Genetic Screen in Chlamydia muridarum Reveals Role for an Interferon-Induced Host Cell Death Program in Antimicrobial Inclusion Rupture

Abstract

Interferon-regulated immune defenses protect mammals from pathogenically diverse obligate intracellular bacterial pathogens of the genus Chlamydia. Interferon gamma (IFN-γ) is especially important in controlling the virulence of Chlamydia species and thus impacts the modeling of human chlamydial infection and disease in mice. How IFN-γ contributes to cell-autonomous defenses against Chlamydia species and how these pathogens evade IFN-γ-mediated immunity in their natural hosts are not well understood. We conducted a genetic screen which identified 31 IFN-γ-sensitive (Igs) mutants of the mouse model pathogen Chlamydia muridarum. Genetic suppressor analysis and lateral gene transfer were used to map the phenotype of one of these mutants, Igs4, to a missense mutation in a putative chlamydial inclusion membrane protein, TC0574. We observed the lytic destruction of Igs4-occupied inclusions and accompanying host cell death in response to IFN-γ priming or various proapoptotic stimuli. However, Igs4 was insensitive to IFN-γ-regulated cell-autonomous defenses previously implicated in anti-Chlamydia trachomatis host defense in mice. Igs4 inclusion integrity was restored by caspase inhibitors, indicating that the IFN-γ-mediated destruction of Igs4 inclusions is dependent upon the function of caspases or related prodeath cysteine proteases. We further demonstrated that the Igs4 mutant is immune restricted in an IFN-γ-dependent manner in a mouse infection model, thereby implicating IFN-γ-mediated inclusion destruction and host cell death as potent in vivo host defense mechanisms to which wild-type C. muridarum is resistant. Overall, our results suggest that C. muridarum evolved resistance mechanisms to counter IFN-γ-elicited programmed cell death and the associated destruction of intravacuolar pathogens. IMPORTANCE Multiple obligatory intracellular bacteria in the genus Chlamydia are important pathogens. In humans, strains of C. trachomatis cause trachoma, chlamydia, and lymphogranuloma venereum. These diseases are all associated with extended courses of infection and reinfection that likely reflect the ability of chlamydiae to evade various aspects of host immune responses. Interferon-stimulated genes, driven in part by the cytokine interferon gamma, restrict the host range of various Chlamydia species, but how these pathogens evade interferon-stimulated genes in their definitive host is poorly understood. Various Chlamydia species can inhibit death of their host cells and may have evolved this strategy to evade prodeath signals elicited by host immune responses. We present evidence that chlamydia-induced programmed cell death resistance evolved to counter interferon- and immune-mediated killing of Chlamydia-infected cells.

Evasion of Innate Lymphoid Cell-Regulated Gamma Interferon Responses by Chlamydia muridarum To Achieve Long-Lasting Colonization in Mouse Colon

“…It is worth noting that besides clone G28.51.1 (86,87), there are other C. muridarum mutants that have been shown to be defective in colonizing the GI tract and susceptible to IFN-␥ inhibition (88,89). A common property of these mutants is that they all carry mutations in chromosomal genes.…”

An Ancient Molecular Arms Race: Chlamydia vs. Membrane Attack Complex/Perforin (MACPF) Domain Proteins

“…Regardless, it seems apparent that Chlamydia avoid Perforin-2 killing in epithelial cells by avoiding and suppressing MPEG1 expression. Although Perforin-2 proteins levels were not shown, inhibition of C. muridarum in IFNγ-primed murine embryonic fibroblasts was unchanged after Perforin-2 knockdown using siRNAs (74). These data may, once again, elude to alternative, species-specific susceptiblity of Chlamydia to Perforin-2.…”

Host cell death during infection withChlamydia: a double-edged sword

“…More recently, a genetic screen in C. muridarum uncovered several mutants that were more sensitive to IFN-γ-mediated restriction of bacterial growth in mouse cells (Giebel et al . 2019 ). A closer analysis of one mutant, which carried a specific point mutation in the putative Inc protein TC0574, revealed premature inclusion lysis, followed by induction of host cell death (Giebel et al .…”

“…A closer analysis of one mutant, which carried a specific point mutation in the putative Inc protein TC0574, revealed premature inclusion lysis, followed by induction of host cell death (Giebel et al . 2019 ). Moreover, levels of inclusion lysis and cell death were strongly enhanced when IFN-γ or IFN-β were added to the culture (Giebel et al .…”

“… 2017 ; Giebel et al . 2019 ). A profound understanding of this complex interplay between host and pathogen is critical for our understanding of Chlamydia diseases and anti-chlamydial host defenses.…”