Didier Soulat

The DEAD-box helicase DDX3X is a critical component of the TANK-binding kinase 1-dependent innate immune response.

TANK‐binding kinase 1 (TBK1) is of central importance for the induction of type‐I interferon (IFN) in response to pathogens. We identified the DEAD‐box helicase DDX3X as an interaction partner of TBK1. TBK1 and DDX3X acted synergistically in their ability to stimulate the IFN promoter, whereas RNAi‐mediated reduction of DDX3X expression led to an impairment of IFN production. Chromatin immunoprecipitation indicated that DDX3X is recruited to the IFN promoter upon infection with Listeria monocytogenes, suggesting a transcriptional mechanism of action. DDX3X was found to be a TBK1 substrate in vitro and in vivo. Phosphorylation‐deficient mutants of DDX3X failed to synergize with TBK1 in their ability to stimulate the IFN promoter. Overall, our data imply that DDX3X is a critical effector of TBK1 that is necessary for type I IFN induction.

Characterization of the interferon-producing cell in mice infected with Listeria monocytogenes.

Production of type I interferons (IFN-I, mainly IFNα and IFNβ) is a hallmark of innate immune responses to all classes of pathogens. When viral infection spreads to lymphoid organs, the majority of systemic IFN-I is produced by a specialized “interferon-producing cell” (IPC) that has been shown to belong to the lineage of plasmacytoid dendritic cells (pDC). It is unclear whether production of systemic IFN-I is generally attributable to pDC irrespective of the nature of the infecting pathogen. We have addressed this question by studying infections of mice with the intracellular bacterium Listeria monocytogenes. Protective innate immunity against this pathogen is weakened by IFN-I activity. In mice infected with L. monocytogenes, systemic IFN-I was amplified via IFN-β, the IFN-I receptor (IFNAR), and transcription factor interferon regulatory factor 7 (IRF7), a molecular circuitry usually characteristic of non-pDC producers. Synthesis of serum IFN-I did not require TLR9. In contrast, in vitro–differentiated pDC infected with L. monocytogenes needed TLR9 to transcribe IFN-I mRNA. Consistent with the assumption that pDC are not the producers of systemic IFN-I, conditional ablation of the IFN-I receptor in mice showed that most systemic IFN-I is produced by myeloid cells. Furthermore, results obtained with FACS-purified splenic cell populations from infected mice confirmed the assumption that a cell type with surface antigens characteristic of macrophages and not of pDC is responsible for bulk IFN-I synthesis. The amount of IFN-I produced in the investigated mouse lines was inversely correlated to the resistance to lethal infection. Based on these data, we propose that the engagement of pDC, the mode of IFN-I mobilization, as well as the shaping of the antimicrobial innate immune response by IFN-I differ between intracellular pathogens.

Type I IFN are host modulators of strain‐specific Listeria monocytogenes virulence

Type I IFN (IFN‐I) increase the sensitivity of cells and mice to lethal infection with Listeria monocytogenes. Therefore the amount of IFN‐I produced during infection might be an important factor determining Listeria virulence. Two commonly used strains of L. monocytogenes, EGD and LO28, were identified as, respectively, low and high inducers of IFN‐I synthesis in infected macrophages. Increased IFN‐I production resulted from the stronger ability of the LO28 strain to trigger the IRF3 signalling pathway and correlated with an increased sensitization of macrophages to lethal infection. In contrast, stimulation of NFκB, MAPK, or inflammasome signalling by the LO28 and EGD strains did not differ significantly. The LO28 strain was more virulent in wild‐type (wt) C57/BL6 mice than the EGD strain whereas both strains were similarly virulent in IFN‐I receptor‐deficient C57/BL6 mice. Together our data suggest that isolates of wt L. monocytogenes differ in their ability to trigger the IRF3 signalling pathway and IFN‐I production, and that the amount of IFN‐I produced during infection is an important determinant of Listeria virulence.

Cytoplasmic Listeria monocytogenes stimulates IFN-β synthesis without requiring the adapter protein MAVS

The mitochondria‐associated adapter protein MAVS (also called IPS‐1, VISA or CARDIF, designated MAVS for reasons of simplicity in our manuscript) relays signals from cytoplasmic sensors of viral RNA to the IRF3 kinase complex and the interferon‐β (IFN‐β) gene. Using siRNA‐mediated knock‐down in macrophages we show that IFN‐β synthesis in response to transfected, intracellular double‐stranded RNA (dsRNA), a pathogen‐associated molecular pattern of viruses, is decreased in absence of MAVS. By contrast, the Gram‐positive bacterium Listeria monocytogenes targets the IFN‐β gene without detectable MAVS requirement. The data show that MAVS is not a central adapter protein for all cytoplasmic pathogen sensors that stimulate IFN‐β synthesis.

LipA, a tyrosine and lipid phosphatase involved in the virulence of Listeria monocytogenes.

ABSTRACT Intracellular bacterial pathogens manipulate host cell functions by producing enzymes that stimulate or antagonize signal transduction. The Listeria monocytogenes genome contains a gene, lmo1800, encoding a protein with a conserved motif of conventional tyrosine phosphatases. Here, we report that the lmo1800-encoded protein LipA is secreted by Listeria and displays tyrosine as well as lipid phosphatase activity in vitro. Bacteria lacking LipA are severely attenuated in virulence in vivo, thus revealing a so-far-undescribed enzymatic activity involved in Listeria infection.