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Title: Identification and characterization of a nontypeable Haemophilus influenzae putative toxin-antitoxin locus

Abstract

Background: Certain strains of an obligate parasite of the human upper respiratory tract, nontypeable Haemophilus influenzae (NTHi), can cause invasive diseases such as septicemia and meningitis, as well as chronic mucosal infections such as otitis media. To do this, the organism must invade and survive within both epithelial and endothelial cells. We have identified a facilitator of NTHi survival inside human cells, virulence-associated protein D (vapDHi, encoded by gene HI0450). Both vapDHi and a flanking gene, HI0451, exhibit the genetic and physical characteristics of a toxin/antitoxin (TA) locus, with VapDHi serving as the toxin moiety and HI0451 as the antitoxin. We propose the name VapXHi for the HI0451 antitoxin protein. Originally identified on plasmids, TA loci have been found on the chromosomes of a number of bacterial pathogens, and have been implicated in the control of translation during stressful conditions. Translation arrest would enhance survival within human cells and facilitate persistent or chronic mucosal infections. Results: Isogenic mutants in vapDHi were attenuated for survival inside human respiratory epithelial cells (NCI-H292) and human brain microvascular endothelial cells (HBMEC), the in vitro models of mucosal infection and the blood-brain barrier, respectively. Transcomplementation with a vapDHi allele restored wild-type NTHi survival within bothmore » cell lines. A PCR survey of 59 H. influenzae strains isolated from various anatomical sites determined the presence of a vapDHiallele in 100% of strains. Two isoforms of the gene were identified in this population; one that was 91 residues in length, and another that was truncated to 45 amino acids due to an in-frame deletion. The truncated allele failed to transcomplement the NTHi vapDHi survival defect in HBMEC. Subunits of full-length VapDHi homodimerized, but subunits of the truncated protein did not. However, truncated protein subunits did interact with full-length subunits, and this interaction resulted in a dominant-negative phenotype. Although Escherichia coli does not contain a homologue of either vapDHi or vapXHi, overexpression of the VapDHi toxin in trans resulted in E. coli cell growth arrest. This arrest could be rescued by providing the VapXHi antitoxin on a compatible plasmid. Conclusion: We conclude that vapDHi and vapXHi may constitute a H. influenzae TA locus that functions to enhance NTHi survival within human epithelial and endothelial cells.« less

Authors:
 [1];  [2];  [2]
  1. Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States). International Security Directorate. Arms Control. Nonproliferation
  2. Seattle Biomedical Research Inst., Seattle, WA (United States). Bacterial Pathogenesis Program
Publication Date:
Research Org.:
Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States)
Sponsoring Org.:
USDOE Office of Science (SC), Biological and Environmental Research (BER). Biological Systems Science Division; National Institutes of Health (NIH)
OSTI Identifier:
1626485
Grant/Contract Number:  
AC52-07NA27344; AI 07276; AI 44002
Resource Type:
Journal Article: Accepted Manuscript
Journal Name:
BMC Microbiology
Additional Journal Information:
Journal Volume: 4; Journal Issue: 1; Journal ID: ISSN 1471-2180
Publisher:
BioMed Central
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; Microbiology

Citation Formats

Daines, Dayle A., Jarisch, Justin, and Smith, Arnold L. Identification and characterization of a nontypeable Haemophilus influenzae putative toxin-antitoxin locus. United States: N. p., 2004. Web. doi:10.1186/1471-2180-4-30.
Daines, Dayle A., Jarisch, Justin, & Smith, Arnold L. Identification and characterization of a nontypeable Haemophilus influenzae putative toxin-antitoxin locus. United States. https://doi.org/10.1186/1471-2180-4-30
Daines, Dayle A., Jarisch, Justin, and Smith, Arnold L. 2004. "Identification and characterization of a nontypeable Haemophilus influenzae putative toxin-antitoxin locus". United States. https://doi.org/10.1186/1471-2180-4-30. https://www.osti.gov/servlets/purl/1626485.
@article{osti_1626485,
title = {Identification and characterization of a nontypeable Haemophilus influenzae putative toxin-antitoxin locus},
author = {Daines, Dayle A. and Jarisch, Justin and Smith, Arnold L.},
abstractNote = {Background: Certain strains of an obligate parasite of the human upper respiratory tract, nontypeable Haemophilus influenzae (NTHi), can cause invasive diseases such as septicemia and meningitis, as well as chronic mucosal infections such as otitis media. To do this, the organism must invade and survive within both epithelial and endothelial cells. We have identified a facilitator of NTHi survival inside human cells, virulence-associated protein D (vapDHi, encoded by gene HI0450). Both vapDHi and a flanking gene, HI0451, exhibit the genetic and physical characteristics of a toxin/antitoxin (TA) locus, with VapDHi serving as the toxin moiety and HI0451 as the antitoxin. We propose the name VapXHi for the HI0451 antitoxin protein. Originally identified on plasmids, TA loci have been found on the chromosomes of a number of bacterial pathogens, and have been implicated in the control of translation during stressful conditions. Translation arrest would enhance survival within human cells and facilitate persistent or chronic mucosal infections. Results: Isogenic mutants in vapDHi were attenuated for survival inside human respiratory epithelial cells (NCI-H292) and human brain microvascular endothelial cells (HBMEC), the in vitro models of mucosal infection and the blood-brain barrier, respectively. Transcomplementation with a vapDHi allele restored wild-type NTHi survival within both cell lines. A PCR survey of 59 H. influenzae strains isolated from various anatomical sites determined the presence of a vapDHiallele in 100% of strains. Two isoforms of the gene were identified in this population; one that was 91 residues in length, and another that was truncated to 45 amino acids due to an in-frame deletion. The truncated allele failed to transcomplement the NTHi vapDHi survival defect in HBMEC. Subunits of full-length VapDHi homodimerized, but subunits of the truncated protein did not. However, truncated protein subunits did interact with full-length subunits, and this interaction resulted in a dominant-negative phenotype. Although Escherichia coli does not contain a homologue of either vapDHi or vapXHi, overexpression of the VapDHi toxin in trans resulted in E. coli cell growth arrest. This arrest could be rescued by providing the VapXHi antitoxin on a compatible plasmid. Conclusion: We conclude that vapDHi and vapXHi may constitute a H. influenzae TA locus that functions to enhance NTHi survival within human epithelial and endothelial cells.},
doi = {10.1186/1471-2180-4-30},
url = {https://www.osti.gov/biblio/1626485}, journal = {BMC Microbiology},
issn = {1471-2180},
number = 1,
volume = 4,
place = {United States},
year = {Mon Jul 26 00:00:00 EDT 2004},
month = {Mon Jul 26 00:00:00 EDT 2004}
}

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