial cells or erythrocytes, and detectably intoxicates them by cAMP. Here, we used CyaA variants with strongly and selectively enhanced or reduced pore-forming activity that, at the same time, exhibited a full capacity to elevate cAMP concentrations in both CR3-expressing and CR3-non-expressing target cells. Using B. pertussis mutants secreting such CyaA variants, we show that a selective enhancement of the cell-permeabilizing activity of CyaA does not increase the overall virulence and lethality of pneumonic B. pertussis infection of mice any further. In turn, a reduction of the cell-permeabilizing activity of CyaA did not reduce B. pertussis virulence any importantly. These results suggest that the phagocyte-paralyzing cAMP-elevating capacity of CyaA prevails over the cell-permeabilizing activity of CyaA that appears to play an auxiliary role in the biological activity of the CyaA toxin in the course of B. pertussis infections in vivo.
Highlights
The Gram-negative coccobacillus Bordetella pertussis causes a highly contagious respiratory disease called pertussis, or whooping cough
In B. pertussis, proCyaA is posttranslationally activated by CyaC-catalyzed acylation of the K860 and K983 residues by the saturated palmitoyl (C16:0) chains [17,18]
This indicated that the pore-forming activity of CyaA and/or its capacity to elevate cAMP in complement receptor 3 (CR3)− cells was required for full virulence of B. pertussis [42]
Summary
The Gram-negative coccobacillus Bordetella pertussis causes a highly contagious respiratory disease called pertussis, or whooping cough. CyaA is a 1706 residues-long polypeptide consisting of an N-terminal adenylate cyclase (AC) enzyme domain (~384 residues) and a pore-forming RTX (Repeats in ToXin) hemolysin (Hly) moiety (~1322 residues) [8,10,11,12]. The Hly comprises (i) a hydrophobic pore-forming domain consisting of residues 500 to 700 [13,14,15,16]; (ii) a fatty acyl-modified segment located between residues 800 to 1000, where the CyaC-mediated activation of proCyaA to CyaA takes place by covalent post-translational acylation of ε-amino groups of lysine residues 860 (K860) and 983 (K983) [17,18]; (iii) a typical calcium-binding RTX domain, harboring the conserved nonapeptide repeats of a consensus sequence X-(L/I/F)-X-G-G-X-G-(N/D)D, which forms ~40 calcium-binding sites [19], and (iv) a carboxy-proximal secretion signal [11,20,21]
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