Aggregatibacter actinomycetemcomitans outer membrane protein 29 (Omp29) induces TGF-β-regulated apoptosis signal in human gingival epithelial cells via fibronectin/integrinβ1/FAK cascade

Cell Microbiol. 2016 Dec;18(12):1723-1738. doi: 10.1111/cmi.12607. Epub 2016 Jun 10.

Abstract

Gingival junctional epithelial cell apoptosis caused by periodontopathic bacteria exacerbates periodontitis. This pathological apoptosis is involved in the activation of transforming growth factor β (TGF-β). However, the molecular mechanisms by which microbes induce the activation of TGF-β remain unclear. We previously reported that Aggregatibacter actinomycetemcomitans (Aa) activated TGF-β receptor (TGF-βR)/smad2 signalling to induce epithelial cell apoptosis, even though Aa cannot bind to TGF-βR. Additionally, outer membrane protein 29 kDa (Omp29), a member of the Aa Omps family, can induce actin rearrangements via focal adhesion kinase (FAK) signalling, which also plays a role in the activation of TGF-β by cooperating with integrin. Accordingly, we hypothesized that Omp29-induced actin rearrangements via FAK activity would enhance the activation of TGF-β, leading to gingival epithelial cell apoptosis in vitro. By using human gingival epithelial cell line OBA9, we found that Omp29 activated TGF-βR/smad2 signalling and decreased active TGF-β protein levels in the extracellular matrix (ECM) of cell culture, suggesting the transactivation of TGF-βR. Inhibition of actin rearrangements by cytochalasin D or blebbistatin and knockdown of FAK or integrinβ1 expression by siRNA transfection attenuated TGF-βR/smad2 signalling activity and reduction of TGF-β levels in the ECM caused by Omp29. Furthermore, Omp29 bound to fibronectin (Fn) to induce its aggregation on integrinβ1, which is associated with TGF-β signalling activity. All the chemical inhibitors and siRNAs tested blocked Omp29-induced OBA9 cells apoptosis. These results suggest that Omp29 binds to Fn in order to facilitate Fn/integrinβ1/FAK signalling-dependent TGF-β release from the ECM, thereby inducing gingival epithelial cell apoptosis via TGF-βR/smad2 pathway.

MeSH terms

  • Aggregatibacter actinomycetemcomitans / genetics*
  • Aggregatibacter actinomycetemcomitans / metabolism
  • Apoptosis / genetics
  • Bacterial Outer Membrane Proteins / genetics*
  • Bacterial Outer Membrane Proteins / metabolism
  • Bacterial Outer Membrane Proteins / pharmacology
  • Cell Line, Transformed
  • Cytochalasin D / pharmacology
  • Epithelial Cells / drug effects
  • Epithelial Cells / metabolism
  • Epithelial Cells / microbiology*
  • Fibronectins / genetics*
  • Fibronectins / metabolism
  • Focal Adhesion Kinase 1 / genetics*
  • Focal Adhesion Kinase 1 / metabolism
  • Gene Expression Regulation
  • Gingiva / metabolism
  • Gingiva / microbiology
  • Heterocyclic Compounds, 4 or More Rings / pharmacology
  • Host-Pathogen Interactions
  • Humans
  • Integrin beta1 / genetics*
  • Integrin beta1 / metabolism
  • Protein Serine-Threonine Kinases / genetics
  • Protein Serine-Threonine Kinases / metabolism
  • RNA, Small Interfering / genetics
  • RNA, Small Interfering / metabolism
  • Receptor, Transforming Growth Factor-beta Type I
  • Receptors, Transforming Growth Factor beta / genetics
  • Receptors, Transforming Growth Factor beta / metabolism
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Recombinant Proteins / pharmacology
  • Signal Transduction
  • Smad2 Protein / antagonists & inhibitors
  • Smad2 Protein / genetics
  • Smad2 Protein / metabolism
  • Transforming Growth Factor beta / antagonists & inhibitors
  • Transforming Growth Factor beta / genetics*
  • Transforming Growth Factor beta / metabolism

Substances

  • Bacterial Outer Membrane Proteins
  • Fibronectins
  • Heterocyclic Compounds, 4 or More Rings
  • Integrin beta1
  • Omp29 protein, Actinobacillus actinomycetemcomitans
  • RNA, Small Interfering
  • Receptors, Transforming Growth Factor beta
  • Recombinant Proteins
  • SMAD2 protein, human
  • Smad2 Protein
  • Transforming Growth Factor beta
  • blebbistatin
  • Cytochalasin D
  • Focal Adhesion Kinase 1
  • PTK2 protein, human
  • Protein Serine-Threonine Kinases
  • Receptor, Transforming Growth Factor-beta Type I