Biology subjects

Nash, Z. M.

Publications and source records attributed to Nash, Z. M..

3 recordsLinked to original sources

bioRxiv2024

Cytochrome oxidase requirements in Bordetella reveal insights into evolution towards life in the mammalian respiratory tract

Little is known about oxygen utilization during infection by bacterial respiratory pathogens. The classical Bordetella species, including B. pertussis, the causal agent of human whooping cough, and B. bronchiseptica, which infects nearly all mammals, are obligate aerobes that use only oxygen as the terminal electron acceptor for electron transport-coupled oxidative phosphorylation. B. bronchiseptica, which occupies many niches, has eight distinct cytochrome oxidase-encoding loci, while B. pertussis, which evolved from a B. bronchiseptica-like ancestor but now only survives only in and between human respiratory tracts, has only three functional cytochrome oxidase-encoding loci: cydAB1, ctaCDFGE1, and cyoABCD1. To test the hypothesis that the three cytochrome oxidases encoded within the B. pertussis genome represent the minimum number and class of cytochrome oxidase required for respiratory infection, we compared B. bronchiseptica strains lacking one or more of the eight possible cytochrome oxidases in vitro and in vivo. No individual cytochrome oxidase was required for growth in ambient air, and all three of the cytochrome oxidases conserved in B. pertussis were sufficient for growth in ambient air and low oxygen. Using a high-dose, large-volume persistence model and a low-dose, small-volume establishment of infection model, we found that B. bronchiseptica producing only the three B. pertussis-conserved cytochrome oxidases was indistinguishable from the wild-type strain for infection. We also showed that CyoABCD1 is sufficient to cause the same level of bacterial burden in mice as the wild-type strain and is thus the primary cytochrome oxidase required for murine infection, and that CydAB1 and CtaCDFGE1 fulfill auxiliary roles or are important for aspects of infection we have not assessed, such as transmission. Our results shed light on respiration requirements for bacteria that colonize the respiratory tract, the environment at the surface of the ciliated epithelium, and the evolution of virulence in bacterial pathogens. AUTHOR SUMMARYCytochrome oxidases, critical components for aerobic respiration, have been shown to be vital for pathogenesis and tissue tropism in several bacterial species. However, the majority of the research has focused on facultative anaerobes and infections of microoxic to anaerobic host environments, like the gut. We sought to understand the role of cytochrome oxidases during respiratory infection by Bordetella bronchiseptica, an obligate aerobe, performing the first analysis of cytochrome oxidases in an extracellular respiratory pathogen that we know of. By comparing B. bronchiseptica to the closely related B. pertussis, a strictly human-specific pathogen and the causative agent of whooping cough, we found three cytochrome oxidases that are important for growth and survival within the mammalian respiratory tract. We also found that a bo3-type cytochrome oxidase, predicted to have a low affinity for oxygen and therefore best suited to ambient air levels of oxygen, was sufficient for both the establishment of infection and persistence in the respiratory tract in mice. Our findings reveal the importance of low affinity cytochrome oxidases in respiratory pathogens, and emphasize the need to study the physiology of diverse pathogens.

microbiology↗

bioRxiv2023

Bordetella Filamentous hemagglutinin (FhaB) and Adenylyl cyclase toxin (ACT) interactions on the bacterial surface are consistent with FhaB-mediated delivery of ACT to phagocytic cells

Bordetella species that cause respiratory infections in mammals include B. pertussis, which causes human whooping cough, and B. bronchiseptica, which infects nearly all mammals. Both bacterial species produce filamentous hemagglutinin (FhaB) and adenylyl cyclase toxin (ACT), prominent surface-associated and secreted virulence factors that contribute to persistence in the lower respiratory tract by inhibiting clearance by phagocytic cells. FhaB and ACT proteins interact with themselves, each other, and host cells. Using immunoblot analyses, we showed that ACT binds to FhaB preferentially on the bacterial surface before being released into the extracellular environment. We showed that SphB1, a surface protease identified based on its ability to cleave FhaB, also cleaves ACT, and we showed that the presence of ACT blocks SphB1-dependent and independent cleavage of FhaB, but the presence of FhaB does not block SphB1-dependent cleavage of ACT. SphB1-dependent cleavage of ACT occurs proximally to ACTs active site, and hence is predicted to inactivate ACT. We also showed that FhaB-bound ACT can intoxicate host cells producing CR3, the receptor for ACT. In addition to increasing our understanding of FhaB, ACT, and ACT interactions on the Bordetella surface, our data support a model in which FhaB functions as a novel toxin delivery system by binding to ACT and allowing its release upon binding of ACT to its receptor, CR3, on phagocytic cells. AUTHOR SUMMARYBacteria need to control the variety, abundance, and conformation of proteins on the cellular surface to survive. Members of the Gram-negative bacterial genus Bordetella include B. pertussis, which causes whooping cough in humans, and B. bronchiseptica, which causes respiratory infections in a broad range of mammals. These extremely closely related species produce two prominent virulence factors, the two-partner secretion (TPS) effector FhaB and Adenylyl cyclase toxin (ACT), that interact with themselves, each other, and host cells. In this work, we showed that ACT binds preferentially to FhaB on the bacterial surface before being released into the extracellular environment. We showed that the exoprotease SphB1, which cleaves FhaB, also cleaves ACT. We showed that ACT that is bound to FhaB can be delivered to CR3+ host cells. Our data support a model in which FhaB functions to deliver ACT specifically to phagocytic cells, and not epithelial cells. This is the first report of a TPS system facilitating delivery of a separate polypeptide toxin to target cells and expands our understanding of how these systems contribute to bacterial pathogenesis.

microbiology↗

bioRxiv2021

DegP initiates regulated processing of filamentous hemagglutinin in Bordetella bronchiseptica

Filamentous hemagglutinin (FhaB) is a critical virulence factor for both Bordetella pertussis, the causal agent of whooping cough, and the closely related species Bordetella bronchiseptica. FhaB is an adhesin, suppresses inflammatory cytokine production, and protects against phagocytic cell clearance during infection. Regulated degradation of the FhaB C-terminal prodomain is required to establish a persistent infection in mice. Two proteases, CtpA in the periplasm and SphB1 on the bacterial surface, are known to mediate FhaB processing, and we recently determined that CtpA functions before, and controls the FhaB cleavage site of, SphB1. However, the data indicate that another periplasmic protease must initiate degradation of the prodomain by removing a portion of the FhaB C terminus that inhibits CtpA-mediated degradation. Using a candidate approach, we identified DegP as the initiating protease. Deletion of degP or substitution of its predicted catalytic residue resulted in reduced creation of FHA', the main product of FhaB processing, and an accumulation of full-length FhaB in whole cell lysates. Also, FHA' was no longer released into culture supernatants in degP mutants. Alterations of the FhaB C terminus that relieve inhibition of CtpA abrogate the need for DegP, consistent with DegP functioning prior to CtpA in the processing pathway. DegP is not required for secretion of FhaB through FhaC or for adherence of the bacteria to host cells, indicating that DegP acts primarily as a protease and not a chaperone for FhaB in B. bronchiseptica. Our results highlight a role for HtrA family proteases in activation of virulence factors in pathogenic bacteria. ImportanceTwo-partner secretion (TPS) systems are broadly distributed among Gram-negative bacteria and play important roles in bacterial pathogenesis. FhaB-FhaC is the prototypical member of the TPS family and here we identified the protease that initiates a processing cascade that controls FhaB function. Our results are significant because they provide insight into the molecular mechanism underlying the ability of Bordetella species to prevent clearance by phagocytic cells, which is critical for bacterial persistence in the lower respiratory tract. Our findings also highlight an underappreciated role for HtrA family proteases in processing specific bacterial virulence factors.

microbiology↗