Dataset Information

The PAS Domain-Containing Protein HeuR Regulates Heme Uptake in Campylobacter jejuni.

ABSTRACT: Campylobacter jejuni is a leading cause of bacterially derived gastroenteritis. A previous mutant screen demonstrated that the heme uptake system (Chu) is required for full colonization of the chicken gastrointestinal tract. Subsequent work identified a PAS domain-containing regulator, termed HeuR, as being required for chicken colonization. Here we confirm that both the heme uptake system and HeuR are required for full chicken gastrointestinal tract colonization, with the heuR mutant being particularly affected during competition with wild-type C. jejuni Transcriptomic analysis identified the chu genes-and those encoding other iron uptake systems-as regulatory targets of HeuR. Purified HeuR bound the chuZA promoter region in electrophoretic mobility shift assays. Consistent with a role for HeuR in chu expression, heuR mutants were unable to efficiently use heme as a source of iron under iron-limiting conditions, and mutants exhibited decreased levels of cell-associated iron by mass spectrometry. Finally, we demonstrate that an heuR mutant of C. jejuni is resistant to hydrogen peroxide and that this resistance correlates to elevated levels of catalase activity. These results indicate that HeuR directly and positively regulates iron acquisition from heme and negatively impacts catalase activity by an as yet unidentified mechanism in C. jejuni IMPORTANCE: Annually, Campylobacter jejuni causes millions of gastrointestinal infections in the United States, due primarily to its ability to reside within the gastrointestinal tracts of poultry, where it can be released during processing and contaminate meat. In the developing world, humans are often infected by consuming contaminated water or by direct contact with livestock. Following consumption of contaminated food or water, humans develop disease that is characterized by mild to severe diarrhea. There is a need to understand both colonization of chickens, to make food safer, and colonization of humans, to better understand disease. Here we demonstrate that to efficiently colonize a host, C. jejuni requires iron from heme, which is regulated by the protein HeuR. Understanding how HeuR functions, we can develop ways to inhibit its function and reduce iron acquisition during colonization, potentially reducing C. jejuni in the avian host, which would make food safer, or limiting human colonization.

SUBMITTER: Johnson JG

PROVIDER: S-EPMC5111405 | biostudies-literature | 2016 Nov

REPOSITORIES: biostudies-literature

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The PAS Domain-Containing Protein HeuR Regulates Heme Uptake in Campylobacter jejuni.

Johnson Jeremiah G JG Gaddy Jennifer A JA DiRita Victor J VJ

mBio 20161115 6

Campylobacter jejuni is a leading cause of bacterially derived gastroenteritis. A previous mutant screen demonstrated that the heme uptake system (Chu) is required for full colonization of the chicken gastrointestinal tract. Subsequent work identified a PAS domain-containing regulator, termed HeuR, as being required for chicken colonization. Here we confirm that both the heme uptake system and HeuR are required for full chicken gastrointestinal tract colonization, with the heuR mutant being part ...[more]

PMID: 27935836

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Project description:Campylobacter jejuni is a major cause of bacterial foodborne illness in humans worldwide. Bacterial entry into a host eukaryotic cell involves the initial steps of adherence and invasion, which generally activate several cell-signaling pathways that induce the activation of innate defense systems, which leads to the release of proinflammatory cytokines and induction of apoptosis. Recent studies have reported that the unfolded protein response (UPR), a system to clear unfolded proteins from the endoplasmic reticulum (ER), also participates in the activation of cellular defense mechanisms in response to bacterial infection. However, no study has yet investigated the role of UPR in C. jejuni infection. Hence, the aim of this study was to deduce the role of UPR signaling via induction of ER stress in the process of C. jejuni infection. The results suggest that C. jejuni infection suppresses global protein translation. Also, 12 h of C. jejuni infection induced activation of the eIF2α pathway and expression of the transcription factor CHOP. Interestingly, bacterial invasion was facilitated by knockdown of UPR-associated signaling factors and treatment with the ER stress inducers, thapsigargin and tunicamycin, decreased the invasive ability of C. jejuni. An investigation into the mechanism of UPR-mediated inhibition of C. jejuni invasion showed that UPR signaling did not affect bacterial adhesion to or survival in the host cells. Further, Salmonella Enteritidis or FITC-dextran intake were not regulated by UPR signaling. These results indicated that the effect of UPR on intracellular intake was specifically found in C. jejuni infection. These findings are the first to describe the role of UPR in C. jejuni infection and revealed the participation of a new signaling pathway in C. jejuni invasion. UPR signaling is involved in defense against the early step of C. jejuni invasion and thus presents a potential therapeutic target for the treatment of C. jejuni infection.

Project description:Periodontal disease (PD) results from a shift in the composition of the microbial community of the subgingival crevice. As the bacterial population transitions from Gram-positive bacteria to predominantly Gram-negative anaerobes and spirochetes, dramatic changes occur in the physiological and immunological environment at diseased sites. Treponema denticola thrives in periodontal pockets, indicating that it has a unique ability to adapt to changing environmental conditions. Hpk2 (tde1970), a Per-Arnt-Sim motif (PAS) domain-containing histidine kinase (HK), is part of the T. denticola Hpk2-Rrp2 (tde1969) two-component regulatory (TCR) system. This TCR system is growth phase regulated and has been postulated to play a key role in adaptive responses. In this study, we employ predictive structural analyses and site-directed mutagenesis to investigate the functional role of specific amino acid residues located within the Hpk2 PAS domain. Specific substitutions impacted autophosphorylation (AP), phosphotransfer (PT), oligomerization, and hemin binding. The AP, PT, hemin binding, and oligomerization potential of some mutated Hpk2 proteins differed under aerobic versus anaerobic reaction conditions. The data presented here suggest that the regulatory activity of Hpk2 is linked to diatomic gas levels. In a broader sense, this study highlights the importance of studying proteins produced by anaerobes under conditions that approximate the environment in which they thrive.IMPORTANCE Periodontal disease affects nearly 60% of the global adult population. Its costs to individuals, and to society as a whole, are enormous. As periodontal disease develops, there is a shift in the composition of the oral microbial community. The bacteria that become dominant are able to cause significant damage to the tissues that support the teeth, leading to tooth loss. Treponema denticola is one of the keystone pathogens associated with periodontal disease. An earlier study demonstrated that the Hpk2 and Rrp2 proteins play an important role in adaptive responses. Here, we explore the role of specific Hpk2 amino acids in environmental sensing and function, using structural analyses and site-directed mutagenesis.

Dataset Information

The PAS Domain-Containing Protein HeuR Regulates Heme Uptake in Campylobacter jejuni.

Publications

The PAS Domain-Containing Protein HeuR Regulates Heme Uptake in Campylobacter jejuni.

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