Chlamydia trachomatis overcomes human cell-autonomous immunity through secretion of a novel inclusion membrane protein IncU

Abstract

Chlamydia is the most common sexually transmitted bacterial infection in the world. In the United States, over 100 million people are infected each year, and from 2015 to 2019 incidence of chlamydia rose by 19%. Of extreme concern, this increase has occurred despite readily available effective antibiotics and national screening recommendations for young women at highest risk of contracting chlamydia. A chlamydia vaccine is needed, however our near-complete lack of knowledge of protective immune responses that could prevent infection is a major barrier for vaccine development. Research to unveil correlates of protection is a significant challenge—studies in humans carry major ethical considerations, and there is no animal model that faithfully recapitulates the natural history of infection as it occurs in humans. Careful design of in vitro research using human cells and tissues and work to identify the utility and limitations of various animal models are critical research priorities. To date, interferon gamma (IFNγ) secreting T-cells represent the strongest plausible correlate of protection against chlamydia. Because the etiological agent of chlamydia, the bacterium Chlamydia trachomatis, is an obligate intracellular pathogen that primarily infects human epithelial cells, an important aspect to the protective capacity of IFNγ lies in the ability of this cytokine to stimulate innate immunological responses in the cells that harbor and support chlamydial growth. The ability of these classically non-immune cells to mount innate immune responses to protect themselves against intracellular pathogens is often termed cell-autonomous immunity. IFNγ-stimulated cell-autonomous immunity against intracellular pathogens is a topic of ongoing investigation in the Chlamydia field and other systems, and its importance as a potent anti-Chlamydia response is only recently appreciated. While murine cell-autonomous immunity renders epithelial cells able to mark C. trachomatis-containing vacuoles for destruction, this pathogen readily evades detection and growth restriction in its native human host cells. I have dedicated my dissertation work to understanding the relationship between human cell-autonomous immunity and Chlamydia trachomatis and, importantly, identifying how this pathogen resists this potent immune pathway. By screening a novel library of chimeric interspecies Chlamydia mutant strains, I identified a genetic locus required for resistance. Follow up analysis of mutant C. trachomatis strains aided my identification of a single virulence gene, incU. Human cells infected with incU-mutants can recognize chlamydial vacuoles, termed inclusions, leading to the recruitment of downstream cell-autonomous immunity proteins. Ultimately, this leads to severe growth restriction of these C. trachomatis mutants. With contributions from collaborators at the University of Washington and Oregon State University, I found that incU mutants failed to replicate in vivo in the nonhuman primate model. In vitro infection of primary NHP fibroblasts revealed that NHP cells exhibit an IFNγ-inducible cell-autonomous immune response that is mechanistically similar to the human pathway, providing supporting evidence that in vivo growth defects were due to incU mutants’ susceptibility to IFNγ-stimulated cell-autonomous immunity. Taken together, these data shed new light on the relationship between IFNγ-stimulated immunity and C. trachomatis, while identifying a relevant animal model in which to further explore the protective capacity of IFNγ in humans.