Project acronym AppSAM
Project A Flexible Platform for the Application of SAM-dependent enzymes
Researcher (PI) Jennifer Nina ANDEXER
Host Institution (HI) ALBERT-LUDWIGS-UNIVERSITAET FREIBURG
Call Details Starting Grant (StG), LS9, ERC-2016-STG
Summary AppSAM will unlock the synthetic capability of S-adenosyl¬methionine (SAM)-dependent methyltransferases and radical SAM enzymes for application in environmentally friendly and fully sustainable reactions. The biotechnological application of these enzymes will provide access to chemo-, regio- and stereoselective methylations and alkylations, as well as to a wide range of complex rearrangement reactions that are currently not possible through traditional approaches. Methylation reactions are of particular interest due to their importance in epigenetics, cancer metabolism and the development of novel pharmaceuticals. As chemical methylation methods often involve toxic compounds and rarely exhibit the desired selectivity and specificity, there is an urgent need for new, environmentally friendly methodologies.
The proposed project will meet these demands by the provision of modular in vitro and in vivo systems that can be tailored to specific applications. In the first phase of AppSAM, efficient in vitro SAM-regeneration systems will be developed for use with methyltransferases as well as radical SAM enzymes. To achieve this aim, enzymes from different biosynthetic pathways will be combined in multi-enzyme cascades; methods from enzyme and reaction engineering will be used for optimisation. The second phase of AppSAM will address the application on a preparative scale. This will include the isolation of pure product from the in vitro systems, reactions using immobilised enzymes and extracts from in vivo productions. In addition to E. coli, the methylotrophic bacterium Methylobacter extorquens AM1 will be used as a host for the in vivo systems. M. extorquens can use C1 building blocks such as methanol as the sole carbon source, thereby initiating the biotechnological methylation process from a green source material and making the process fully sustainable, as well as being compatible with an envisaged “methanol economy”.
Summary
AppSAM will unlock the synthetic capability of S-adenosyl¬methionine (SAM)-dependent methyltransferases and radical SAM enzymes for application in environmentally friendly and fully sustainable reactions. The biotechnological application of these enzymes will provide access to chemo-, regio- and stereoselective methylations and alkylations, as well as to a wide range of complex rearrangement reactions that are currently not possible through traditional approaches. Methylation reactions are of particular interest due to their importance in epigenetics, cancer metabolism and the development of novel pharmaceuticals. As chemical methylation methods often involve toxic compounds and rarely exhibit the desired selectivity and specificity, there is an urgent need for new, environmentally friendly methodologies.
The proposed project will meet these demands by the provision of modular in vitro and in vivo systems that can be tailored to specific applications. In the first phase of AppSAM, efficient in vitro SAM-regeneration systems will be developed for use with methyltransferases as well as radical SAM enzymes. To achieve this aim, enzymes from different biosynthetic pathways will be combined in multi-enzyme cascades; methods from enzyme and reaction engineering will be used for optimisation. The second phase of AppSAM will address the application on a preparative scale. This will include the isolation of pure product from the in vitro systems, reactions using immobilised enzymes and extracts from in vivo productions. In addition to E. coli, the methylotrophic bacterium Methylobacter extorquens AM1 will be used as a host for the in vivo systems. M. extorquens can use C1 building blocks such as methanol as the sole carbon source, thereby initiating the biotechnological methylation process from a green source material and making the process fully sustainable, as well as being compatible with an envisaged “methanol economy”.
Max ERC Funding
1 499 219 €
Duration
Start date: 2017-10-01, End date: 2022-09-30
Project acronym BacBio
Project Mechanistic and functional studies of Bacillus biofilms assembly on plants, and their impact in sustainable agriculture and food safety
Researcher (PI) Diego Francisco Romero Hinojosa
Host Institution (HI) UNIVERSIDAD DE MALAGA
Call Details Starting Grant (StG), LS9, ERC-2014-STG
Summary Sustainable agriculture is an ambitious concept conceived to improve productivity but minimizing side effects. Why the efficiency of a biocontrol agent is so variable? How can different therapies be efficiently exploited in a combined way to combat microbial diseases? These are questions that need investigation to convey with criteria of sustainability. What I present is an integral proposal aim to study the microbial ecology and specifically bacterial biofilms as a central axis of two differential but likely interconnected scenarios in plant health: i) the beneficial interaction of the biocontrol agent (BCA) Bacillus subtilis, and ii) the non-conventional interaction of the food-borne pathogen Bacillus cereus.
I will start working with B. subtilis, and reasons are: 1) Different isolates are promising BCAs and are commercialized for such purpose, 2) There exist vast information of the genetics circuitries that govern important aspects of B. subtilis physiology as antibiotic production, cell differentiation, and biofilm formation. In parallel I propose to study the way B. cereus, a food-borne pathogenic bacterium interacts with vegetables. I am planning to set up a multidisciplinary approach that will combine genetics, biochemistry, proteomics, cell biology and molecular biology to visualize how these bacterial population interacts, communicates with plants and other microorganisms, or how all these factors trigger or inhibit the developmental program ending in biofilm formation. I am also interested on knowing if structural components of the bacterial extracellular matrix (exopolysaccharides or amyloid proteins) are important for bacterial fitness. If this were the case, I will also investigate which external factors affect their expression and assembly in functional biofilms. The insights get on these studies are committed to impulse our knowledge on microbial ecology and their biotechnological applicability to sustainable agriculture and food safety.
Summary
Sustainable agriculture is an ambitious concept conceived to improve productivity but minimizing side effects. Why the efficiency of a biocontrol agent is so variable? How can different therapies be efficiently exploited in a combined way to combat microbial diseases? These are questions that need investigation to convey with criteria of sustainability. What I present is an integral proposal aim to study the microbial ecology and specifically bacterial biofilms as a central axis of two differential but likely interconnected scenarios in plant health: i) the beneficial interaction of the biocontrol agent (BCA) Bacillus subtilis, and ii) the non-conventional interaction of the food-borne pathogen Bacillus cereus.
I will start working with B. subtilis, and reasons are: 1) Different isolates are promising BCAs and are commercialized for such purpose, 2) There exist vast information of the genetics circuitries that govern important aspects of B. subtilis physiology as antibiotic production, cell differentiation, and biofilm formation. In parallel I propose to study the way B. cereus, a food-borne pathogenic bacterium interacts with vegetables. I am planning to set up a multidisciplinary approach that will combine genetics, biochemistry, proteomics, cell biology and molecular biology to visualize how these bacterial population interacts, communicates with plants and other microorganisms, or how all these factors trigger or inhibit the developmental program ending in biofilm formation. I am also interested on knowing if structural components of the bacterial extracellular matrix (exopolysaccharides or amyloid proteins) are important for bacterial fitness. If this were the case, I will also investigate which external factors affect their expression and assembly in functional biofilms. The insights get on these studies are committed to impulse our knowledge on microbial ecology and their biotechnological applicability to sustainable agriculture and food safety.
Max ERC Funding
1 453 563 €
Duration
Start date: 2015-03-01, End date: 2021-02-28
Project acronym bloodANDbone
Project Blood and bone – conjoined twins in health and disease: bone marrow analogs for hematological and musculoskeletal diseases
Researcher (PI) Cornelia Lee-Thedieck
Host Institution (HI) GOTTFRIED WILHELM LEIBNIZ UNIVERSITAET HANNOVER
Call Details Starting Grant (StG), LS9, ERC-2017-STG
Summary Blood and bone are closely intertwined. Their intrinsic regenerative capacities are disturbed in many hematological and musculoskeletal diseases. Re-establishing the regenerative potential is the key to cure these diseases by regenerative medicine. Multipotent stem cells of both tissues – hematopoietic stem cells (HSCs) for blood and mesenchymal stem/stromal (MSCs) for bone – are the basis for their regenerative capacity. While it is well established that HSCs are influenced by the bone marrow in their natural environment including MSCs and their progeny, surprisingly little attention has been paid to the reciprocal relationship. The hypothesis of the current proposal is that only when taking both tissues and their mutual crosstalk into account, we will be able to understand how the regenerative potential of blood and bone is impaired in disease and how it can be re-established with novel treatment strategies. For this purpose we need to understand the early events of disease onset and progression. Due to the limitations of such studies in human beings and animals, I propose to develop human in vitro models of healthy bone marrow, which can be induced to develop hematological and musculoskeletal diseases with high incidence, namely leukemia, multiple myeloma and bone metastasis. Previously my team and I developed a simplified bone marrow analog that bases on macroporous, cell-laden biomaterials with tunable physical, biochemical and biological properties. This versatility will enable us to create biomimetic human in vitro models of the human bone marrow in health and disease, which are ground-breaking in their applicability to investigate how the regenerative balance of bone marrow is maintained in health and disturbed in the different kinds of diseases – a prerequisite to develop novel regenerative treatments – as well as their scalability and thus suitability as in vitro test systems for screening of novel drugs or treatments.
Summary
Blood and bone are closely intertwined. Their intrinsic regenerative capacities are disturbed in many hematological and musculoskeletal diseases. Re-establishing the regenerative potential is the key to cure these diseases by regenerative medicine. Multipotent stem cells of both tissues – hematopoietic stem cells (HSCs) for blood and mesenchymal stem/stromal (MSCs) for bone – are the basis for their regenerative capacity. While it is well established that HSCs are influenced by the bone marrow in their natural environment including MSCs and their progeny, surprisingly little attention has been paid to the reciprocal relationship. The hypothesis of the current proposal is that only when taking both tissues and their mutual crosstalk into account, we will be able to understand how the regenerative potential of blood and bone is impaired in disease and how it can be re-established with novel treatment strategies. For this purpose we need to understand the early events of disease onset and progression. Due to the limitations of such studies in human beings and animals, I propose to develop human in vitro models of healthy bone marrow, which can be induced to develop hematological and musculoskeletal diseases with high incidence, namely leukemia, multiple myeloma and bone metastasis. Previously my team and I developed a simplified bone marrow analog that bases on macroporous, cell-laden biomaterials with tunable physical, biochemical and biological properties. This versatility will enable us to create biomimetic human in vitro models of the human bone marrow in health and disease, which are ground-breaking in their applicability to investigate how the regenerative balance of bone marrow is maintained in health and disturbed in the different kinds of diseases – a prerequisite to develop novel regenerative treatments – as well as their scalability and thus suitability as in vitro test systems for screening of novel drugs or treatments.
Max ERC Funding
1 499 920 €
Duration
Start date: 2018-02-01, End date: 2023-01-31
Project acronym BLOODCELLSCROSSTALK
Project The Crosstalk Between Red And White Blood Cells: The Case Of Fish
Researcher (PI) Maria del Mar Ortega-Villaizan Romo
Host Institution (HI) UNIVERSIDAD MIGUEL HERNANDEZ DE ELCHE
Call Details Starting Grant (StG), LS9, ERC-2014-STG
Summary Fish are the phylogenetically oldest vertebrate group with an immune system with clear similarities to the immune system of mammals. However, it is an actual matter of fact that the current knowledge of the fish immune system seems to lack the key piece to complete the puzzle.
In 1953 Nelson described a new role of human red blood cells (RBCs) which would go beyond the simple transport of O2 to the tissues. This new role, involved in the defence against microbes, described the antibody and complement-dependent binding of microbial immune complexes to RBCs. Regardless of the importance of this finding in the field of microbial infection, this phenomenon has been poorly evaluated. Just recently, a set of biological processes relevant to immunity have been described in the RBCs of a diverse group of organisms, which include: pathogen recognition, pathogen binding and clearance and cytokines production. Furthermore, it has been demonstrated that nucleated erythrocytes from fish and avian species develop specific responses to different pathogen associated molecular patterns and produce soluble factors that modulate leukocyte activity.
In the light of these pieces of evidences, and in an attempt to improve the knowledge of the immune mechanism(s) responsible for fish protection against viral infections, we raised the question: could nucleated fish erythrocytes be the key mediators of the antiviral responses? To answer this question we decided to focus our project on the evaluation of the crosstalk between red and white blood cells in the scenario of fish viral infections and prophylaxis. For that a working model composed of the rainbow trout and the viral haemorrhagic septicaemia virus (VHSV) was chosen, being the objectives of the project to evaluate: i) the implication trout RBCs (tRBCs) in the clearance of VHSV, and ii) the involvement of tRBCs in the blood transportation of the glycoprotein G of VHSV (GVHSV), the antigen encoded by the DNA vaccine.
Summary
Fish are the phylogenetically oldest vertebrate group with an immune system with clear similarities to the immune system of mammals. However, it is an actual matter of fact that the current knowledge of the fish immune system seems to lack the key piece to complete the puzzle.
In 1953 Nelson described a new role of human red blood cells (RBCs) which would go beyond the simple transport of O2 to the tissues. This new role, involved in the defence against microbes, described the antibody and complement-dependent binding of microbial immune complexes to RBCs. Regardless of the importance of this finding in the field of microbial infection, this phenomenon has been poorly evaluated. Just recently, a set of biological processes relevant to immunity have been described in the RBCs of a diverse group of organisms, which include: pathogen recognition, pathogen binding and clearance and cytokines production. Furthermore, it has been demonstrated that nucleated erythrocytes from fish and avian species develop specific responses to different pathogen associated molecular patterns and produce soluble factors that modulate leukocyte activity.
In the light of these pieces of evidences, and in an attempt to improve the knowledge of the immune mechanism(s) responsible for fish protection against viral infections, we raised the question: could nucleated fish erythrocytes be the key mediators of the antiviral responses? To answer this question we decided to focus our project on the evaluation of the crosstalk between red and white blood cells in the scenario of fish viral infections and prophylaxis. For that a working model composed of the rainbow trout and the viral haemorrhagic septicaemia virus (VHSV) was chosen, being the objectives of the project to evaluate: i) the implication trout RBCs (tRBCs) in the clearance of VHSV, and ii) the involvement of tRBCs in the blood transportation of the glycoprotein G of VHSV (GVHSV), the antigen encoded by the DNA vaccine.
Max ERC Funding
1 823 250 €
Duration
Start date: 2015-04-01, End date: 2020-03-31
Project acronym CUSTOM-SENSE
Project Custom-made biosensors – Accelerating the transition to a bio-based economy
Researcher (PI) Jan Marienhagen
Host Institution (HI) FORSCHUNGSZENTRUM JULICH GMBH
Call Details Starting Grant (StG), LS9, ERC-2014-STG
Summary How will we meet the globally growing demand for pharmaceutically active compounds, nutrients and fine chemicals when crude oil resources are dwindling? For decades, biotechnologists have been engineering microorganisms to produce valuable compounds from sugar and biomass. However, a lack of knowledge regarding the host cell metabolism as well as long and laborious development times render this approach challenging to this day.
I want to establish a platform to engineer transcriptional biosensors for the intracellular detection of heterologous compounds in single cells. The application of these sensors in combination with flow cytometry and next-generation sequencing will enable high-throughput engineering of microorganisms at the single-cell level with unprecedented speed and simplicity.
In the field of biotechnology, this new technology will be a powerful tool for the (i) accelerated directed evolution of genes and pathways in vivo, (ii) functional integration of heterologous genes or whole synthetic pathways into the metabolism of microorganisms for the production of small valuable metabolites, (iii) genome engineering of industrially relevant microorganisms and (iv) adaptation of production strains to process conditions.
Furthermore, during CUSTOM-SENSE, biosensors will also prove to be a valuable tool to answer questions in basic science because they will help to elucidate the function of unknown genes and aid the discovery of novel and unexpected functional links in cellular metabolism.
I am in an exclusive position to pursue this goal of developing an engineering platform for custom-made biosensors due to the previous invention of biosensors at IBG-1. The starting grant would allow me to compete with Patrick D. Cirino (University of Houston, USA), who is working on a similar approach, and Christina D. Smolke (Stanford University/Caltech, USA), who is focusing on RNA devices for metabolite detection.
Summary
How will we meet the globally growing demand for pharmaceutically active compounds, nutrients and fine chemicals when crude oil resources are dwindling? For decades, biotechnologists have been engineering microorganisms to produce valuable compounds from sugar and biomass. However, a lack of knowledge regarding the host cell metabolism as well as long and laborious development times render this approach challenging to this day.
I want to establish a platform to engineer transcriptional biosensors for the intracellular detection of heterologous compounds in single cells. The application of these sensors in combination with flow cytometry and next-generation sequencing will enable high-throughput engineering of microorganisms at the single-cell level with unprecedented speed and simplicity.
In the field of biotechnology, this new technology will be a powerful tool for the (i) accelerated directed evolution of genes and pathways in vivo, (ii) functional integration of heterologous genes or whole synthetic pathways into the metabolism of microorganisms for the production of small valuable metabolites, (iii) genome engineering of industrially relevant microorganisms and (iv) adaptation of production strains to process conditions.
Furthermore, during CUSTOM-SENSE, biosensors will also prove to be a valuable tool to answer questions in basic science because they will help to elucidate the function of unknown genes and aid the discovery of novel and unexpected functional links in cellular metabolism.
I am in an exclusive position to pursue this goal of developing an engineering platform for custom-made biosensors due to the previous invention of biosensors at IBG-1. The starting grant would allow me to compete with Patrick D. Cirino (University of Houston, USA), who is working on a similar approach, and Christina D. Smolke (Stanford University/Caltech, USA), who is focusing on RNA devices for metabolite detection.
Max ERC Funding
1 482 220 €
Duration
Start date: 2015-05-01, End date: 2020-04-30
Project acronym DARKSIDE
Project Harnessing the Dark Side of Protein Folding: Manipulating Aggregation for Recombinant Protein Production
Researcher (PI) Daniel Kaganovich
Host Institution (HI) UNIVERSITAETSMEDIZIN GOETTINGEN - GEORG-AUGUST-UNIVERSITAET GOETTINGEN - STIFTUNG OEFFENTLICHEN RECHTS
Call Details Starting Grant (StG), LS9, ERC-2013-StG
Summary Nearly all desirable biological activities, whether for the purposes of nutrition, pharmacology, biofuel production, or waste disposal, can be carried out by proteins. Nature has furnished a vast array of bioactive and biocatalytic tools, and with the advent of rational protein design nearly any imaginable bioactivity is at our fingertips. There is, therefore, a pressing need for cost-effective, safe, and easily scalable strategies for generating Recombinant Proteins (rProteins). The main bottleneck for mass-producing a whole host of valuable biologically active rProteins is the difficulty of recovering functional proteins from expression hosts.
This difficulty stems largely from the lack of sufficient know-how for manipulating protein biogenesis in the cell. The key component of protein biology, whether in the context of rProtein production or cell viability, is enabling a protein to achieve its proper folding state. Most proteins do not fold on their own – they require the assistance of a vast network of folding managers, or chaperones. The cellular chaperone machinery not only assists protein folding, it also carries out quality control, ensuring that proteins that are damaged or unable to fold for other reasons are properly disposed of through degradation or protective aggregation.
The aim of this proposal is to understand the protein biosynthetic pathway in sufficient detail, so as to be able to manipulate its overall function. My eventual goal is to exert control over folding and aggregation in order to produce higher yields of functional rProteins in eukaryotes. The biotechnological strategy will consist of: 1. Manipulating aggregation to remove damaged endogenous proteins from the folding proteome, thus diverting more resources to the folding of rProteins; 2. Manipulating the allocation of cellular chaperone resources between folding, degradation, and aggregation; 3. Utilizing aggregates to produce substantially higher amounts of functional rProteins.
Summary
Nearly all desirable biological activities, whether for the purposes of nutrition, pharmacology, biofuel production, or waste disposal, can be carried out by proteins. Nature has furnished a vast array of bioactive and biocatalytic tools, and with the advent of rational protein design nearly any imaginable bioactivity is at our fingertips. There is, therefore, a pressing need for cost-effective, safe, and easily scalable strategies for generating Recombinant Proteins (rProteins). The main bottleneck for mass-producing a whole host of valuable biologically active rProteins is the difficulty of recovering functional proteins from expression hosts.
This difficulty stems largely from the lack of sufficient know-how for manipulating protein biogenesis in the cell. The key component of protein biology, whether in the context of rProtein production or cell viability, is enabling a protein to achieve its proper folding state. Most proteins do not fold on their own – they require the assistance of a vast network of folding managers, or chaperones. The cellular chaperone machinery not only assists protein folding, it also carries out quality control, ensuring that proteins that are damaged or unable to fold for other reasons are properly disposed of through degradation or protective aggregation.
The aim of this proposal is to understand the protein biosynthetic pathway in sufficient detail, so as to be able to manipulate its overall function. My eventual goal is to exert control over folding and aggregation in order to produce higher yields of functional rProteins in eukaryotes. The biotechnological strategy will consist of: 1. Manipulating aggregation to remove damaged endogenous proteins from the folding proteome, thus diverting more resources to the folding of rProteins; 2. Manipulating the allocation of cellular chaperone resources between folding, degradation, and aggregation; 3. Utilizing aggregates to produce substantially higher amounts of functional rProteins.
Max ERC Funding
1 639 400 €
Duration
Start date: 2013-11-01, End date: 2019-10-31
Project acronym E3
Project E3 - Extreme Event Ecology
Researcher (PI) Annette Menzel
Host Institution (HI) TECHNISCHE UNIVERSITAET MUENCHEN
Call Details Starting Grant (StG), LS9, ERC-2011-StG_20101109
Summary With anthropogenic warming, extreme events have already increased in magnitude and frequency and are likely to continue to do so in the near future. These extreme events play decisive roles in climate change impacts. Natural and managed systems, such as agriculture and forestry, are more strongly affected by extremes than by a change in average conditions. Classical parameters considered have included temperature, precipitation and wind speed, but here we will concentrate on multi-factorial complex situations, such as drought, and subsequent ecological events, such as pests. Novel methods from finance mathematics and statistics will be transferred for application to natural systems in order to assess risks of extremes in past, present and future conditions. Special emphasis will be given to deriving critical thresholds and prediction for when they will be crossed. Here, analyses of long-term ecoclimatological data from dendrology, phenology, seed quality, as well as both manipulated experiments and simulations are needed to provide information on the effects stemming from multiple stressors and extremes. In contrast, real data, no matter how long-term, cannot model the risk of new threatening combinations of climatological and ecological parameters. Adaptation should therefore focus not only on retrospective but also on new extremes, in other words, should look forward to the future. In particular, low probabilities and high risk scenarios have to be taken into account. Adaptation measures can range from breeding, and selection of suitable species and varieties to management options, such as sanitation and forest protection. Insurance also needs to adapt to changes in climate and ecology and accurate forecasting becomes more critical in the face of unforeseen extremes and calamities. Thus, future risk management must be based on both adaptation and insurance, with new products, such as index insurance, facilitating the handling of customer claims.
Summary
With anthropogenic warming, extreme events have already increased in magnitude and frequency and are likely to continue to do so in the near future. These extreme events play decisive roles in climate change impacts. Natural and managed systems, such as agriculture and forestry, are more strongly affected by extremes than by a change in average conditions. Classical parameters considered have included temperature, precipitation and wind speed, but here we will concentrate on multi-factorial complex situations, such as drought, and subsequent ecological events, such as pests. Novel methods from finance mathematics and statistics will be transferred for application to natural systems in order to assess risks of extremes in past, present and future conditions. Special emphasis will be given to deriving critical thresholds and prediction for when they will be crossed. Here, analyses of long-term ecoclimatological data from dendrology, phenology, seed quality, as well as both manipulated experiments and simulations are needed to provide information on the effects stemming from multiple stressors and extremes. In contrast, real data, no matter how long-term, cannot model the risk of new threatening combinations of climatological and ecological parameters. Adaptation should therefore focus not only on retrospective but also on new extremes, in other words, should look forward to the future. In particular, low probabilities and high risk scenarios have to be taken into account. Adaptation measures can range from breeding, and selection of suitable species and varieties to management options, such as sanitation and forest protection. Insurance also needs to adapt to changes in climate and ecology and accurate forecasting becomes more critical in the face of unforeseen extremes and calamities. Thus, future risk management must be based on both adaptation and insurance, with new products, such as index insurance, facilitating the handling of customer claims.
Max ERC Funding
1 487 000 €
Duration
Start date: 2012-01-01, End date: 2016-12-31
Project acronym ELONGAN
Project Gene editing and in vitro approaches to understand conceptus elongation in ungulates
Researcher (PI) Pablo BERMEJO-ÁLVAREZ
Host Institution (HI) INSTITUTO NACIONAL DE INVESTIGACION Y TECNOLOGIA AGRARIA Y ALIMENTARIA OA MP
Call Details Starting Grant (StG), LS9, ERC-2017-STG
Summary In contrast to human or rodent embryos, ungulate embryos do not implant into the uterus right after blastocyst hatching. Before implantation, the hatched ungulate blastocyst must undergo dramatic morphological changes characterized by cell differentiation, proliferation and migration processes leading to the development of extra-embryonic membranes, the appearance of a flat embryonic disc and gastrulation. This prolonged preimplantation development is termed conceptus elongation and deficiencies on this process constitute the most frequent cause of reproductive failures in ungulates, including the 4 most relevant mammalian livestock species in Europe. The purpose of this project is to elucidate the factors involved in conceptus elongation by gene editing and in vitro culture approaches. A first objective will be to identify key genes involved in differentiation processes by RNA-seq analysis of different embryo derivatives from bovine conceptuses at different developmental stages. Subsequently, the function of some of the genes identified as well as others known to play a crucial role in mouse development or putatively involved in embryo-maternal interactions will be assessed. For this aim, bovine embryos in which a candidate gene has been ablated (KO) will be generated by CRISPR and transferred to recipient females to assess in vivo the function of such particular gene on conceptus development. A second set of experiments pursue the development of an in vitro system for conceptus elongation that would bypass the requirement for in vivo experiments. For this aim we will perform metabolomics and proteomics analyses of bovine uterine fluid at different stages and will use these data to rationally develop a culture system able to sustain conceptus development. The knowledge generated by this project will serve to develop strategies to enhance farming profitability by reducing embryonic loss and to understand Developmental Biology questions unanswered by the mouse model.
Summary
In contrast to human or rodent embryos, ungulate embryos do not implant into the uterus right after blastocyst hatching. Before implantation, the hatched ungulate blastocyst must undergo dramatic morphological changes characterized by cell differentiation, proliferation and migration processes leading to the development of extra-embryonic membranes, the appearance of a flat embryonic disc and gastrulation. This prolonged preimplantation development is termed conceptus elongation and deficiencies on this process constitute the most frequent cause of reproductive failures in ungulates, including the 4 most relevant mammalian livestock species in Europe. The purpose of this project is to elucidate the factors involved in conceptus elongation by gene editing and in vitro culture approaches. A first objective will be to identify key genes involved in differentiation processes by RNA-seq analysis of different embryo derivatives from bovine conceptuses at different developmental stages. Subsequently, the function of some of the genes identified as well as others known to play a crucial role in mouse development or putatively involved in embryo-maternal interactions will be assessed. For this aim, bovine embryos in which a candidate gene has been ablated (KO) will be generated by CRISPR and transferred to recipient females to assess in vivo the function of such particular gene on conceptus development. A second set of experiments pursue the development of an in vitro system for conceptus elongation that would bypass the requirement for in vivo experiments. For this aim we will perform metabolomics and proteomics analyses of bovine uterine fluid at different stages and will use these data to rationally develop a culture system able to sustain conceptus development. The knowledge generated by this project will serve to develop strategies to enhance farming profitability by reducing embryonic loss and to understand Developmental Biology questions unanswered by the mouse model.
Max ERC Funding
1 480 880 €
Duration
Start date: 2017-10-01, End date: 2022-09-30
Project acronym EXPLOGEN
Project Exploitation of actinomycetes genomics using synthetic and system biology approaches
Researcher (PI) Andriy Luzhetskyy
Host Institution (HI) HELMHOLTZ-ZENTRUM FUR INFEKTIONSFORSCHUNG GMBH
Call Details Starting Grant (StG), LS9, ERC-2011-StG_20101109
Summary "Actinomycetes produce a wealth of important natural products, which play a pivotal role in modern drug-based therapy of various diseases. Recent whole-genome sequencing projects have revealed that the number of biosynthetic gene clusters significantly outnumbers the natural products produced by actinomycetes under laboratory conditions. Only a minority of biosynthetic gene clusters are expressed under known laboratory conditions. The major challenge in the field now is to exploit this untapped reservoir of potentially active and useful compounds. Using synthetic and system biology approaches we will access these so far unavailable natural products.
The overall strategy of the EXPLOGEN project will be implemented through the activities of four different work packages.
WP1. Synthetic biobricks. Construction of different synthetic biobricks, which can be mixed and matched to build the synthetic devices and systems in actinomycetes. Generation of streptomyces strains with minimized genomes.
WP2. Systems biology. The metabolic reconstruction of Streptomyces albus and S. lividans and its simulation in order to deliver gene targets for knockouts and overexpression experiments. Predicted mutants should accumulate main precursors of natural products, particularly polyketides.
WP3. Regulatory network identification. Using systematic in vivo transposon mutagenesis combined with GFP-based flow cytometry assay and gusA based screening, we will identify gene networks responsible for the regulation and “silencing” of natural product biosynthesis.
WP4. Metabolic engineering of the hosts. Generation of S. albus and S. lividans hosts accumulating main precursors for the heterologous production of natural products. Heterologous expression of the aranciamycin, phenalinolactone, and two ""cryptic"" biosynthetic gene clusters in the developed hosts.
The outcome of this project will be a new platform for the production of novel natural products including valuable pharmaceuticals."
Summary
"Actinomycetes produce a wealth of important natural products, which play a pivotal role in modern drug-based therapy of various diseases. Recent whole-genome sequencing projects have revealed that the number of biosynthetic gene clusters significantly outnumbers the natural products produced by actinomycetes under laboratory conditions. Only a minority of biosynthetic gene clusters are expressed under known laboratory conditions. The major challenge in the field now is to exploit this untapped reservoir of potentially active and useful compounds. Using synthetic and system biology approaches we will access these so far unavailable natural products.
The overall strategy of the EXPLOGEN project will be implemented through the activities of four different work packages.
WP1. Synthetic biobricks. Construction of different synthetic biobricks, which can be mixed and matched to build the synthetic devices and systems in actinomycetes. Generation of streptomyces strains with minimized genomes.
WP2. Systems biology. The metabolic reconstruction of Streptomyces albus and S. lividans and its simulation in order to deliver gene targets for knockouts and overexpression experiments. Predicted mutants should accumulate main precursors of natural products, particularly polyketides.
WP3. Regulatory network identification. Using systematic in vivo transposon mutagenesis combined with GFP-based flow cytometry assay and gusA based screening, we will identify gene networks responsible for the regulation and “silencing” of natural product biosynthesis.
WP4. Metabolic engineering of the hosts. Generation of S. albus and S. lividans hosts accumulating main precursors for the heterologous production of natural products. Heterologous expression of the aranciamycin, phenalinolactone, and two ""cryptic"" biosynthetic gene clusters in the developed hosts.
The outcome of this project will be a new platform for the production of novel natural products including valuable pharmaceuticals."
Max ERC Funding
1 484 016 €
Duration
Start date: 2012-01-01, End date: 2016-12-31
Project acronym FUME
Project Functional Metagenomics – Harnessing the Biotechnological Potential of Completely Novel Protein Families
Researcher (PI) Lars Ingo Ole Leichert
Host Institution (HI) RUHR-UNIVERSITAET BOCHUM
Call Details Starting Grant (StG), LS9, ERC-2011-StG_20101109
Summary Today the vast amount of newly published genomic data is generated by metagenomic projects. The annotation of these sequences of organisms, whose existence was not even known before their DNA was extracted from environmental samples, has led to the identification of thousands of new protein families with no detectable homology to any proteins in sequenced and cultivable organisms. While these protein families now make up a major part of the protein universe, we do not know anything about their functions or biocatalytic activity. In this grant application, we propose to lay the groundwork for studying the function of this vast set of proteins and exploit their biocatalytic potential. However, many of the techniques traditionally used to elucidate protein function, such as genetics and “omics”-technologies are not applicable, because the organisms in which these proteins exist are as of yet unknown. Therefore we plan to:
1. Computationally analyze the newly discovered protein families to identify one paradigmatic member of every novel family.
2. Create an expression-plasmid library of those paradigmatic representatives by de novo gene-synthesis.
3. Use this library, combined with the power of Escherichia coli genetics, for specific complementation studies and biochemical assays to assign functions to novel protein families.
This combination of synthetic biology and metagenomics will provide the starting point for a novel systematic approach to harness the biocatalytic potential and to understand the function of an unknown sector of biology that has only very recently been discovered.
Summary
Today the vast amount of newly published genomic data is generated by metagenomic projects. The annotation of these sequences of organisms, whose existence was not even known before their DNA was extracted from environmental samples, has led to the identification of thousands of new protein families with no detectable homology to any proteins in sequenced and cultivable organisms. While these protein families now make up a major part of the protein universe, we do not know anything about their functions or biocatalytic activity. In this grant application, we propose to lay the groundwork for studying the function of this vast set of proteins and exploit their biocatalytic potential. However, many of the techniques traditionally used to elucidate protein function, such as genetics and “omics”-technologies are not applicable, because the organisms in which these proteins exist are as of yet unknown. Therefore we plan to:
1. Computationally analyze the newly discovered protein families to identify one paradigmatic member of every novel family.
2. Create an expression-plasmid library of those paradigmatic representatives by de novo gene-synthesis.
3. Use this library, combined with the power of Escherichia coli genetics, for specific complementation studies and biochemical assays to assign functions to novel protein families.
This combination of synthetic biology and metagenomics will provide the starting point for a novel systematic approach to harness the biocatalytic potential and to understand the function of an unknown sector of biology that has only very recently been discovered.
Max ERC Funding
1 499 442 €
Duration
Start date: 2011-10-01, End date: 2016-11-30
