This project will examine global protein expression changes in oral microbial communities and the community response to model host cells. We will select as model organisms the periodontal pathogen P. gingivalis, along with F. nucleatum and S. gordonii as representative of organism commonly found in dental biofilms and capable of synergistic interactions with P. gingivalis. In addition to differential protein expression as these organisms assemble into communities, we will also determine changes in protein expression induced by contact with gingival epithelial cells. The long-term goals of the study involve improving our fundamental understanding at the molecular level of events surrounding heterotypic oral biofilm formation and the interaction of these biofilms with host cells.
Specific Aim 1 defines reference proteomes of P. gingivalis, S. gordonii, and F. nucleatum individually, pairwise and all together. This will include global protein modification analysis using data mining techniques, and a chemical approach to global phosphorylation specifically. We will define protein relative abundance changes (relative to the single organism in isolation) that can be attributed to community microbial interactions. We will thus establish the proteome of the P. gingivalis-F. nucleatum-S. gordonii community, and the differential responses to community development in comparison to individual proteomes.
Specific Aim 2 will define the effects of gingival epithelial cells on the P. gingivalis-F. nucleatum-S. gordonii community. This will involve running the same experiments as in Aim 1, but in the presence of epithelial cells, also with comparative posttranslational modification (PTM) analysis on the microbial side. This will define the response of the community to the presence of host cells. As in Aim 1, this includes mining the datasets globally for PTMs for all known modifications using computational approaches, and in addition using a chemically based experimental approach to measuring differential global phosphorylation.
In Aim 3 we will prioritize the regulated proteins identified in Aims 1 and 2, as discovery warrants, for additional corroboration and for construction of mutants in the corresponding genes. The phenotype of the mutants in community development and responses to epithelial cells will further elucidate functional meaning and biological relevance.

Public Health Relevance

Periodontal, or gum, diseases are the major cause of tooth loss in developed countries and result from the concerted activities of groups of microorganisms. These organisms adjust expression of their proteins as they assemble into communities and interact with host cells. We will define these protein expression changes and assess their importance for contribution to the disease process. Gaining an improved understanding of protein changes is a basic requirement for improved therapeutic and diagnostic strategies.

Agency
National Institute of Health (NIH)
Institute
National Institute of Dental & Craniofacial Research (NIDCR)
Type
Research Project (R01)
Project #
5R01DE014372-09
Application #
8471012
Study Section
Special Emphasis Panel (ZRG1-MOSS-B (02))
Program Officer
Lunsford, Dwayne
Project Start
2002-01-01
Project End
2014-05-31
Budget Start
2013-06-01
Budget End
2014-05-31
Support Year
9
Fiscal Year
2013
Total Cost
$539,723
Indirect Cost
$162,817
Name
University of Washington
Department
Engineering (All Types)
Type
Schools of Engineering
DUNS #
605799469
City
Seattle
State
WA
Country
United States
Zip Code
98195
Hutcherson, J A; Gogeneni, H; Yoder-Himes, D et al. (2016) Comparison of inherently essential genes of Porphyromonas gingivalis identified in two transposon-sequencing libraries. Mol Oral Microbiol 31:354-64
Wright, C J; Wu, H; Melander, R J et al. (2014) Disruption of heterotypic community development by Porphyromonas gingivalis with small molecule inhibitors. Mol Oral Microbiol 29:185-93
Wang, Huizhi; Zhou, Huaxin; Duan, Xiaoxian et al. (2014) Porphyromonas gingivalis-induced reactive oxygen species activate JAK2 and regulate production of inflammatory cytokines through c-Jun. Infect Immun 82:4118-26
Wright, Christopher J; Xue, Peng; Hirano, Takanori et al. (2014) Characterization of a bacterial tyrosine kinase in Porphyromonas gingivalis involved in polymicrobial synergy. Microbiologyopen 3:383-94
Hendrickson, Erik L; Wang, Tiansong; Beck, David A C et al. (2014) Proteomics of Fusobacterium nucleatum within a model developing oral microbial community. Microbiologyopen 3:729-51
Wright, C J; Burns, L H; Jack, A A et al. (2013) Microbial interactions in building of communities. Mol Oral Microbiol 28:83-101
Hirano, T; Beck, D A C; Wright, C J et al. (2013) Regulon controlled by the GppX hybrid two component system in Porphyromonas gingivalis. Mol Oral Microbiol 28:70-81
Kuboniwa, Masae; Tribble, Gena D; Hendrickson, Erik L et al. (2012) Insights into the virulence of oral biofilms: discoveries from proteomics. Expert Rev Proteomics 9:311-23
Hendrickson, Erik L; Wang, Tiansong; Dickinson, Brittany C et al. (2012) Proteomics of Streptococcus gordonii within a model developing oral microbial community. BMC Microbiol 12:211
Hirano, Takanori; Beck, David A C; Demuth, Donald R et al. (2012) Deep Sequencing of Porphyromonas gingivalis and Comparative Transcriptome Analysis of a LuxS Mutant. Front Cell Infect Microbiol 2:79

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