Identification of T-cell epitopes in Francisella tularensis using an ordered protein array of serological targets

Michael D. Valentino, Zachary J. Maben, Lucinda L. Hensley, Matthew D. Woolard, Thomas H. Kawula, Jeffrey A Frelinger, John G. Frelinger

Research output: Contribution to journalArticle

18 Citations (Scopus)

Abstract

Francisella tularensis is a Gram-negative intracellular bacterium that is the causative agent of tularaemia. Concerns regarding its use as a bioterrorism agent have led to a renewed interest in the biology of infection, host response and pathogenesis. A robust T-cell response is critical to confer protection against F. tularensis. However, characterization of the cellular immune response has been hindered by the paucity of tools to examine the anti-Francisella immune response at the molecular level. We set out to combine recent advances of genomics with solid-phase antigen delivery coupled with a T-cell functional assay to identify T-cell epitopes. A subset of clones, encoding serological targets, was selected from an F. tularensis SchuS4 ordered genomic library and subcloned into a bacterial expression vector to test the feasibility of this approach. Proteins were expressed and purified individually employing the BioRobot 3000 in a semi-automated purification method. The purified proteins were coupled to beads, delivered to antigen-presenting cells for processing, and screened with Francisella-specific T-cell hybridomas of unknown specificity. We identified cellular reactivity against the pathogenicity protein IglB, and the chaperone proteins GroEL and DnaK. Further analyses using genetic deletions and synthetic peptides were performed to identify the minimal peptide epitopes. Priming with the peptide epitopes before infection with F. tularensis LVS increased the frequency of antigen-specific CD4 T cells as assessed by intracellular interferon-γ staining. These results illustrate the feasibility of screening an arrayed protein library that should be applicable to a variety of pathogens.

Original languageEnglish (US)
Pages (from-to)348-360
Number of pages13
JournalImmunology
Volume132
Issue number3
DOIs
StatePublished - Mar 2011

Fingerprint

Francisella tularensis
Protein Array Analysis
T-Lymphocyte Epitopes
Francisella
Tularemia
T-Lymphocytes
Peptides
Epitopes
Proteins
Biological Warfare Agents
Chaperonin 60
CD4 Antigens
Genomic Library
Hybridomas
Antigen-Presenting Cells
Genomics
Gram-Negative Bacteria
Cellular Immunity
Interferons
Virulence

Keywords

  • Epitope discovery
  • Francisella tularensis
  • Gateway genomic array
  • Proteomic array
  • T lymphocyte

ASJC Scopus subject areas

  • Immunology
  • Immunology and Allergy

Cite this

Valentino, M. D., Maben, Z. J., Hensley, L. L., Woolard, M. D., Kawula, T. H., Frelinger, J. A., & Frelinger, J. G. (2011). Identification of T-cell epitopes in Francisella tularensis using an ordered protein array of serological targets. Immunology, 132(3), 348-360. https://doi.org/10.1111/j.1365-2567.2010.03387.x

Identification of T-cell epitopes in Francisella tularensis using an ordered protein array of serological targets. / Valentino, Michael D.; Maben, Zachary J.; Hensley, Lucinda L.; Woolard, Matthew D.; Kawula, Thomas H.; Frelinger, Jeffrey A; Frelinger, John G.

In: Immunology, Vol. 132, No. 3, 03.2011, p. 348-360.

Research output: Contribution to journalArticle

Valentino, Michael D. ; Maben, Zachary J. ; Hensley, Lucinda L. ; Woolard, Matthew D. ; Kawula, Thomas H. ; Frelinger, Jeffrey A ; Frelinger, John G. / Identification of T-cell epitopes in Francisella tularensis using an ordered protein array of serological targets. In: Immunology. 2011 ; Vol. 132, No. 3. pp. 348-360.
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