AMANDA BRINKWORTH, PhD
As an obligate intracellular bacterium with a highly-reduced genome, Chlamydia trachomatis relies upon the acquisition of nutrients and metabolic substrates from its host to enable bacterial replication and proliferation. In human females, C. trachomatis infections primarily become established in the endocervix, which consists of a monolayer of polarized epithelial cells. Given that the first tissue Chlamydia comes into contact with during male-to-female sexual transmission is the stratified epithelium of the ectocervix, it is likely that these cells also become infected. The apical surface of the fully stratified epithelium of the ectocervix is thought to be nutrient-poor due to their lack of proximity to nutrient-laden serum. These cells are typically metabolically inactive and undergo frequent exfoliation. Thus, it is expected that infection of apical cells of the endocervix will result in nutrient-starved Chlamydia. Notably, the phenomenon of Chlamydia becoming persistent during starvation of key nutrients (eg. iron, amino acids) has been well-characterized in cultured cells and is typically reversible upon removal of the stressor or supplementation with the missing nutrients. We expect that chlamydiae are able to establish infection in the stratified epithelium and can serve as a slow-growing or even quiescent reservoir for recurring infections.
In collaboration with the Carabeo laboratory, we are using the in vitro stratified epithelium model to study chlamydial growth and survival in the ectocervix. In fact, Nogueira et al. (Front Cell Infect Microbiol. 2017. doi: 10.3389/fcimb.2017.00438.) revealed that infection of the apical surface of fully stratified epithelium resulted in low numbers of intracellular Chlamydia that did not complete development. We are using single-cell transcriptomic, proteomic and metabolic approaches to characterize differences between basal, central, and apical cells of the stratified epithelium. By revealing which nutrient acquisition factors and metabolic enzymes are absent from cells lining the apical surface, we can anticipate alternate metabolic factors Chlamydia will need to scavenge from dying cells or nearby microflora to survive. We will use targeted-metabolic labeling approaches to determine which metabolites can be acquired and utilized by Chlamydia and define the metabolic requirement for restoration of chlamydial development in the upper layers of the stratified epithelium. Taken together these approaches will define the metabolic requirements for ectocervical colonization by C. trachomatis and indicate the ectocervix as a reservoir for chronic recurring infections.
Murphy KN, Brinkworth AJ. Manipulation of Focal Adhesion Signaling by Pathogenic Microbes. International Journal of Molecular Sciences. 2021; 22(3):1358. doi: 10.3390/ijms22031358
Pedrosa AT, Murphy KN, Nogueira AT, Brinkworth AJ, Thwaites TR, Aaron J, Chew TL, Carabeo RA. A post-invasion role for Chlamydia type III effector TarP in modulating the dynamics and organization of host cell focal adhesions. J Biol Chem. 2020 Oct 23;295(43):14763-14779. doi:10.1074/jbc.RA120.015219. Epub 2020 Aug 25. PMID: 32843479
Drecktrah D, Hall LS, Brinkworth AJ, Comstock JR, Wassarman KM, Samuels DS. Characterization of 6S RNA in the Lyme disease spirochete. Mol Microbiol. 2020 Feb;113(2):399-417. doi: 10.1111/mmi.14427. Epub 2019 Dec 11. PMID: 31742773
Pokorzynski ND, Brinkworth AJ, Carabeo R. A bipartite iron-dependent transcriptional regulation of the tryptophan salvage pathway in Chlamydia trachomatis. Elife. 2019 Apr 2;8: e42295. doi: 10.7554/eLife.42295. PMID: 30938288
Brinkworth AJ, Wildung MA, Carabeo, RA. Genomewide Transcriptional Responses of Iron-Starved Chlamydia trachomatis Reveal Prioritization of Metabolic Precursor Synthesis over Protein Translation. mSystems. 2018 Feb 13:3(1): e00184-17.
Brinkworth AJ, Hammer CH, Olano LR, Kobayashi SD, Chen L, Kreiswirth BN, DeLeo FR. (2015) Identification of outer membrane and exoproteins of the carbapenem-resistant strain K.pneumoniae ST258. PLoS One. 2015 Apr 20; 10(4): e0123219.
Brinkworth, AJ.; Malcom, DS.; Pedrosa, AT.; Roguska, K; Shahbazian, S.; Graham, JE; Hayward, RD; Carabeo, RA. (2011) Chlamydia trachomatis Slc1 is a type III secretion chaperone that enhances the translocation of its invasion effector substrate TARP. Mol Microbiol 82(1): 134-144.