Project acronym ANTIViR
Project Molecular mechanisms of interferon-induced antiviral restriction and signalling
Researcher (PI) Caroline GOUJON
Host Institution (HI) INSTITUT NATIONAL DE LA SANTE ET DE LA RECHERCHE MEDICALE
Call Details Starting Grant (StG), LS6, ERC-2017-STG
Summary Interferons (IFNs), which are signalling proteins produced by infected cells, are the first line of defence against viral infections. IFNs induce, in infected and neighbouring cells, the expression of hundreds of IFN-stimulated genes (ISGs). The ISGs in turn induce in cells a potent antiviral state, capable of preventing replication of most viruses, including Human Immunodeficiency Virus type 1 (HIV-1) and influenza A virus (FLUAV). Identifying the antiviral ISGs and understanding their mechanisms of action is therefore crucial to progress in the fight against viruses.
ISGs playing a role in the antiviral state have been identified, such as human MX1, a well-known antiviral factor able to restrict numerous viruses including FLUAV, and MX2, an HIV-1 inhibitor. Both proteins bind to viral components but their detailed mechanisms of action, as well as the consequences of restriction on the activation of the innate immune system, remain unclear. Moreover, our preliminary work shows that additional anti-HIV-1 and anti-FLUAV ISGs remain to identify.
In this context, this proposal seeks an ERC StG funding to explore 3 major aims: 1) unravelling the mechanisms of antiviral action of MX proteins, by taking advantage of their similar structure and engineered chimeric proteins, and by using functional genetic screens to identify their cofactors; 2) investigating the consequences of incoming virus recognition by MX proteins on innate immune signalling, by altering their expression in target cells and measuring the cell response in terms of gene induction and cytokine production; 3) identifying and characterizing new ISGs able to inhibit viral replication with a combination of powerful approaches, including a whole-genome CRISPR/Cas9 knock-out screen.
Overall, this proposal will provide a better understanding of the molecular mechanisms involved in the antiviral effect of IFN, and may guide future efforts to identify novel therapeutic targets against major pathogenic viruses.
Summary
Interferons (IFNs), which are signalling proteins produced by infected cells, are the first line of defence against viral infections. IFNs induce, in infected and neighbouring cells, the expression of hundreds of IFN-stimulated genes (ISGs). The ISGs in turn induce in cells a potent antiviral state, capable of preventing replication of most viruses, including Human Immunodeficiency Virus type 1 (HIV-1) and influenza A virus (FLUAV). Identifying the antiviral ISGs and understanding their mechanisms of action is therefore crucial to progress in the fight against viruses.
ISGs playing a role in the antiviral state have been identified, such as human MX1, a well-known antiviral factor able to restrict numerous viruses including FLUAV, and MX2, an HIV-1 inhibitor. Both proteins bind to viral components but their detailed mechanisms of action, as well as the consequences of restriction on the activation of the innate immune system, remain unclear. Moreover, our preliminary work shows that additional anti-HIV-1 and anti-FLUAV ISGs remain to identify.
In this context, this proposal seeks an ERC StG funding to explore 3 major aims: 1) unravelling the mechanisms of antiviral action of MX proteins, by taking advantage of their similar structure and engineered chimeric proteins, and by using functional genetic screens to identify their cofactors; 2) investigating the consequences of incoming virus recognition by MX proteins on innate immune signalling, by altering their expression in target cells and measuring the cell response in terms of gene induction and cytokine production; 3) identifying and characterizing new ISGs able to inhibit viral replication with a combination of powerful approaches, including a whole-genome CRISPR/Cas9 knock-out screen.
Overall, this proposal will provide a better understanding of the molecular mechanisms involved in the antiviral effect of IFN, and may guide future efforts to identify novel therapeutic targets against major pathogenic viruses.
Max ERC Funding
1 499 794 €
Duration
Start date: 2017-12-01, End date: 2022-11-30
Project acronym ASTHMACRYSTALCLEAR
Project Role of protein crystallization in type 2 immunity and asthma
Researcher (PI) Bart LAMBRECHT
Host Institution (HI) VIB VZW
Call Details Advanced Grant (AdG), LS6, ERC-2017-ADG
Summary Spontaneous protein crystallization is a rare event in biology. Eosinophilic inflammation such as seen in the airways in asthma, chronic rhinosinusitis and helminth infection is however accompanied by accumulation of large amounts of extracellular Charcot-Leyden crystals. These are made of Galectin-10, a protein of unknown function produced by eosinophils, hallmark cells of type 2 immunity. In mice, eosinophilic inflammation is also accompanied by protein crystal build up, composed of the chitinase-like proteins Ym1 and Ym2, produced by alternatively activated macrophages. Here we challenge the current view that these crystals are just markers of eosinophil demise or macrophages activation. We hypothesize that protein crystallization serves an active role in immunoregulation of type 2 immunity. On the one hand, crystallization might turn a harmless protein into a danger signal. On the other hand, crystallization might sequester and eliminate the physiological function of soluble Galectin-10 and Ym1, or prolong it via slow release elution. For full understanding, we therefore need to understand the function of the proteins in a soluble and crystalline state. Our program at the frontline of immunology, molecular structural biology and clinical science combines innovative tool creation and integrative research to investigate the structure, function, and physiology of galectin-10 and related protein crystals. We chose to study asthma as the crystallizing proteins are abundantly present in human and murine disease. There is still a large medical need for novel therapies that could benefit patients with chronic steroid-resistant disease, and are alternatives to eosinophil-depleting antibodies whose long term effects are unknown.
Summary
Spontaneous protein crystallization is a rare event in biology. Eosinophilic inflammation such as seen in the airways in asthma, chronic rhinosinusitis and helminth infection is however accompanied by accumulation of large amounts of extracellular Charcot-Leyden crystals. These are made of Galectin-10, a protein of unknown function produced by eosinophils, hallmark cells of type 2 immunity. In mice, eosinophilic inflammation is also accompanied by protein crystal build up, composed of the chitinase-like proteins Ym1 and Ym2, produced by alternatively activated macrophages. Here we challenge the current view that these crystals are just markers of eosinophil demise or macrophages activation. We hypothesize that protein crystallization serves an active role in immunoregulation of type 2 immunity. On the one hand, crystallization might turn a harmless protein into a danger signal. On the other hand, crystallization might sequester and eliminate the physiological function of soluble Galectin-10 and Ym1, or prolong it via slow release elution. For full understanding, we therefore need to understand the function of the proteins in a soluble and crystalline state. Our program at the frontline of immunology, molecular structural biology and clinical science combines innovative tool creation and integrative research to investigate the structure, function, and physiology of galectin-10 and related protein crystals. We chose to study asthma as the crystallizing proteins are abundantly present in human and murine disease. There is still a large medical need for novel therapies that could benefit patients with chronic steroid-resistant disease, and are alternatives to eosinophil-depleting antibodies whose long term effects are unknown.
Max ERC Funding
2 499 846 €
Duration
Start date: 2018-08-01, End date: 2023-07-31
Project acronym BASILIC
Project Decoding at systems-level the crosstalk between the T cell antigen receptor, the CD28 costimulator and the PD-1 coinhibitor under physiological and pathological conditions.
Researcher (PI) Bernard MALISSEN
Host Institution (HI) CENTRE NATIONAL DE LA RECHERCHE SCIENTIFIQUE CNRS
Call Details Advanced Grant (AdG), LS6, ERC-2017-ADG
Summary Although the T cell antigen receptor (TCR) occupies a central place in T cell physiology, it does not work in isolation and the signals it triggers are tuned by receptors that convey positive (costimulators) and negative (coinhibitors) informations. We lack a satisfying comprehension of the way T cells integrate such multiple inputs to make informed decisions. The proteomics-based methodology we developed around the TCR places us in a favorable situation to decode at systems-level the crosstalk between the TCR, the CD28 costimulator and the PD-1 coinhibitor signaling pathways. The novelty of our approach stems from (1) its use of primary T cells, (2) its capacity to probe the architecture and dynamics of signalosomes resulting from T cell-antigen presenting cell encounters, (3) the attention we pay to the stoichiometry of the studied signalosomes, a key parameter largely ignored in previous studies, and (4) its multidisciplinary nature straddling molecular and organismal scales.
Our specific aims are:
Aim 1. To understand how the TCR and CD28 signaling pathways cooperate to achieve optimal T cell responses.
Aim 2. To determine whether CD28 is the sole target of the PD-1 coinhibitor.
Aim 3. To determine how under inflammatory conditions CD28 functions can be superseded by those of OX40, a costimulator of the TNFR superfamily.
Aim 4. To unveil how malfunctions of LAT, a key signaling hub used by the TCR, disrupt the TCR-CD28 crosstalk and result in unique pathogenic T cells that by becoming ‘autistic’ to TCR signals and addicted to CD28 signals lead to severe immunopathologies.
We think that combining genetic, epigenomics, proteomics, and computational approaches creates ideal experimental conditions to understand at system-levels how TCR, costimulatory, coinhibitory and inflammatory signals are integrated during T cell clonal expansion. Although of fundamental nature, our project should help understanding the harmful role PD-1 plays during anti-tumoral responses.
Summary
Although the T cell antigen receptor (TCR) occupies a central place in T cell physiology, it does not work in isolation and the signals it triggers are tuned by receptors that convey positive (costimulators) and negative (coinhibitors) informations. We lack a satisfying comprehension of the way T cells integrate such multiple inputs to make informed decisions. The proteomics-based methodology we developed around the TCR places us in a favorable situation to decode at systems-level the crosstalk between the TCR, the CD28 costimulator and the PD-1 coinhibitor signaling pathways. The novelty of our approach stems from (1) its use of primary T cells, (2) its capacity to probe the architecture and dynamics of signalosomes resulting from T cell-antigen presenting cell encounters, (3) the attention we pay to the stoichiometry of the studied signalosomes, a key parameter largely ignored in previous studies, and (4) its multidisciplinary nature straddling molecular and organismal scales.
Our specific aims are:
Aim 1. To understand how the TCR and CD28 signaling pathways cooperate to achieve optimal T cell responses.
Aim 2. To determine whether CD28 is the sole target of the PD-1 coinhibitor.
Aim 3. To determine how under inflammatory conditions CD28 functions can be superseded by those of OX40, a costimulator of the TNFR superfamily.
Aim 4. To unveil how malfunctions of LAT, a key signaling hub used by the TCR, disrupt the TCR-CD28 crosstalk and result in unique pathogenic T cells that by becoming ‘autistic’ to TCR signals and addicted to CD28 signals lead to severe immunopathologies.
We think that combining genetic, epigenomics, proteomics, and computational approaches creates ideal experimental conditions to understand at system-levels how TCR, costimulatory, coinhibitory and inflammatory signals are integrated during T cell clonal expansion. Although of fundamental nature, our project should help understanding the harmful role PD-1 plays during anti-tumoral responses.
Max ERC Funding
2 000 000 €
Duration
Start date: 2018-08-01, End date: 2022-07-31
Project acronym CANCER-DC
Project Dissecting Regulatory Networks That Mediate Dendritic Cell Suppression
Researcher (PI) Oren PARNAS
Host Institution (HI) THE HEBREW UNIVERSITY OF JERUSALEM
Call Details Starting Grant (StG), LS6, ERC-2017-STG
Summary Recent advances have shown that therapeutic manipulations of key cell-cell interactions can have dramatic clinical outcomes. Most notable are several early successes in cancer immunotherapy that target the tumor-T cell interface. However, these successes were only partial. This is likely because the few known interactions are just a few pieces of a much larger puzzle, involving additional signaling molecules and cell types. Dendritic cells (DCs), play critical roles in the induction/suppression of T cells. At early cancer stages, DCs capture tumor antigens and present them to T cells. However, in advanced cancers, the tumor microenvironment (TME) disrupts the crosstalk between DCs and T cells.
We will take a multi-step approach to explore how the TME imposes a suppressive effect on DCs and how to reverse this hazardous effect. First, we will use single cell RNA-seq to search for genes in aggressive human and mouse ovarian tumors that are highly expressed in advanced tumors compared to early tumors and that encode molecules that suppress DC activity. Second, we will design a set of CRISPR screens to find genes that are expressed in DCs and regulate the transfer of the suppressive signals. The screens will be performed in the presence of suppressive molecules to mimic the TME and are expected to uncover many key genes in DCs biology. We will develop a new strategy to find synergistic combinations of genes to target (named Perturb-comb), thereby reversing the effect of local tumor immunosuppressive signals. Lastly, we will examine the effect of modified DCs on T cell activation and proliferation in-vivo, and on tumor growth.
We expect to find: (1) Signaling molecules in the TME that affect the immune system. (2) New cytokines and cell surface receptors that are expressed in DCs and signal to T cells. (3) New key regulators in DC biology and their mechanisms. (4) Combinations of genes to target in DCs that reverse the TME’s hazardous effects.
Summary
Recent advances have shown that therapeutic manipulations of key cell-cell interactions can have dramatic clinical outcomes. Most notable are several early successes in cancer immunotherapy that target the tumor-T cell interface. However, these successes were only partial. This is likely because the few known interactions are just a few pieces of a much larger puzzle, involving additional signaling molecules and cell types. Dendritic cells (DCs), play critical roles in the induction/suppression of T cells. At early cancer stages, DCs capture tumor antigens and present them to T cells. However, in advanced cancers, the tumor microenvironment (TME) disrupts the crosstalk between DCs and T cells.
We will take a multi-step approach to explore how the TME imposes a suppressive effect on DCs and how to reverse this hazardous effect. First, we will use single cell RNA-seq to search for genes in aggressive human and mouse ovarian tumors that are highly expressed in advanced tumors compared to early tumors and that encode molecules that suppress DC activity. Second, we will design a set of CRISPR screens to find genes that are expressed in DCs and regulate the transfer of the suppressive signals. The screens will be performed in the presence of suppressive molecules to mimic the TME and are expected to uncover many key genes in DCs biology. We will develop a new strategy to find synergistic combinations of genes to target (named Perturb-comb), thereby reversing the effect of local tumor immunosuppressive signals. Lastly, we will examine the effect of modified DCs on T cell activation and proliferation in-vivo, and on tumor growth.
We expect to find: (1) Signaling molecules in the TME that affect the immune system. (2) New cytokines and cell surface receptors that are expressed in DCs and signal to T cells. (3) New key regulators in DC biology and their mechanisms. (4) Combinations of genes to target in DCs that reverse the TME’s hazardous effects.
Max ERC Funding
1 500 000 €
Duration
Start date: 2018-01-01, End date: 2022-12-31
Project acronym ChronosAntibiotics
Project Exploring the bacterial cell cycle to re-sensitize antibiotic-resistant bacteria
Researcher (PI) MARIANA LUISA TOMAS GOMES DE PINHO
Host Institution (HI) UNIVERSIDADE NOVA DE LISBOA
Call Details Consolidator Grant (CoG), LS6, ERC-2017-COG
Summary Over the next 35 years, antibiotic resistant bacteria are expected to kill more than 300 million people. The need to find alternative strategies for antimicrobial therapies remains a global challenge with several bottlenecks in the antibiotic discovery process. Using Staphylococcus aureus, the most common multidrug-resistant bacterium in the European Union and an excellent model organism for cell division in cocci, we propose:
(i) to find new pathways to re-sensitize resistant bacteria. Bacteria undergo major morphology changes during the cell cycle. We hypothesize that these changes generate windows of opportunity during which bacteria are more susceptible or more tolerant to the action of antibiotics. We will identify key regulators of the cell cycle in order to manipulate the duration of windows of opportunity for the action of existing antibiotics.
(ii) to develop new fluorescence-based reporters for whole-cell screenings of antimicrobial compounds with new modes of action, including compounds that arrest or delay the cell cycle; compounds that target non-essential pathways that are required for expression of resistance against existing antibiotics and therefore can be used as synergistic drugs for combination therapies; compounds that inhibit the production of virulence factors and compounds that revert persister states that are phenotypically resistant to antibiotics.
(iii) to unravel new modes of action of antibiotics by using the constructed reporter strains as powerful tools to learn how antibiotics act at the single cell level.
Over the past years, my group has become expert on the biology of S. aureus, has constructed powerful biological tools to study cell division and synthesis of the cell surface and has studied mechanisms of action of various antimicrobial compounds. We are therefore in a privileged position to quickly unravel the function of new players in the bacterial cell cycle and simultaneously contribute to accelerate antibiotic discovery.
Summary
Over the next 35 years, antibiotic resistant bacteria are expected to kill more than 300 million people. The need to find alternative strategies for antimicrobial therapies remains a global challenge with several bottlenecks in the antibiotic discovery process. Using Staphylococcus aureus, the most common multidrug-resistant bacterium in the European Union and an excellent model organism for cell division in cocci, we propose:
(i) to find new pathways to re-sensitize resistant bacteria. Bacteria undergo major morphology changes during the cell cycle. We hypothesize that these changes generate windows of opportunity during which bacteria are more susceptible or more tolerant to the action of antibiotics. We will identify key regulators of the cell cycle in order to manipulate the duration of windows of opportunity for the action of existing antibiotics.
(ii) to develop new fluorescence-based reporters for whole-cell screenings of antimicrobial compounds with new modes of action, including compounds that arrest or delay the cell cycle; compounds that target non-essential pathways that are required for expression of resistance against existing antibiotics and therefore can be used as synergistic drugs for combination therapies; compounds that inhibit the production of virulence factors and compounds that revert persister states that are phenotypically resistant to antibiotics.
(iii) to unravel new modes of action of antibiotics by using the constructed reporter strains as powerful tools to learn how antibiotics act at the single cell level.
Over the past years, my group has become expert on the biology of S. aureus, has constructed powerful biological tools to study cell division and synthesis of the cell surface and has studied mechanisms of action of various antimicrobial compounds. We are therefore in a privileged position to quickly unravel the function of new players in the bacterial cell cycle and simultaneously contribute to accelerate antibiotic discovery.
Max ERC Funding
2 533 500 €
Duration
Start date: 2018-03-01, End date: 2023-02-28
Project acronym DC_Nutrient
Project Investigating nutrients as key determinants of DC-induced CD8 T cell responses
Researcher (PI) David FINLAY
Host Institution (HI) THE PROVOST, FELLOWS, FOUNDATION SCHOLARS & THE OTHER MEMBERS OF BOARD OF THE COLLEGE OF THE HOLY & UNDIVIDED TRINITY OF QUEEN ELIZABETH NEAR DUBLIN
Call Details Consolidator Grant (CoG), LS6, ERC-2017-COG
Summary A new immunoregulatory axis has emerged in recent years demonstrating that cellular metabolism is crucial in controlling immune responses. This regulatory axis is acutely sensitive to nutrients that fuel metabolic pathways and support nutrient sensitive signalling pathways. My recent research demonstrates that nutrients are dynamically controlled and are not equally available to all immune cells. The data shows that activated T cells, clustered around a dendritic cell (DC), can consume the available nutrients, leaving the DC nutrient deprived in vitro. This local regulation of the DC nutrient microenvironment by neighbouring cells has profound effects on DC function and T cell responses. Nutrient deprived DC have altered signalling (decreased mTORC1 activity), increased pro-inflammatory functions (IL12 and costimulatory molecule expression) and induce enhanced T cell responses (proliferation, IFNγ production). However, proving this, particularly in vivo, is a major challenge as the tools to investigate nutrient dynamics within complex microenvironments have not yet been developed. This research programme will generate innovative new technologies to measure the local distribution of glucose, glutamine and leucine (all of which control mTORC1 signalling) to be visualised and quantified. These technologies will pioneer a new era of in vivo nutrient analysis. Nutrient deprivation of antigen presenting DC will then be investigated (using our new technologies) in response to various stimuli within the inflammatory lymph node and correlated to CD8 T cell responses. We will generate state-of-the-art transgenic mice to specifically knock-down nutrient transporters for glucose, glutamine, or leucine in DC to definitively prove that the availability of these nutrients to antigen presenting DC is a key mechanism for controlling CD8 T cells responses. This would be a paradigm shifting discovery that would open new horizons for the study of nutrient-regulated immune responses.
Summary
A new immunoregulatory axis has emerged in recent years demonstrating that cellular metabolism is crucial in controlling immune responses. This regulatory axis is acutely sensitive to nutrients that fuel metabolic pathways and support nutrient sensitive signalling pathways. My recent research demonstrates that nutrients are dynamically controlled and are not equally available to all immune cells. The data shows that activated T cells, clustered around a dendritic cell (DC), can consume the available nutrients, leaving the DC nutrient deprived in vitro. This local regulation of the DC nutrient microenvironment by neighbouring cells has profound effects on DC function and T cell responses. Nutrient deprived DC have altered signalling (decreased mTORC1 activity), increased pro-inflammatory functions (IL12 and costimulatory molecule expression) and induce enhanced T cell responses (proliferation, IFNγ production). However, proving this, particularly in vivo, is a major challenge as the tools to investigate nutrient dynamics within complex microenvironments have not yet been developed. This research programme will generate innovative new technologies to measure the local distribution of glucose, glutamine and leucine (all of which control mTORC1 signalling) to be visualised and quantified. These technologies will pioneer a new era of in vivo nutrient analysis. Nutrient deprivation of antigen presenting DC will then be investigated (using our new technologies) in response to various stimuli within the inflammatory lymph node and correlated to CD8 T cell responses. We will generate state-of-the-art transgenic mice to specifically knock-down nutrient transporters for glucose, glutamine, or leucine in DC to definitively prove that the availability of these nutrients to antigen presenting DC is a key mechanism for controlling CD8 T cells responses. This would be a paradigm shifting discovery that would open new horizons for the study of nutrient-regulated immune responses.
Max ERC Funding
1 995 861 €
Duration
Start date: 2018-05-01, End date: 2023-04-30
Project acronym DCPOIESIS
Project Steady-state and demand-driven dendritic cell generation
Researcher (PI) Caetano Maria Pacheco Pais Dos Reis e Sousa
Host Institution (HI) THE FRANCIS CRICK INSTITUTE LIMITED
Call Details Advanced Grant (AdG), LS6, ERC-2017-ADG
Summary Classical dendritic cells (cDCs) are leucocytes that play a key role in innate immunity as well as the initiation and regulation of T cell responses. cDCpoiesis starts with commitment of a bone marrow (BM) haematopoietic progenitor, known as the classical DC precursor (CDP), to the cDC lineage. CDPs then give rise to pre-cDCs that exit the BM via the blood and seed tissues to give rise to the two major types of fully-differentiated cDCs, the cDC1 and cDC2 subsets. The key parameters of cDCpoiesis are poorly understood. We propose to characterise the niche in which cDCs develop within the BM and to study how pre-cDCs seed tissues and establish local clones of differentiated cDC1 and cDC2. We further wish to ask how the activity of CDPs and pre-cDCs is altered following infection, inflammation or tissue damage. Finally, we want to know to what extent cDCpoiesis is affected by direct sensing of infection or cell damage by cDC precursors. All these objectives will be addressed in a mouse lineage tracing model in which cDC precursors are genetically labelled through the activity of a Cre recombinase driven by the Clec9a locus. These mice will be crossed to fluorescent protein reporter mice, including Confetti mice that allow for clonal analysis, and the appearance of labelled cDCs and cDC clones in tissues will be followed over time in the steady-state or after induction of infection or inflammation. The dependence of cDC precursor activity on specific pathogen and damage sensing pathways will be assessed by loss-of-function experiments. The interactions of cDC precursors with their BM niche will be analysed in steady-state or inflammatory conditions by visualising the cells in situ. Finally, the consequences of demand-driven cDCpoiesis for immunity will be assessed. The results from this project will lead to a greater understanding of the influence of environmental signals on cDCpoiesis and may have applications in the design of better vaccines and immunotherapies.
Summary
Classical dendritic cells (cDCs) are leucocytes that play a key role in innate immunity as well as the initiation and regulation of T cell responses. cDCpoiesis starts with commitment of a bone marrow (BM) haematopoietic progenitor, known as the classical DC precursor (CDP), to the cDC lineage. CDPs then give rise to pre-cDCs that exit the BM via the blood and seed tissues to give rise to the two major types of fully-differentiated cDCs, the cDC1 and cDC2 subsets. The key parameters of cDCpoiesis are poorly understood. We propose to characterise the niche in which cDCs develop within the BM and to study how pre-cDCs seed tissues and establish local clones of differentiated cDC1 and cDC2. We further wish to ask how the activity of CDPs and pre-cDCs is altered following infection, inflammation or tissue damage. Finally, we want to know to what extent cDCpoiesis is affected by direct sensing of infection or cell damage by cDC precursors. All these objectives will be addressed in a mouse lineage tracing model in which cDC precursors are genetically labelled through the activity of a Cre recombinase driven by the Clec9a locus. These mice will be crossed to fluorescent protein reporter mice, including Confetti mice that allow for clonal analysis, and the appearance of labelled cDCs and cDC clones in tissues will be followed over time in the steady-state or after induction of infection or inflammation. The dependence of cDC precursor activity on specific pathogen and damage sensing pathways will be assessed by loss-of-function experiments. The interactions of cDC precursors with their BM niche will be analysed in steady-state or inflammatory conditions by visualising the cells in situ. Finally, the consequences of demand-driven cDCpoiesis for immunity will be assessed. The results from this project will lead to a greater understanding of the influence of environmental signals on cDCpoiesis and may have applications in the design of better vaccines and immunotherapies.
Max ERC Funding
2 500 000 €
Duration
Start date: 2018-09-01, End date: 2023-08-31
Project acronym DecodingInfection
Project Decoding the host-pathogen interspecies crosstalk at a multiparametric single-cell level
Researcher (PI) Roi AVRAHAM
Host Institution (HI) WEIZMANN INSTITUTE OF SCIENCE
Call Details Starting Grant (StG), LS6, ERC-2017-STG
Summary Bacterial pathogens remain a significant threat to global health, necessitating a better understanding of host-pathogen biology. While various evidence point to early infection as a key event in the eventual progression to disease, our recent preliminary data show that during this stage, highly adaptable and dynamic host cells and bacteria engage in complex, diverse interactions that contribute to well-documented heterogeneous outcomes of infection. However, current methodologies rely on measurements of bulk populations, thereby overlooking this diversity that can trigger different outcomes. This application focuses on understanding heterogeneity during the first stages of infection in order to reduce the complexity of these interactions into informative readouts of population physiology and predictors of infection outcome. We will apply multiparametric single-cell analysis to obtain an accurate and complete description of infection with the enteric intracellular pathogen Salmonella of macrophages in vitro, and in early stages of mice colonization. We will characterize the molecular details that underlie distinct infection outcomes of individual encounters, to reconstruct the repertoire of host and pathogen strategies that prevail at critical stages of early infection.
We propose the following three objectives: (1) Develop methodologies to simultaneously profile host and pathogen transcriptional changes on a single cell level; 2) Characterizing the molecular details that underlie the formation of subpopulations during macrophage infection; and (3) Determine how host and pathogen encounters in vivo result in emergence of specialized subpopulations, recruitment of immune cells and pathogen dissemination.
We anticipate that this work will fundamentally shift our paradigms of infectious disease pathogenesis and lay the groundwork for the development of a new generation of therapeutic agents targeting the specific host-pathogen interactions ultimately driving disease.
Summary
Bacterial pathogens remain a significant threat to global health, necessitating a better understanding of host-pathogen biology. While various evidence point to early infection as a key event in the eventual progression to disease, our recent preliminary data show that during this stage, highly adaptable and dynamic host cells and bacteria engage in complex, diverse interactions that contribute to well-documented heterogeneous outcomes of infection. However, current methodologies rely on measurements of bulk populations, thereby overlooking this diversity that can trigger different outcomes. This application focuses on understanding heterogeneity during the first stages of infection in order to reduce the complexity of these interactions into informative readouts of population physiology and predictors of infection outcome. We will apply multiparametric single-cell analysis to obtain an accurate and complete description of infection with the enteric intracellular pathogen Salmonella of macrophages in vitro, and in early stages of mice colonization. We will characterize the molecular details that underlie distinct infection outcomes of individual encounters, to reconstruct the repertoire of host and pathogen strategies that prevail at critical stages of early infection.
We propose the following three objectives: (1) Develop methodologies to simultaneously profile host and pathogen transcriptional changes on a single cell level; 2) Characterizing the molecular details that underlie the formation of subpopulations during macrophage infection; and (3) Determine how host and pathogen encounters in vivo result in emergence of specialized subpopulations, recruitment of immune cells and pathogen dissemination.
We anticipate that this work will fundamentally shift our paradigms of infectious disease pathogenesis and lay the groundwork for the development of a new generation of therapeutic agents targeting the specific host-pathogen interactions ultimately driving disease.
Max ERC Funding
1 499 999 €
Duration
Start date: 2017-10-01, End date: 2022-09-30
Project acronym DrySeasonPf
Project Dry season P. falciparum reservoir
Researcher (PI) Silvia VILAR PORTUGAL
Host Institution (HI) UNIVERSITATSKLINIKUM HEIDELBERG
Call Details Starting Grant (StG), LS6, ERC-2017-STG
Summary The mosquito-borne Plasmodium falciparum parasite is responsible for over 200 million malaria cases and nearly half a million deaths each year among African children. Dependent on Anopheles mosquito for transmission, the parasite faces a challenge during the dry season in the regions where rain seasonality limits vector availability for several months. While malaria cases are restricted to the wet season, clinically silent P. falciparum infections can persist through the dry season and are an important reservoir for transmission. Our preliminary data provides unequivocal evidence that P. falciparum modulates its transcription during the dry season, while the host immune response seems to be minimally affected, suggesting that the parasite has the ability to adapt to a vector-free environment for long periods of time. Understanding the mechanisms which allow the parasite to remain undetectable in absence of mosquito vector, and to restart transmission in the ensuing rainy season will reveal complex interactions between P. falciparum and its host. To that end I propose to: (i) Identify the Plasmodium signalling pathway(s) and metabolic profile associated with long-term maintenance of low parasitaemias during the dry season, (ii) Determine which PfEMP1 are expressed by parasites during the dry season and how effectively they are detected by the immune system, and (iii) Investigate the kinetics of P. falciparum gametocytogenesis, its ability to transmit during the dry season, and uncover sensing molecules and mechanisms of the disappearance and return of the mosquito vector Undoubtedly, results arising from the present multidisciplinary proposal will provide novel insights into the cell biology of dry season P. falciparum parasites, will increase our understanding of their interactions with their hosts and environment. Furthermore, it may benefit the international development agenda goals to design public health strategies to fight malaria.
Summary
The mosquito-borne Plasmodium falciparum parasite is responsible for over 200 million malaria cases and nearly half a million deaths each year among African children. Dependent on Anopheles mosquito for transmission, the parasite faces a challenge during the dry season in the regions where rain seasonality limits vector availability for several months. While malaria cases are restricted to the wet season, clinically silent P. falciparum infections can persist through the dry season and are an important reservoir for transmission. Our preliminary data provides unequivocal evidence that P. falciparum modulates its transcription during the dry season, while the host immune response seems to be minimally affected, suggesting that the parasite has the ability to adapt to a vector-free environment for long periods of time. Understanding the mechanisms which allow the parasite to remain undetectable in absence of mosquito vector, and to restart transmission in the ensuing rainy season will reveal complex interactions between P. falciparum and its host. To that end I propose to: (i) Identify the Plasmodium signalling pathway(s) and metabolic profile associated with long-term maintenance of low parasitaemias during the dry season, (ii) Determine which PfEMP1 are expressed by parasites during the dry season and how effectively they are detected by the immune system, and (iii) Investigate the kinetics of P. falciparum gametocytogenesis, its ability to transmit during the dry season, and uncover sensing molecules and mechanisms of the disappearance and return of the mosquito vector Undoubtedly, results arising from the present multidisciplinary proposal will provide novel insights into the cell biology of dry season P. falciparum parasites, will increase our understanding of their interactions with their hosts and environment. Furthermore, it may benefit the international development agenda goals to design public health strategies to fight malaria.
Max ERC Funding
1 500 000 €
Duration
Start date: 2018-01-01, End date: 2022-12-31
Project acronym DynaMO_TB
Project Spatiotemporal regulation of localization and replication of M. tuberculosis in humanmacrophages
Researcher (PI) Maximiliano Gabriel GUTIERREZ
Host Institution (HI) THE FRANCIS CRICK INSTITUTE LIMITED
Call Details Consolidator Grant (CoG), LS6, ERC-2017-COG
Summary Mycobacterium tuberculosis (Mtb) is a very successful intracellular pathogen: in 2014, tuberculosis (TB) caused 1.5 million human deaths (World Health Organisation). To cause disease and disseminate to other hosts, Mtb needs to replicate within human cells. In spite of its enormous relevance for TB pathogenesis, the precise sites of Mtb replication in host cells remain unknown. This surprising gap in knowledge is in part due to the lack of appropriate imaging technologies that have precluded comprehensive understanding of the fundamental biology that underpins Mtb-host cell interactions critical to design rational interventions. Here, we propose to use a series of cutting-edge imaging approaches in human macrophages to: (1) define how the dynamic interactions between Mtb populations and organelles impact Mtb replication; (2) identify critical host and bacterial components that regulate Mtb replication and (3) characterise the host cell death pathways that control Mtb replication. For this, we will benefit from technologies developed in our group to image and quantify Mtb localisation and replication, such as live cell imaging, super resolution (SR) microscopy and correlative live cell 3D- electron microscopy (CLEM). We will refine these approaches to challenge the current limits of cell-based, high content imaging by combining human stem cell-derived macrophages with adhesive micropattern technologies for single cell analysis; this allows us to identify where and when Mtb replicate and how the interplay between host cells and Mtb impacts this process. Together, this proposal can uncover novel cellular pathways defining the intracellular sites that allow or restrict Mtb replication in human macrophages, thereby advancing the fields of both cell and infection biology. The characterization of the site of intracellular replication of Mtb can open avenues for a deeper understanding of human TB pathogenesis and facilitate development of vaccines and antibioo be here soon
Summary
Mycobacterium tuberculosis (Mtb) is a very successful intracellular pathogen: in 2014, tuberculosis (TB) caused 1.5 million human deaths (World Health Organisation). To cause disease and disseminate to other hosts, Mtb needs to replicate within human cells. In spite of its enormous relevance for TB pathogenesis, the precise sites of Mtb replication in host cells remain unknown. This surprising gap in knowledge is in part due to the lack of appropriate imaging technologies that have precluded comprehensive understanding of the fundamental biology that underpins Mtb-host cell interactions critical to design rational interventions. Here, we propose to use a series of cutting-edge imaging approaches in human macrophages to: (1) define how the dynamic interactions between Mtb populations and organelles impact Mtb replication; (2) identify critical host and bacterial components that regulate Mtb replication and (3) characterise the host cell death pathways that control Mtb replication. For this, we will benefit from technologies developed in our group to image and quantify Mtb localisation and replication, such as live cell imaging, super resolution (SR) microscopy and correlative live cell 3D- electron microscopy (CLEM). We will refine these approaches to challenge the current limits of cell-based, high content imaging by combining human stem cell-derived macrophages with adhesive micropattern technologies for single cell analysis; this allows us to identify where and when Mtb replicate and how the interplay between host cells and Mtb impacts this process. Together, this proposal can uncover novel cellular pathways defining the intracellular sites that allow or restrict Mtb replication in human macrophages, thereby advancing the fields of both cell and infection biology. The characterization of the site of intracellular replication of Mtb can open avenues for a deeper understanding of human TB pathogenesis and facilitate development of vaccines and antibioo be here soon
Max ERC Funding
2 000 000 €
Duration
Start date: 2018-09-01, End date: 2023-08-31
