Argonaute-mediated immune subversion in Leishmania-infected macrophages
ArgoLeish leverages the protozoan parasite Leishmania as a unique model to uncover how intracellular pathogens subvert macrophage innate immune functions and favor their own survival by targeting the host Argonaute/RISC/miRNA axis. The key strength of this proposal is the complementary expertise of the German and French teams in Argonaute biology and Leishmania/macrophage interaction, respectively. Their collaboration has already yielded important preliminary results, jointly pre-published on bioRxiv. Argonaute (Ago) proteins are fundamental regulators of mammalian gene expression. As core components of the RNA-induced silencing complex (RISC), they mediate the functions of microRNAs (miRNAs), a class of non-coding RNAs that regulate gene expression post-transcriptionally by promoting mRNA degradation or inhibiting protein translation. Dysregulated RISC activity is linked to various pathologies – including microbial infection - revealing miRNAs as promising targets for RNA-based therapeutics. However, how pathogenic microbes exploit the Ago/RISC/miRNA axis to reprogram host cells remains largely unknown. Based on our robust, pre-published findings, we hypothesize that Leishmania exploits Ago2/RISC complexes to drive the significant changes we have seen in infected macrophages, reprogramming these highly toxic innate immune cells into tamed metabolic factories that promote chronic infection. We will apply a highly multidisciplinary approach to investigate this hypothesis through two objectives: First, we will assess infection-induced changes in the Ago2/RISC/miRNA axis and analyze their impact on macrophage phenotype and parasite survival, combining biochemical, cell biological and genetic loss-of-function approaches with in vitro and in vivo infection studies. This will provide unprecedented insights into the dynamic host miRNA machinery and its manipulation by intracellular Leishmania, while evaluating the Ago2/RISC axis as a potential therapeutic target. Second, we will map and functionally validate Ago2-dependent miRNA-mRNAs interactions during Leishmania infection using single cell SLAM-seq, Ago2-enhanced crosslinking and immunoprecipitation (eCLIP), and transfection of miRNA mimic (agomirs, antagomirs). This will uncover novel regulatory mechanisms of macrophage immune subversion and test non-canonical miRNA-mRNA nodes as novel targets for host-directed anti-leishmanial intervention. In conclusion, by harnesses the complementary expertise of its German and French partners, ArgoLeish will yield unprecedented functional insights into the pathologic feedback loop driven by parasite-induced, miRNA-dependent immune silencing, which in turn further sustains parasite persistence. Our project establishes an innovative experimental framework for investigating miRNA-dependent expression control in host-pathogen interactions that (i) provides a powerful blueprint for other intracellular pathogens targeting macrophages and causing severe immunopathologies, such as tuberculosis, candidiasis, AIDS or COVID, (ii) fills important knowledge gaps on macrophage immune subversion, and (ii) provides new impetus for the stalled anti-leishmanial drug discovery process by targeting the Ago/RISC/miRNA axis for host-directed, anti-microbial therapy.
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Imported from OpenAIRE Germany | Grant ANR-25-CE15-5898.
Open sourceOverview
Grant agreement ANR-25-CE15-5898 | ARGOLEISH
Open sourceArgoLeish leverages the protozoan parasite Leishmania as a unique model to uncover how intracellular pathogens subvert macrophage innate immune functions and favor their own survival by targeting the host Argonaute/RISC/miRNA axis. The key strength of this proposal is the complementary expertise of the German and French teams in Argonaute biology and Leishmania/macrophage interaction, respectively. Their collaboration has already yielded important preliminary results, jointly pre-published on bioRxiv. Argonaute (Ago) proteins are fundamental regulators of mammalian gene expression. As core components of the RNA-induced silencing complex (RISC), they mediate the functions of microRNAs (miRNAs), a class of non-coding RNAs that regulate gene expression post-transcriptionally by promoting mRNA degradation or inhibiting protein translation. Dysregulated RISC activity is linked to various pathologies – including microbial infection - revealing miRNAs as promising targets for RNA-based therapeutics. However, how pathogenic microbes exploit the Ago/RISC/miRNA axis to reprogram host cells remains largely unknown. Based on our robust, pre-published findings, we hypothesize that Leishmania exploits Ago2/RISC complexes to drive the significant changes we have seen in infected macrophages, reprogramming these highly toxic innate immune cells into tamed metabolic factories that promote chronic infection. We will apply a highly multidisciplinary approach to investigate this hypothesis through two objectives: First, we will assess infection-induced changes in the Ago2/RISC/miRNA axis and analyze their impact on macrophage phenotype and parasite survival, combining biochemical, cell biological and genetic loss-of-function approaches with in vitro and in vivo infection studies. This will provide unprecedented insights into the dynamic host miRNA machinery and its manipulation by intracellular Leishmania, while evaluating the Ago2/RISC axis as a potential therapeutic target. Second, we will map and functionally validate Ago2-dependent miRNA-mRNAs interactions during Leishmania infection using single cell SLAM-seq, Ago2-enhanced crosslinking and immunoprecipitation (eCLIP), and transfection of miRNA mimic (agomirs, antagomirs). This will uncover novel regulatory mechanisms of macrophage immune subversion and test non-canonical miRNA-mRNA nodes as novel targets for host-directed anti-leishmanial intervention. In conclusion, by harnesses the complementary expertise of its German and French partners, ArgoLeish will yield unprecedented functional insights into the pathologic feedback loop driven by parasite-induced, miRNA-dependent immune silencing, which in turn further sustains parasite persistence. Our project establishes an innovative experimental framework for investigating miRNA-dependent expression control in host-pathogen interactions that (i) provides a powerful blueprint for other intracellular pathogens targeting macrophages and causing severe immunopathologies, such as tuberculosis, candidiasis, AIDS or COVID, (ii) fills important knowledge gaps on macrophage immune subversion, and (ii) provides new impetus for the stalled anti-leishmanial drug discovery process by targeting the Ago/RISC/miRNA axis for host-directed, anti-microbial therapy.