Abstract
Interferon-gamma (IFN-γ) inhibits intracellular replication of Francisella tularensis in human monocyte-derived macrophages (HMDM) and in mice, but the mechanisms of this protective effect are poorly characterized. We used genome-wide RNA interference (RNAi) screening in the human macrophage cell line THP-1 to identify genes that mediate the beneficial effects of IFN-γ on F. tularensis infection. A primary screen identified ∼200 replicated candidate genes. These were prioritized according to mRNA expression in IFN-γ-primed and F. tularensis-challenged macrophages. A panel of 20 top hits was further assessed by re-testing using individual shRNAs or siRNAs in THP-1 cells, HMDMs and primary human lung macrophages. Six of eight validated genes tested were also found to confer resistance to Listeria monocytogenes infection, suggesting a broadly shared host gene program for intracellular pathogens. The F. tularensis-validated hits included ‘druggable’ targets such as TNFRSF9, which encodes CD137. Treating HMDM with a blocking antibody to CD137 confirmed a beneficial role of CD137 in macrophage clearance of F. tularensis. These studies reveal a number of important mediators of IFN-γ activated host defense against intracellular pathogens, and implicate CD137 as a potential therapeutic target and regulator of macrophage interactions with Francisella tularensis.
Highlights
Francisella tularensis is a highly virulent, facultative intracellular bacterium that causes tularemia [1,2]
We first tested our assumption that IFN-c activation would enable the human macrophage cell line THP-1 to inhibit intracellular replication of F. tularensis
THP-1 cells were differentiated by phorbol-12-myristate acetate (PMA), followed by pre-treatment with IFN-c (100 U/ml) for 24 h, and the cells were infected with green fluorescent protein labeled F. tularensis live vaccine strain (LVS) (GFP-LVS) as described in Methods
Summary
Francisella tularensis is a highly virulent, facultative intracellular bacterium that causes tularemia [1,2]. Macrophages are the main site of replication of F. tularensis [4], and lung macrophages are especially targeted in inhalational tularemia [5]. Intracellular killing of F. tularensis by macrophages depends on IFN-c-induced activation [7,8,9], but the specific mechanisms of IFN-c induced killing are not fully understood. More recent data implicates autophagy [16,17] and inflammasomes [18,19,20], the importance of these in the specific context of IFN-c stimulation has not been demonstrated. We hypothesized that IFN-c limits intracellular pathogen F. tularensis by activation of specific genes, and that functional genomic screening could identify these genes
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
