Inhibition of tRNA Synthetases Induces Persistence inChlamydia

Abstract

AbstractChlamydia trachomatisis the leading cause of bacterial sexually transmitted infections, andC. pneumoniaecauses community-acquired respiratory infections.In vivo, the host immune system will release interferon-gamma (IFNγ) to combat infection. IFNγ activates human cells to produce the tryptophan (trp) catabolizing enzyme, IDO. Consequently, there is a reduction in cytosolic trp in IFNγ-activated host cells. In evolving to obligate intracellular dependence,Chlamydiahas significantly reduced its genome size and content as it relies on the host cell for various nutrients. Importantly,C. trachomatisandC. pneumoniaeare trp auxotrophs and are starved for this essential nutrient when the human host cell is exposed to IFNγ. To survive this, chlamydiae enter an alternative growth state referred to as persistence. Chlamydial persistence is characterized by a halt in the division cycle, aberrant morphology, and, in the case of IFNγ-induced persistence, trp codon-dependent changes in transcription. We hypothesize that these changes in transcription are dependent on the particular amino acid starvation state. To investigate the chlamydial response mechanisms acting when other amino acids become limiting, we tested the efficacy of prokaryotic specific tRNA synthetase inhibitors, indolmycin and AN3365, to mimic starvation of trp and leucine, respectively. We show that these drugs block chlamydial growth and induce changes in morphology and transcription consistent with persistence. Importantly, growth inhibition was reversed when the compounds were removed from the medium. With these data, we find that indolmycin and AN3365 are valid tools that can be used to mimic the persistent state independently of IFNγ.ImportanceThe obligate intracellular pathogenChlamydia trachomatis, although treatable, remains a major public health concern due to rising infection rates. The asymptomatic nature of mostChlamydiainfections is hypothesized to be a product of its ability to transition into a slow-growing state referred to as persistence. The most physiologically relevant inducer of persistence is the immune cytokine IFNγ, which in humans activates an enzyme that degrades tryptophan, an essential amino acid thatChlamydiascavenges from the host cell. Unfortunately, the exact timing at whichChlamydiais starved after IFNγ treatment is inexact. To mechanistically study persistence using genetic tools, an experimental model where amino acid starvation can be induced at specific times is needed. Here, we demonstrate the capability of tRNA synthetase inhibitors, indolmycin and AN3365, to model persistence independently from the use of IFNγ. These tools will also allow comparisons between amino acid stress responses in this unique bacterium.