Project acronym 3DCellPhase-
Project In situ Structural Analysis of Molecular Crowding and Phase Separation
Researcher (PI) Julia MAHAMID
Host Institution (HI) EUROPEAN MOLECULAR BIOLOGY LABORATORY
Call Details Starting Grant (StG), LS1, ERC-2017-STG
Summary This proposal brings together two fields in biology, namely the emerging field of phase-separated assemblies in cell biology and state-of-the-art cellular cryo-electron tomography, to advance our understanding on a fundamental, yet illusive, question: the molecular organization of the cytoplasm.
Eukaryotes organize their biochemical reactions into functionally distinct compartments. Intriguingly, many, if not most, cellular compartments are not membrane enclosed. Rather, they assemble dynamically by phase separation, typically triggered upon a specific event. Despite significant progress on reconstituting such liquid-like assemblies in vitro, we lack information as to whether these compartments in vivo are indeed amorphous liquids, or whether they exhibit structural features such as gels or fibers. My recent work on sample preparation of cells for cryo-electron tomography, including cryo-focused ion beam thinning, guided by 3D correlative fluorescence microscopy, shows that we can now prepare site-specific ‘electron-transparent windows’ in suitable eukaryotic systems, which allow direct examination of structural features of cellular compartments in their cellular context. Here, we will use these techniques to elucidate the structural principles and cytoplasmic environment driving the dynamic assembly of two phase-separated compartments: Stress granules, which are RNA bodies that form rapidly in the cytoplasm upon cellular stress, and centrosomes, which are sites of microtubule nucleation. We will combine these studies with a quantitative description of the crowded nature of cytoplasm and of its local variations, to provide a direct readout of the impact of excluded volume on molecular assembly in living cells. Taken together, these studies will provide fundamental insights into the structural basis by which cells form biochemical compartments.
Summary
This proposal brings together two fields in biology, namely the emerging field of phase-separated assemblies in cell biology and state-of-the-art cellular cryo-electron tomography, to advance our understanding on a fundamental, yet illusive, question: the molecular organization of the cytoplasm.
Eukaryotes organize their biochemical reactions into functionally distinct compartments. Intriguingly, many, if not most, cellular compartments are not membrane enclosed. Rather, they assemble dynamically by phase separation, typically triggered upon a specific event. Despite significant progress on reconstituting such liquid-like assemblies in vitro, we lack information as to whether these compartments in vivo are indeed amorphous liquids, or whether they exhibit structural features such as gels or fibers. My recent work on sample preparation of cells for cryo-electron tomography, including cryo-focused ion beam thinning, guided by 3D correlative fluorescence microscopy, shows that we can now prepare site-specific ‘electron-transparent windows’ in suitable eukaryotic systems, which allow direct examination of structural features of cellular compartments in their cellular context. Here, we will use these techniques to elucidate the structural principles and cytoplasmic environment driving the dynamic assembly of two phase-separated compartments: Stress granules, which are RNA bodies that form rapidly in the cytoplasm upon cellular stress, and centrosomes, which are sites of microtubule nucleation. We will combine these studies with a quantitative description of the crowded nature of cytoplasm and of its local variations, to provide a direct readout of the impact of excluded volume on molecular assembly in living cells. Taken together, these studies will provide fundamental insights into the structural basis by which cells form biochemical compartments.
Max ERC Funding
1 228 125 €
Duration
Start date: 2018-02-01, End date: 2023-01-31
Project acronym 3DEpi
Project Transgenerational epigenetic inheritance of chromatin states : the role of Polycomb and 3D chromosome architecture
Researcher (PI) Giacomo CAVALLI
Host Institution (HI) CENTRE NATIONAL DE LA RECHERCHE SCIENTIFIQUE CNRS
Call Details Advanced Grant (AdG), LS2, ERC-2017-ADG
Summary Epigenetic inheritance entails transmission of phenotypic traits not encoded in the DNA sequence and, in the most extreme case, Transgenerational Epigenetic Inheritance (TEI) involves transmission of memory through multiple generations. Very little is known on the mechanisms governing TEI and this is the subject of the present proposal. By transiently enhancing long-range chromatin interactions, we recently established isogenic Drosophila epilines that carry stable alternative epialleles, defined by differential levels of the Polycomb-dependent H3K27me3 mark. Furthermore, we extended our paradigm to natural phenotypes. These are ideal systems to study the role of Polycomb group (PcG) proteins and other components in regulating nuclear organization and epigenetic inheritance of chromatin states. The present project conjugates genetics, epigenomics, imaging and molecular biology to reach three critical aims.
Aim 1: Analysis of the molecular mechanisms regulating Polycomb-mediated TEI. We will identify the DNA, protein and RNA components that trigger and maintain transgenerational chromatin inheritance as well as their mechanisms of action.
Aim 2: Role of 3D genome organization in the regulation of TEI. We will analyze the developmental dynamics of TEI-inducing long-range chromatin interactions, identify chromatin components mediating 3D chromatin contacts and characterize their function in the TEI process.
Aim 3: Identification of a broader role of TEI during development. TEI might reflect a normal role of PcG components in the transmission of parental chromatin onto the next embryonic generation. We will explore this possibility by establishing other TEI paradigms and by relating TEI to the normal PcG function in these systems and in normal development.
This research program will unravel the biological significance and the molecular underpinnings of TEI and lead the way towards establishing this area of research into a consolidated scientific discipline.
Summary
Epigenetic inheritance entails transmission of phenotypic traits not encoded in the DNA sequence and, in the most extreme case, Transgenerational Epigenetic Inheritance (TEI) involves transmission of memory through multiple generations. Very little is known on the mechanisms governing TEI and this is the subject of the present proposal. By transiently enhancing long-range chromatin interactions, we recently established isogenic Drosophila epilines that carry stable alternative epialleles, defined by differential levels of the Polycomb-dependent H3K27me3 mark. Furthermore, we extended our paradigm to natural phenotypes. These are ideal systems to study the role of Polycomb group (PcG) proteins and other components in regulating nuclear organization and epigenetic inheritance of chromatin states. The present project conjugates genetics, epigenomics, imaging and molecular biology to reach three critical aims.
Aim 1: Analysis of the molecular mechanisms regulating Polycomb-mediated TEI. We will identify the DNA, protein and RNA components that trigger and maintain transgenerational chromatin inheritance as well as their mechanisms of action.
Aim 2: Role of 3D genome organization in the regulation of TEI. We will analyze the developmental dynamics of TEI-inducing long-range chromatin interactions, identify chromatin components mediating 3D chromatin contacts and characterize their function in the TEI process.
Aim 3: Identification of a broader role of TEI during development. TEI might reflect a normal role of PcG components in the transmission of parental chromatin onto the next embryonic generation. We will explore this possibility by establishing other TEI paradigms and by relating TEI to the normal PcG function in these systems and in normal development.
This research program will unravel the biological significance and the molecular underpinnings of TEI and lead the way towards establishing this area of research into a consolidated scientific discipline.
Max ERC Funding
2 500 000 €
Duration
Start date: 2018-11-01, End date: 2023-10-31
Project acronym 3DPROTEINPUZZLES
Project Shape-directed protein assembly design
Researcher (PI) Lars Ingemar ANDRÉ
Host Institution (HI) LUNDS UNIVERSITET
Call Details Consolidator Grant (CoG), LS9, ERC-2017-COG
Summary Large protein complexes carry out some of the most complex functions in biology. Such structures are often assembled spontaneously from individual components through the process of self-assembly. If self-assembled protein complexes could be engineered from first principle it would enable a wide range of applications in biomedicine, nanotechnology and materials science. Recently, approaches to rationally design proteins to self-assembly into predefined structures have emerged. The highlight of this work is the design of protein cages that may be engineered into protein containers. However, current approaches for self-assembly design does not result in the assemblies with the required structural complexity to encode many of the sophisticated functions found in nature. To move forward, we have to learn how to engineer protein subunits with more than one designed interface that can assemble into tightly interacting complexes. In this proposal we propose a new protein design paradigm, shape directed protein design, in order to address shortcomings of the current methodology. The proposed method combines geometric shape matching and computational protein design. Using this approach we will de novo design assemblies with a wide variety of structural states, including protein complexes with cyclic and dihedral symmetry as well as icosahedral protein capsids built from novel protein building blocks. To enable these two design challenges we also develop a high-throughput assay to measure assembly stability in vivo that builds on a three-color fluorescent assay. This method will not only facilitate the screening of orders of magnitude more design constructs, but also enable the application of directed evolution to experimentally improve stable and assembly properties of designed containers as well as other designed assemblies.
Summary
Large protein complexes carry out some of the most complex functions in biology. Such structures are often assembled spontaneously from individual components through the process of self-assembly. If self-assembled protein complexes could be engineered from first principle it would enable a wide range of applications in biomedicine, nanotechnology and materials science. Recently, approaches to rationally design proteins to self-assembly into predefined structures have emerged. The highlight of this work is the design of protein cages that may be engineered into protein containers. However, current approaches for self-assembly design does not result in the assemblies with the required structural complexity to encode many of the sophisticated functions found in nature. To move forward, we have to learn how to engineer protein subunits with more than one designed interface that can assemble into tightly interacting complexes. In this proposal we propose a new protein design paradigm, shape directed protein design, in order to address shortcomings of the current methodology. The proposed method combines geometric shape matching and computational protein design. Using this approach we will de novo design assemblies with a wide variety of structural states, including protein complexes with cyclic and dihedral symmetry as well as icosahedral protein capsids built from novel protein building blocks. To enable these two design challenges we also develop a high-throughput assay to measure assembly stability in vivo that builds on a three-color fluorescent assay. This method will not only facilitate the screening of orders of magnitude more design constructs, but also enable the application of directed evolution to experimentally improve stable and assembly properties of designed containers as well as other designed assemblies.
Max ERC Funding
2 325 292 €
Duration
Start date: 2018-06-01, End date: 2023-05-31
Project acronym 4D-GenEx
Project Spatio-temporal Organization and Expression of the Genome
Researcher (PI) Antoine COULON
Host Institution (HI) CENTRE NATIONAL DE LA RECHERCHE SCIENTIFIQUE CNRS
Call Details Starting Grant (StG), LS2, ERC-2017-STG
Summary This project investigates the two-way relationship between spatio-temporal genome organization and coordinated gene regulation, through an approach at the interface between physics, computer science and biology.
In the nucleus, preferred positions are observed from chromosomes to single genes, in relation to normal and pathological cellular states. Evidence indicates a complex spatio-temporal coupling between co-regulated genes: e.g. certain genes cluster spatially when responding to similar factors and transcriptional noise patterns suggest domain-wide mechanisms. Yet, no individual experiment allows probing transcriptional coordination in 4 dimensions (FISH, live locus tracking, Hi-C...). Interpreting such data also critically requires theory (stochastic processes, statistical physics…). A lack of appropriate experimental/analytical approaches is impairing our understanding of the 4D genome.
Our proposal combines cutting-edge single-molecule imaging, signal-theory data analysis and physical modeling to study how genes coordinate in space and time in a single nucleus. Our objectives are to understand (a) competition/recycling of shared resources between genes within subnuclear compartments, (b) how enhancers communicate with genes domain-wide, and (c) the role of local conformational dynamics and supercoiling in gene co-regulation. Our organizing hypothesis is that, by acting on their microenvironment, genes shape their co-expression with other genes.
Building upon my expertise, we will use dual-color MS2/PP7 RNA labeling to visualize for the first time transcription and motion of pairs of hormone-responsive genes in real time. With our innovative signal analysis tools, we will extract spatio-temporal signatures of underlying processes, which we will investigate with stochastic modeling and validate through experimental perturbations. We expect to uncover how the functional organization of the linear genome relates to its physical properties and dynamics in 4D.
Summary
This project investigates the two-way relationship between spatio-temporal genome organization and coordinated gene regulation, through an approach at the interface between physics, computer science and biology.
In the nucleus, preferred positions are observed from chromosomes to single genes, in relation to normal and pathological cellular states. Evidence indicates a complex spatio-temporal coupling between co-regulated genes: e.g. certain genes cluster spatially when responding to similar factors and transcriptional noise patterns suggest domain-wide mechanisms. Yet, no individual experiment allows probing transcriptional coordination in 4 dimensions (FISH, live locus tracking, Hi-C...). Interpreting such data also critically requires theory (stochastic processes, statistical physics…). A lack of appropriate experimental/analytical approaches is impairing our understanding of the 4D genome.
Our proposal combines cutting-edge single-molecule imaging, signal-theory data analysis and physical modeling to study how genes coordinate in space and time in a single nucleus. Our objectives are to understand (a) competition/recycling of shared resources between genes within subnuclear compartments, (b) how enhancers communicate with genes domain-wide, and (c) the role of local conformational dynamics and supercoiling in gene co-regulation. Our organizing hypothesis is that, by acting on their microenvironment, genes shape their co-expression with other genes.
Building upon my expertise, we will use dual-color MS2/PP7 RNA labeling to visualize for the first time transcription and motion of pairs of hormone-responsive genes in real time. With our innovative signal analysis tools, we will extract spatio-temporal signatures of underlying processes, which we will investigate with stochastic modeling and validate through experimental perturbations. We expect to uncover how the functional organization of the linear genome relates to its physical properties and dynamics in 4D.
Max ERC Funding
1 499 750 €
Duration
Start date: 2018-04-01, End date: 2023-03-31
Project acronym Acclimatize
Project Hypothalamic mechanisms of thermal homeostasis and adaptation
Researcher (PI) Jan SIEMENS
Host Institution (HI) UNIVERSITATSKLINIKUM HEIDELBERG
Call Details Consolidator Grant (CoG), LS5, ERC-2017-COG
Summary Mammalian organisms possess the remarkable ability to maintain internal body temperature (Tcore) within a narrow range close to 37°C despite wide environmental temperature variations. The brain’s neural “thermostat” is made up by central circuits in the hypothalamic preoptic area (POA), which orchestrate peripheral thermoregulatory responses to maintain Tcore. Thermogenesis requires metabolic fuel, suggesting intricate connections between the thermoregulatory centre and hypothalamic circuits controlling energy balance. How the POA detects and integrates temperature and metabolic information to achieve thermal balance is largely unknown. A major question is whether this circuitry could be harnessed therapeutically to treat obesity.
We have recently identified the first known molecular temperature sensor in thermoregulatory neurons of the POA, transient receptor potential melastatin 2 (TRPM2), a thermo-sensitive ion channel. I aim to use TRPM2 as a molecular marker to gain access to and probe the function of thermoregulatory neurons in vivo. I propose a multidisciplinary approach, combining local, in vivo POA temperature stimulation with optogenetic circuit-mapping to uncover the molecular and cellular logic of the hypothalamic thermoregulatory centre and to assess its medical potential to counteract metabolic syndrome.
Acclimation is a beneficial adaptive process that fortifies thermal responses upon environmental temperature challenges. Thermoregulatory neuron plasticity is thought to mediate acclimation. Conversely, maladaptive thermoregulatory changes affect obesity. The cell-type-specific neuronal plasticity mechanisms underlying these changes within the POA, however, are unknown.
Using ex-vivo slice electrophysiology and in vivo imaging, I propose to characterize acclimation- and obesity-induced plasticity of thermoregulatory neurons. Ultimately, I aim to manipulate thermoregulatory neuron plasticity to test its potential counter-balancing effect on obesity.
Summary
Mammalian organisms possess the remarkable ability to maintain internal body temperature (Tcore) within a narrow range close to 37°C despite wide environmental temperature variations. The brain’s neural “thermostat” is made up by central circuits in the hypothalamic preoptic area (POA), which orchestrate peripheral thermoregulatory responses to maintain Tcore. Thermogenesis requires metabolic fuel, suggesting intricate connections between the thermoregulatory centre and hypothalamic circuits controlling energy balance. How the POA detects and integrates temperature and metabolic information to achieve thermal balance is largely unknown. A major question is whether this circuitry could be harnessed therapeutically to treat obesity.
We have recently identified the first known molecular temperature sensor in thermoregulatory neurons of the POA, transient receptor potential melastatin 2 (TRPM2), a thermo-sensitive ion channel. I aim to use TRPM2 as a molecular marker to gain access to and probe the function of thermoregulatory neurons in vivo. I propose a multidisciplinary approach, combining local, in vivo POA temperature stimulation with optogenetic circuit-mapping to uncover the molecular and cellular logic of the hypothalamic thermoregulatory centre and to assess its medical potential to counteract metabolic syndrome.
Acclimation is a beneficial adaptive process that fortifies thermal responses upon environmental temperature challenges. Thermoregulatory neuron plasticity is thought to mediate acclimation. Conversely, maladaptive thermoregulatory changes affect obesity. The cell-type-specific neuronal plasticity mechanisms underlying these changes within the POA, however, are unknown.
Using ex-vivo slice electrophysiology and in vivo imaging, I propose to characterize acclimation- and obesity-induced plasticity of thermoregulatory neurons. Ultimately, I aim to manipulate thermoregulatory neuron plasticity to test its potential counter-balancing effect on obesity.
Max ERC Funding
1 902 500 €
Duration
Start date: 2018-09-01, End date: 2023-08-31
Project acronym activeFly
Project Circuit mechanisms of self-movement estimation during walking
Researcher (PI) M Eugenia CHIAPPE
Host Institution (HI) FUNDACAO D. ANNA SOMMER CHAMPALIMAUD E DR. CARLOS MONTEZ CHAMPALIMAUD
Call Details Starting Grant (StG), LS5, ERC-2017-STG
Summary The brain evolves, develops, and operates in the context of animal movements. As a consequence, fundamental brain functions such as spatial perception and motor control critically depend on the precise knowledge of the ongoing body motion. An accurate internal estimate of self-movement is thought to emerge from sensorimotor integration; nonetheless, which circuits perform this internal estimation, and exactly how motor-sensory coordination is implemented within these circuits are basic questions that remain to be poorly understood. There is growing evidence suggesting that, during locomotion, motor-related and visual signals interact at early stages of visual processing. In mammals, however, it is not clear what the function of this interaction is. Recently, we have shown that a population of Drosophila optic-flow processing neurons —neurons that are sensitive to self-generated visual flow, receives convergent visual and walking-related signals to form a faithful representation of the fly’s walking movements. Leveraging from these results, and combining quantitative analysis of behavior with physiology, optogenetics, and modelling, we propose to investigate circuit mechanisms of self-movement estimation during walking. We will:1) use cell specific manipulations to identify what cells are necessary to generate the motor-related activity in the population of visual neurons, 2) record from the identified neurons and correlate their activity with specific locomotor parameters, and 3) perturb the activity of different cell-types within the identified circuits to test their role in the dynamics of the visual neurons, and on the fly’s walking behavior. These experiments will establish unprecedented causal relationships among neural activity, the formation of an internal representation, and locomotor control. The identified sensorimotor principles will establish a framework that can be tested in other scenarios or animal systems with implications both in health and disease.
Summary
The brain evolves, develops, and operates in the context of animal movements. As a consequence, fundamental brain functions such as spatial perception and motor control critically depend on the precise knowledge of the ongoing body motion. An accurate internal estimate of self-movement is thought to emerge from sensorimotor integration; nonetheless, which circuits perform this internal estimation, and exactly how motor-sensory coordination is implemented within these circuits are basic questions that remain to be poorly understood. There is growing evidence suggesting that, during locomotion, motor-related and visual signals interact at early stages of visual processing. In mammals, however, it is not clear what the function of this interaction is. Recently, we have shown that a population of Drosophila optic-flow processing neurons —neurons that are sensitive to self-generated visual flow, receives convergent visual and walking-related signals to form a faithful representation of the fly’s walking movements. Leveraging from these results, and combining quantitative analysis of behavior with physiology, optogenetics, and modelling, we propose to investigate circuit mechanisms of self-movement estimation during walking. We will:1) use cell specific manipulations to identify what cells are necessary to generate the motor-related activity in the population of visual neurons, 2) record from the identified neurons and correlate their activity with specific locomotor parameters, and 3) perturb the activity of different cell-types within the identified circuits to test their role in the dynamics of the visual neurons, and on the fly’s walking behavior. These experiments will establish unprecedented causal relationships among neural activity, the formation of an internal representation, and locomotor control. The identified sensorimotor principles will establish a framework that can be tested in other scenarios or animal systems with implications both in health and disease.
Max ERC Funding
1 500 000 €
Duration
Start date: 2017-11-01, End date: 2022-10-31
Project acronym AdaptiveResponse
Project The evolution of adaptive response mechanisms
Researcher (PI) Franz WEISSING
Host Institution (HI) RIJKSUNIVERSITEIT GRONINGEN
Call Details Advanced Grant (AdG), LS8, ERC-2017-ADG
Summary In an era of rapid climate change there is a pressing need to understand whether and how organisms are able to adapt to novel environments. Such understanding is hampered by a major divide in the life sciences. Disciplines like systems biology or neurobiology make rapid progress in unravelling the mechanisms underlying the responses of organisms to their environment, but this knowledge is insufficiently integrated in eco-evolutionary theory. Current eco-evolutionary models focus on the response patterns themselves, largely neglecting the structures and mechanisms producing these patterns. Here I propose a new, mechanism-oriented framework that views the architecture of adaptation, rather than the resulting responses, as the primary target of natural selection. I am convinced that this change in perspective will yield fundamentally new insights, necessitating the re-evaluation of many seemingly well-established eco-evolutionary principles.
My aim is to develop a comprehensive theory of the eco-evolutionary causes and consequences of the architecture underlying adaptive responses. In three parallel lines of investigation, I will study how architecture is shaped by selection, how evolved response strategies reflect the underlying architecture, and how these responses affect the eco-evolutionary dynamics and the capacity to adapt to novel conditions. All three lines have the potential of making ground-breaking contributions to eco-evolutionary theory, including: the specification of evolutionary tipping points; resolving the puzzle that real organisms evolve much faster than predicted by current theory; a new and general explanation for the evolutionary emergence of individual variation; and a framework for studying the evolution of learning and other general-purpose mechanisms. By making use of concepts from information theory and artificial intelligence, the project will also introduce various methodological innovations.
Summary
In an era of rapid climate change there is a pressing need to understand whether and how organisms are able to adapt to novel environments. Such understanding is hampered by a major divide in the life sciences. Disciplines like systems biology or neurobiology make rapid progress in unravelling the mechanisms underlying the responses of organisms to their environment, but this knowledge is insufficiently integrated in eco-evolutionary theory. Current eco-evolutionary models focus on the response patterns themselves, largely neglecting the structures and mechanisms producing these patterns. Here I propose a new, mechanism-oriented framework that views the architecture of adaptation, rather than the resulting responses, as the primary target of natural selection. I am convinced that this change in perspective will yield fundamentally new insights, necessitating the re-evaluation of many seemingly well-established eco-evolutionary principles.
My aim is to develop a comprehensive theory of the eco-evolutionary causes and consequences of the architecture underlying adaptive responses. In three parallel lines of investigation, I will study how architecture is shaped by selection, how evolved response strategies reflect the underlying architecture, and how these responses affect the eco-evolutionary dynamics and the capacity to adapt to novel conditions. All three lines have the potential of making ground-breaking contributions to eco-evolutionary theory, including: the specification of evolutionary tipping points; resolving the puzzle that real organisms evolve much faster than predicted by current theory; a new and general explanation for the evolutionary emergence of individual variation; and a framework for studying the evolution of learning and other general-purpose mechanisms. By making use of concepts from information theory and artificial intelligence, the project will also introduce various methodological innovations.
Max ERC Funding
2 500 000 €
Duration
Start date: 2018-12-01, End date: 2023-11-30
Project acronym AGRISCENTS
Project Scents and sensibility in agriculture: exploiting specificity in herbivore- and pathogen-induced plant volatiles for real-time crop monitoring
Researcher (PI) Theodoor Turlings
Host Institution (HI) UNIVERSITE DE NEUCHATEL
Call Details Advanced Grant (AdG), LS9, ERC-2017-ADG
Summary Plants typically release large quantities of volatiles in response to attack by herbivores or pathogens. I may claim to have contributed to various breakthroughs in this research field, including the discovery that the volatile blends induced by different attackers are astonishingly specific, resulting in characteristic, readily distinguishable odour blends. Using maize as our model plant, I wish to take several leaps forward in our understanding of this signal specificity and use this knowledge to develop sensors for the real-time detection of crop pests and diseases. For this, three interconnected work-packages will aim to:
• Develop chemical analytical techniques and statistical models to decipher the odorous vocabulary of plants, and to create a complete inventory of “odour-prints” for a wide range of herbivore-plant and pathogen-plant combinations, including simultaneous infestations.
• Develop and optimize nano-mechanical sensors for the detection of specific plant volatile mixtures. For this, we will initially adapt a prototype sensor that has been successfully developed for the detection of cancer-related volatiles in human breath.
• Genetically manipulate maize plants to release a unique blend of root-produced volatiles upon herbivory. For this, we will engineer gene cassettes that combine recently identified P450 (CYP) genes from poplar with inducible, root-specific promoters from maize. This will result in maize plants that, in response to pest attack, release easy-to-detect aldoximes and nitriles from their roots.
In short, by investigating and manipulating the specificity of inducible odour blends we will generate the necessary knowhow to develop a novel odour-detection device. The envisioned sensor technology will permit real-time monitoring of the pests and enable farmers to apply crop protection treatments at the right time and in the right place.
Summary
Plants typically release large quantities of volatiles in response to attack by herbivores or pathogens. I may claim to have contributed to various breakthroughs in this research field, including the discovery that the volatile blends induced by different attackers are astonishingly specific, resulting in characteristic, readily distinguishable odour blends. Using maize as our model plant, I wish to take several leaps forward in our understanding of this signal specificity and use this knowledge to develop sensors for the real-time detection of crop pests and diseases. For this, three interconnected work-packages will aim to:
• Develop chemical analytical techniques and statistical models to decipher the odorous vocabulary of plants, and to create a complete inventory of “odour-prints” for a wide range of herbivore-plant and pathogen-plant combinations, including simultaneous infestations.
• Develop and optimize nano-mechanical sensors for the detection of specific plant volatile mixtures. For this, we will initially adapt a prototype sensor that has been successfully developed for the detection of cancer-related volatiles in human breath.
• Genetically manipulate maize plants to release a unique blend of root-produced volatiles upon herbivory. For this, we will engineer gene cassettes that combine recently identified P450 (CYP) genes from poplar with inducible, root-specific promoters from maize. This will result in maize plants that, in response to pest attack, release easy-to-detect aldoximes and nitriles from their roots.
In short, by investigating and manipulating the specificity of inducible odour blends we will generate the necessary knowhow to develop a novel odour-detection device. The envisioned sensor technology will permit real-time monitoring of the pests and enable farmers to apply crop protection treatments at the right time and in the right place.
Max ERC Funding
2 498 086 €
Duration
Start date: 2018-09-01, End date: 2023-08-31
Project acronym AltCheM
Project In vivo functional screens to decipher mechanisms of stochastically- and mutationally-induced chemoresistance in Acute Myeloid Leukemia
Researcher (PI) Alexandre PUISSANT
Host Institution (HI) INSTITUT NATIONAL DE LA SANTE ET DE LA RECHERCHE MEDICALE
Call Details Starting Grant (StG), LS4, ERC-2017-STG
Summary Acute Myeloid Leukemia (AML), the most common leukemia diagnosed in adults, represents the paradigm of resistance to front-line therapies in hematology. Indeed, AML is so genetically complex that only few targeted therapies are currently tested in this disease and chemotherapy remains the only standard treatment for AML since the past four decades. Despite an initial sustained remission achieved by chemotherapeutic agents, almost all patients relapse with a chemoresistant minimal residual disease (MRD). The goal of my proposal is to characterize the still poorly understood biological mechanisms underlying persistence and emergence of MRD.
MRD is the consequence of the re-expansion of leukemia-initiating cells that are intrinsically more resistant to chemotherapy. This cell fraction may be stochastically more prone to survive front-line therapy regardless of their mutational status (the stochastic model), or genetically predetermined to resist by virtue of a collection of chemoprotective mutations (the mutational model).
I have already generated in mice, by consecutive rounds of chemotherapy, a stochastic MLL-AF9-driven chemoresistance model that I examined by RNA-sequencing. I will pursue the comprehensive cell autonomous and cell non-autonomous characterization of this chemoresistant AML disease using whole-exome and ChIP-sequencing.
To establish a mutationally-induced chemoresistant mouse model, I will conduct an innovative in vivo screen using pooled mutant open reading frame and shRNA libraries in order to predict which combinations of mutations, among those already known in AML, actively promote chemoresistance.
Finally, by combining genomic profiling and in vivo shRNA screening experiments, I will decipher the molecular mechanisms and identify the functional effectors of these two modes of resistance. Ultimately, I will then be able to firmly establish the fundamental relevance of the stochastic and/or the mutational model of chemoresistance for MRD genesis.
Summary
Acute Myeloid Leukemia (AML), the most common leukemia diagnosed in adults, represents the paradigm of resistance to front-line therapies in hematology. Indeed, AML is so genetically complex that only few targeted therapies are currently tested in this disease and chemotherapy remains the only standard treatment for AML since the past four decades. Despite an initial sustained remission achieved by chemotherapeutic agents, almost all patients relapse with a chemoresistant minimal residual disease (MRD). The goal of my proposal is to characterize the still poorly understood biological mechanisms underlying persistence and emergence of MRD.
MRD is the consequence of the re-expansion of leukemia-initiating cells that are intrinsically more resistant to chemotherapy. This cell fraction may be stochastically more prone to survive front-line therapy regardless of their mutational status (the stochastic model), or genetically predetermined to resist by virtue of a collection of chemoprotective mutations (the mutational model).
I have already generated in mice, by consecutive rounds of chemotherapy, a stochastic MLL-AF9-driven chemoresistance model that I examined by RNA-sequencing. I will pursue the comprehensive cell autonomous and cell non-autonomous characterization of this chemoresistant AML disease using whole-exome and ChIP-sequencing.
To establish a mutationally-induced chemoresistant mouse model, I will conduct an innovative in vivo screen using pooled mutant open reading frame and shRNA libraries in order to predict which combinations of mutations, among those already known in AML, actively promote chemoresistance.
Finally, by combining genomic profiling and in vivo shRNA screening experiments, I will decipher the molecular mechanisms and identify the functional effectors of these two modes of resistance. Ultimately, I will then be able to firmly establish the fundamental relevance of the stochastic and/or the mutational model of chemoresistance for MRD genesis.
Max ERC Funding
1 500 000 €
Duration
Start date: 2018-03-01, End date: 2023-02-28
Project acronym Amitochondriates
Project Life without mitochondrion
Researcher (PI) Vladimir HAMPL
Host Institution (HI) UNIVERZITA KARLOVA
Call Details Consolidator Grant (CoG), LS8, ERC-2017-COG
Summary Mitochondria are often referred to as the “power houses” of eukaryotic cells. All eukaryotes were thought to have mitochondria of some form until 2016, when the first eukaryote thriving without mitochondria was discovered by our laboratory – a flagellate Monocercomonoides. Understanding cellular functions of these cells, which represent a new functional type of eukaryotes, and understanding the circumstances of the unique event of mitochondrial loss are motivations for this proposal. The first objective focuses on the cell physiology. We will perform a metabolomic study revealing major metabolic pathways and concentrate further on elucidating its unique system of iron-sulphur cluster assembly. In the second objective, we will investigate in details the unique case of mitochondrial loss. We will examine two additional potentially amitochondriate lineages by means of genomics and transcriptomics, conduct experiments simulating the moments of mitochondrial loss and try to induce the mitochondrial loss in vitro by knocking out or down genes for mitochondrial biogenesis. We have chosen Giardia intestinalis and Entamoeba histolytica as models for the latter experiments, because their mitochondria are already reduced to minimalistic “mitosomes” and because some genetic tools are already available for them. Successful mitochondrial knock-outs would enable us to study mitochondrial loss in ‘real time’ and in vivo. In the third objective, we will focus on transforming Monocercomonoides into a tractable laboratory model by developing methods of axenic cultivation and genetic manipulation. This will open new possibilities in the studies of this organism and create a cell culture representing an amitochondriate model for cell biological studies enabling the dissection of mitochondrial effects from those of other compartments. The team is composed of the laboratory of PI and eight invited experts and we hope it has the ability to address these challenging questions.
Summary
Mitochondria are often referred to as the “power houses” of eukaryotic cells. All eukaryotes were thought to have mitochondria of some form until 2016, when the first eukaryote thriving without mitochondria was discovered by our laboratory – a flagellate Monocercomonoides. Understanding cellular functions of these cells, which represent a new functional type of eukaryotes, and understanding the circumstances of the unique event of mitochondrial loss are motivations for this proposal. The first objective focuses on the cell physiology. We will perform a metabolomic study revealing major metabolic pathways and concentrate further on elucidating its unique system of iron-sulphur cluster assembly. In the second objective, we will investigate in details the unique case of mitochondrial loss. We will examine two additional potentially amitochondriate lineages by means of genomics and transcriptomics, conduct experiments simulating the moments of mitochondrial loss and try to induce the mitochondrial loss in vitro by knocking out or down genes for mitochondrial biogenesis. We have chosen Giardia intestinalis and Entamoeba histolytica as models for the latter experiments, because their mitochondria are already reduced to minimalistic “mitosomes” and because some genetic tools are already available for them. Successful mitochondrial knock-outs would enable us to study mitochondrial loss in ‘real time’ and in vivo. In the third objective, we will focus on transforming Monocercomonoides into a tractable laboratory model by developing methods of axenic cultivation and genetic manipulation. This will open new possibilities in the studies of this organism and create a cell culture representing an amitochondriate model for cell biological studies enabling the dissection of mitochondrial effects from those of other compartments. The team is composed of the laboratory of PI and eight invited experts and we hope it has the ability to address these challenging questions.
Max ERC Funding
1 935 500 €
Duration
Start date: 2018-05-01, End date: 2023-04-30
