Application of β-lactamase Reporter Fusions as an Indicator of Effector Protein Secretion During Infections With the Obligate Intracellular Pathogen Chlamydia Trachomatis

PLoS One. 2015 Aug 10;10(8):e0135295. doi: 10.1371/journal.pone.0135295. eCollection 2015.

Abstract

Chlamydia spp. utilize multiple secretion systems, including the type III secretion system (T3SS), to deploy host-interactive effector proteins into infected host cells. Elucidation of secreted proteins has traditionally required ectopic expression in a surrogate T3SS followed by immunolocalization of endogenous candidate effectors to confirm secretion by chlamydiae. The ability to transform Chlamydia and achieve stable expression of recombinant gene products has enabled a more direct assessment of secretion. We adapted TEM-1 β-lactamase as a reporter system for assessment of chlamydial protein secretion. We provide evidence that this system facilitates visualization of secretion in the context of infection. Specifically, our findings provide definitive evidence that C. trachomatis CT695 is secreted during infection. Follow-up indirect immunofluorescence studies confirmed CT695 secretion and indicate that this effector can be secreted at multiple points during the chlamydial developmental cycle. Our results indicate that the BlaM-fusion reporter assay will allow efficacious identification of novel secreted proteins. Moreover, this approach can easily be adapted to enable more sophisticated studies of the secretion process in Chlamydia.

Publication types

  • Research Support, N.I.H., Extramural

MeSH terms

  • Bacterial Proteins / genetics*
  • Bacterial Proteins / metabolism
  • Base Sequence
  • Chlamydia trachomatis / genetics*
  • Chlamydia trachomatis / metabolism
  • Chlamydia trachomatis / pathogenicity
  • Gene Expression Regulation, Bacterial*
  • Genes, Reporter
  • Green Fluorescent Proteins / genetics
  • Green Fluorescent Proteins / metabolism
  • HeLa Cells
  • Humans
  • Luminescent Proteins / genetics
  • Luminescent Proteins / metabolism
  • Molecular Sequence Data
  • Neisseria meningitidis / chemistry
  • Neisseria meningitidis / genetics
  • Plasmids / chemistry
  • Plasmids / metabolism
  • Promoter Regions, Genetic
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism
  • Transformation, Bacterial
  • Type III Secretion Systems / genetics*
  • Type III Secretion Systems / metabolism
  • beta-Lactamases / genetics*
  • beta-Lactamases / metabolism

Substances

  • Bacterial Proteins
  • Luminescent Proteins
  • Recombinant Fusion Proteins
  • Type III Secretion Systems
  • red fluorescent protein
  • Green Fluorescent Proteins
  • beta-Lactamases
  • beta-lactamase TEM-1