The immune system, by and large, plays a primary role to minimize and/or prevent infection, in general, and periodontal disease, in particular. We propose that immunoregulatory abnormalities contribute to the pathogenesis of and susceptibility to periodontal disease. The fundamental hypothesis of our studies is that periodontal pathogens produce immunotoxins that mediate local and/or systemic immunosuppression, thereby enhancing their own virulence and/or that of other opportunistic organisms. In this regard, our investigations have demonstrated that Aggregatibacter actinomycetemcomitans produces the cytolethal distending toxin (Cdt) which is capable of impairing human T- and B-cell function by inducing cell cycle arrest and apoptosis. The studies described in this proposal will elucidate the molecular events associated with Cdt activity and, therefore, the pathogenesis of disease resulting from Cdt-producing bacteria. We have advanced the hypothesis that the mode of action of the Cdt active subunit, CdtB, involves phosphatidylinositol-3,4,5-triphosphate (PIP3) depletion and inactivation of the PI-3K/PIP3/Akt signaling pathway leading to the activation of downstream targets and conditions that favor growth arrest and apoptosis. Furthermore, it is our hypothesis that G2 arrest is linked to CdtB's PIP3 phosphatase activity and the convergence of three synergistic downstream events: FOXO3 activation leading to expression of cyclin G1 and/or G2, activation of Chk1 and the G2 checkpoint and accumulation of PI-3,4-P2 and localization of the adaptor protein, TAPP1. Our studies have also established that in order for CdtB to perturb PIP3 metabolism, the toxin binds to cells via the CdtC subunit which recognizes cholesterol in the context of lipid rafts. More recently, we have determined the CdtB contains a cholesterol recognition sequence. Therefore, we further propose that cellular entry of CdtB and ultimately, localization to PIP3 pools is dependent upon association with cholesterol-rich lipid rafts and cellugyrin. The objectives of this proposal are to extend our observations to (1) determine the downstream events triggered by CdtB-mediated PIP3 depletion which result in G2 arrest and activation of the apoptotic cascade and (2) determine the role of cholesterol and lipid rafts in CdtB-mediated PIP3 depletion and toxicity. The study is divided into three specific aims: (1) to determine the mechanism by which CdtB-mediated PIP3 depletion leads to activation of the G2 checkpoint and cell cycle arrest, (2) to determine if CdtB-induced apoptosis results from activation of the intrinsic apoptotic pathway mediated by FOXO3a activation, and (3) to determine the role of cholesterol in CdtB mediated PIP3 depletion leading to cell cycle arrest and apoptosis. It is anticipated that these studies will lead to a more detailed understanding of Cdt and provide: (1) important insight into the pathogenesis of disease caused by Cdt-producing bacteria and (2) a rationale on which therapeutic intervention may be developed to prevent or limit disease.

Public Health Relevance

The fundamental goal of the proposed study is to determine the underlying mechanism by which a bacterial derived toxin, the cytolethal distending toxin (Cdt), acts to inhibit cell growth and induce cell death and thereby perturb host defenses. It is anticipated that these studies will lead to a more detailed understanding of the toxin and provide: (1) important insight into the pathogenesis of disease caused by Cdt-producing bacteria and (2) a rationale on which therapeutic intervention may be developed to prevent or limit disease

Agency
National Institute of Health (NIH)
Institute
National Institute of Dental & Craniofacial Research (NIDCR)
Type
Research Project (R01)
Project #
5R01DE006014-30
Application #
8293022
Study Section
Special Emphasis Panel (ZRG1-MOSS-K (02))
Program Officer
Lunsford, Dwayne
Project Start
1982-08-01
Project End
2016-06-30
Budget Start
2012-07-01
Budget End
2013-06-30
Support Year
30
Fiscal Year
2012
Total Cost
$400,000
Indirect Cost
$150,000
Name
University of Pennsylvania
Department
Pathology
Type
Schools of Dentistry
DUNS #
042250712
City
Philadelphia
State
PA
Country
United States
Zip Code
19104
Boesze-Battaglia, Kathleen; Walker, Lisa P; Dhingra, Anuradha et al. (2017) Internalization of the Active Subunit of the Aggregatibacter actinomycetemcomitans Cytolethal Distending Toxin Is Dependent upon Cellugyrin (Synaptogyrin 2), a Host Cell Non-Neuronal Paralog of the Synaptic Vesicle Protein, Synaptogyrin 1. Front Cell Infect Microbiol 7:469
Leonard, Stephanie A; Petito, Lucia C; Stephansson, Olof et al. (2017) Weight gain during pregnancy and the black-white disparity in preterm birth. Ann Epidemiol 27:323-328.e1
Scuron, Monika D; Boesze-Battaglia, Kathleen; Dlaki?, Mensur et al. (2016) The Cytolethal Distending Toxin Contributes to Microbial Virulence and Disease Pathogenesis by Acting As a Tri-Perditious Toxin. Front Cell Infect Microbiol 6:168
Shenker, B J; Walker, L P; Zekavat, A et al. (2016) Lymphoid susceptibility to the Aggregatibacter actinomycetemcomitans cytolethal distending toxin is dependent upon baseline levels of the signaling lipid, phosphatidylinositol-3,4,5-triphosphate. Mol Oral Microbiol 31:33-42
Shenker, Bruce J; Boesze-Battaglia, Kathleen; Scuron, Monika Damek et al. (2016) The toxicity of the Aggregatibacter actinomycetemcomitans cytolethal distending toxin correlates with its phosphatidylinositol-3,4,5-triphosphate phosphatase activity. Cell Microbiol 18:223-43
Boesze-Battaglia, Kathleen; Walker, Lisa P; Zekavat, Ali et al. (2015) The Aggregatibacter actinomycetemcomitans Cytolethal Distending Toxin Active Subunit CdtB Contains a Cholesterol Recognition Sequence Required for Toxin Binding and Subunit Internalization. Infect Immun 83:4042-55
Shenker, Bruce J; Ojcius, David M; Walker, Lisa P et al. (2015) Aggregatibacter actinomycetemcomitans cytolethal distending toxin activates the NLRP3 inflammasome in human macrophages, leading to the release of proinflammatory cytokines. Infect Immun 83:1487-96
Shenker, Bruce J; Walker, Lisa P; Zekavat, Ali et al. (2014) Blockade of the PI-3K signalling pathway by the Aggregatibacter actinomycetemcomitans cytolethal distending toxin induces macrophages to synthesize and secrete pro-inflammatory cytokines. Cell Microbiol 16:1391-404
Shenker, Bruce J; Ali, Hydar; Boesze-Battaglia, Kathleen (2011) PIP3 regulation as promising targeted therapy of mast-cell-mediated diseases. Curr Pharm Des 17:3815-22
Shenker, Bruce J; Boesze-Battaglia, Kathleen; Zekavat, Ali et al. (2010) Inhibition of mast cell degranulation by a chimeric toxin containing a novel phosphatidylinositol-3,4,5-triphosphate phosphatase. Mol Immunol 48:203-10