Turning on Persistence: Novel Molecular Determinants that Underpin P. gingivalis' Intracellular Survival In Epithelial Cells

SUMMARY of The PARENT AWARD A growing multidisciplinary evidence critically underpins that Porphyromonas gingivalis, a leading pathobiont of the oral cavity that successfully remodels oral microbial communities to a pathophysiological state, can live in concert with human gingival epithelial cells (GECs). Epithelial cells are emerged as an integrally important arm

of innate defenses in the oral mucosa, while recent observations suggest that these cells can be exploited as privileged growth niches and a reservoir by P. gingivalis, which can intracellularly multiply and remain largely unharmed in GECs. Despite, extensive systems level molecular knowledge exists on the P. gingivalis and GEC

interaction, there is considerably little known on the intracellular life of the organism in this central cell type. We recently revealed that formation of autophagosomes is critical for the P. gingivalis’ intracellular replication and evasion of the anti-microbial degradation pathways in the GECs. Our novel preliminary findings also support

that lipidation of LC3-C, a key molecule in the ‘selective autophagy’ pathway, which targets intracellular pathogens is significantly modulated by P. gingivalis under the control of an anti-stress molecule, HSP27. Further, glutathione peroxidase (GpX1), a major host redox balance enzyme and a regulator of autophagic flux

largely impacted on the global LC3 lipidation state of GECs upon infection. The inhibition of either HSP27 or GpX1 appears to severely affect the intracellular trafficking and viability of the microorganism. The central hypothesis is that P. gingivalis induces a distinct form of selective autophagy, which results in

protection of bacterial life and ultimately securing of P. gingivalis’ persistence in the oral mucosa. To test this novel hypothesis, we will pursue two-pronged approach, where we propose the selective autophagy requires tightly coordinated actions of HSP27 and GpX1 to form autophagosomes that fully function as

protected replicative niches for P. gingivalis. Aim 1 will define the selective molecular machinery that drives P. gingivalis-containing autophagosome assembly under the control of HSP27 and the mechanisms that disrupt autophagic flux for the evasion of cellular degradation pathways. Aim 2 will establish the role of GpX1 in

regulating the selective autophagy in infection via redox homeostasis and suppressing autophagolysosomal machinery. Both aims will employ reductionist primary GECs culture systems to functionally dissect out the mechanisms and phenotypically characterize the molecular events and sub-cellular components. Aim 3 will

establish the dual significance of these two components using oral epithelial-tissue-specific knockout mice models. Thus, this proposal aims to fill a significant gap in our fundamental knowledge that is how P. gingivalis, a facultatively intracellular pathogen, establishes a privileged cellular environment and converts

nutritionally rich epithelial cells into potentially a central reservoir for bacterial growth and persistence in the oral mucosa. Ultimately, the knowledge gained may translate into molecular strategies that can control or reduce the intracellular colonization and survival methods employed by this important opportunistic pathogen.