Project acronym Agglomerates
Project Infinite Protein Self-Assembly in Health and Disease
Researcher (PI) Emmanuel Doram LEVY
Host Institution (HI) WEIZMANN INSTITUTE OF SCIENCE
Call Details Consolidator Grant (CoG), LS2, ERC-2018-COG
Summary Understanding how proteins respond to mutations is of paramount importance to biology and disease. While protein stability and misfolding have been instrumental in rationalizing the impact of mutations, we recently discovered that an alternative route is also frequent, where mutations at the surface of symmetric proteins trigger novel self-interactions that lead to infinite self-assembly. This mechanism can be involved in disease, as in sickle-cell anemia, but may also serve in adaptation. Importantly, it differs fundamentally from aggregation, because misfolding does not drive it. Thus, we term it “agglomeration”. The ease with which agglomeration can occur, even by single point mutations, shifts the paradigm of how quickly new protein assemblies can emerge, both in health and disease. This prompts us to determine the basic principles of protein agglomeration and explore its implications in cell physiology and human disease.
We propose an interdisciplinary research program bridging atomic and cellular scales to explore agglomeration in three aims: (i) Map the landscape of protein agglomeration in response to mutation in endogenous yeast proteins; (ii) Characterize how yeast physiology impacts agglomeration by changes in gene expression or cell state, and, conversely, how protein agglomerates impact yeast fitness. (iii) Analyze agglomeration in relation to human disease via two approaches. First, by predicting single nucleotide polymorphisms that trigger agglomeration, prioritizing them using knowledge from Aims 1 & 2, and characterizing them experimentally. Second, by providing a proof-of-concept that agglomeration can be exploited in drug design, whereby drugs induce its formation, like mutations can do.
Overall, through this research, we aim to establish agglomeration as a paradigm for protein assembly, with implications for our understanding of evolution, physiology, and disease.
Summary
Understanding how proteins respond to mutations is of paramount importance to biology and disease. While protein stability and misfolding have been instrumental in rationalizing the impact of mutations, we recently discovered that an alternative route is also frequent, where mutations at the surface of symmetric proteins trigger novel self-interactions that lead to infinite self-assembly. This mechanism can be involved in disease, as in sickle-cell anemia, but may also serve in adaptation. Importantly, it differs fundamentally from aggregation, because misfolding does not drive it. Thus, we term it “agglomeration”. The ease with which agglomeration can occur, even by single point mutations, shifts the paradigm of how quickly new protein assemblies can emerge, both in health and disease. This prompts us to determine the basic principles of protein agglomeration and explore its implications in cell physiology and human disease.
We propose an interdisciplinary research program bridging atomic and cellular scales to explore agglomeration in three aims: (i) Map the landscape of protein agglomeration in response to mutation in endogenous yeast proteins; (ii) Characterize how yeast physiology impacts agglomeration by changes in gene expression or cell state, and, conversely, how protein agglomerates impact yeast fitness. (iii) Analyze agglomeration in relation to human disease via two approaches. First, by predicting single nucleotide polymorphisms that trigger agglomeration, prioritizing them using knowledge from Aims 1 & 2, and characterizing them experimentally. Second, by providing a proof-of-concept that agglomeration can be exploited in drug design, whereby drugs induce its formation, like mutations can do.
Overall, through this research, we aim to establish agglomeration as a paradigm for protein assembly, with implications for our understanding of evolution, physiology, and disease.
Max ERC Funding
2 574 819 €
Duration
Start date: 2019-04-01, End date: 2024-03-31
Project acronym BiomeRiskFactors
Project Discovering microbiome-based disease risk factors
Researcher (PI) Eran Segal
Host Institution (HI) WEIZMANN INSTITUTE OF SCIENCE
Call Details Advanced Grant (AdG), LS2, ERC-2017-ADG
Summary Identifying risk factors for diseases that can be prevented or delayed by early intervention is of major importance, and numerous genetic, lifestyle, anthropometric and clinical risk factors were found for many different diseases. Another source of potentially pertinent disease risk factors is the human microbiome - the collective genome of trillions of bacteria, viruses, fungi, and parasites that reside in the human gut. However, very few microbiome disease markers were found to date.
Here, we aim to develop risk prediction tools based on the human microbiome that predict the likelihood of an individual to develop a particular condition or disease within 5-10 years. We will use a cohort of >2200 individuals that my group previously assembled, for whom we have clinical profiles, gut microbiome data, and banked blood and stool samples. We will invite people 5-10 years after their initial recruitment time, profile disease status and blood markers, and develop algorithms for predicting 5-10 year onset of Type 2 diabetes, cardiovascular disease, and obesity, using microbiome data from recruitment time.
To increase the likelihood of finding microbiome markers predictive of disease onset, we will develop novel experimental and computational methods for in-depth characterization of microbial gene function, the metabolites produced by the microbiome, the underexplored fungal microbiome members, and the interactions between the gut microbiota and the host adaptive immune system. We will then apply these methods to >2200 banked samples from cohort recruitment time and use the resulting data in devising our microbiome-based risk prediction tools. In themselves, these novel assays and their application to >2200 samples should greatly advance the microbiome field.
If successful, our proposal will identify new disease risk factors and risk prediction tools based on the microbiome, paving the way towards using the microbiome in early disease detection and prevention.
Summary
Identifying risk factors for diseases that can be prevented or delayed by early intervention is of major importance, and numerous genetic, lifestyle, anthropometric and clinical risk factors were found for many different diseases. Another source of potentially pertinent disease risk factors is the human microbiome - the collective genome of trillions of bacteria, viruses, fungi, and parasites that reside in the human gut. However, very few microbiome disease markers were found to date.
Here, we aim to develop risk prediction tools based on the human microbiome that predict the likelihood of an individual to develop a particular condition or disease within 5-10 years. We will use a cohort of >2200 individuals that my group previously assembled, for whom we have clinical profiles, gut microbiome data, and banked blood and stool samples. We will invite people 5-10 years after their initial recruitment time, profile disease status and blood markers, and develop algorithms for predicting 5-10 year onset of Type 2 diabetes, cardiovascular disease, and obesity, using microbiome data from recruitment time.
To increase the likelihood of finding microbiome markers predictive of disease onset, we will develop novel experimental and computational methods for in-depth characterization of microbial gene function, the metabolites produced by the microbiome, the underexplored fungal microbiome members, and the interactions between the gut microbiota and the host adaptive immune system. We will then apply these methods to >2200 banked samples from cohort recruitment time and use the resulting data in devising our microbiome-based risk prediction tools. In themselves, these novel assays and their application to >2200 samples should greatly advance the microbiome field.
If successful, our proposal will identify new disease risk factors and risk prediction tools based on the microbiome, paving the way towards using the microbiome in early disease detection and prevention.
Max ERC Funding
2 500 000 €
Duration
Start date: 2019-03-01, End date: 2024-02-29
Project acronym CancerFluxome
Project Cancer Cellular Metabolism across Space and Time
Researcher (PI) Tomer Shlomi
Host Institution (HI) TECHNION - ISRAEL INSTITUTE OF TECHNOLOGY
Call Details Starting Grant (StG), LS2, ERC-2016-STG
Summary The metabolism of cancer cells is altered to meet cellular requirements for growth, providing novel means to selectively target tumorigenesis. While extensively studied, our current view of cancer cellular metabolism is fundamentally limited by lack of information on variability in metabolic activity between distinct subcellular compartments and cells.
We propose to develop a spatio-temporal fluxomics approach for quantifying metabolic fluxes in the cytoplasm vs. mitochondria as well as their cell-cycle dynamics, combining mass-spectrometry based isotope tracing with cell synchronization, rapid cellular fractionation, and computational metabolic network modelling.
Spatio-temporal fluxomics will be used to revisit and challenge our current understanding of central metabolism and its induced adaptation to oncogenic events – an important endeavour considering that mitochondrial bioenergetics and biosynthesis are required for tumorigenesis and accumulating evidences for metabolic alterations throughout the cell-cycle.
Our preliminary results show intriguing oscillations between oxidative and reductive TCA cycle flux throughout the cell-cycle. We will explore the extent to which cells adapt their metabolism to fulfil the changing energetic and anabolic demands throughout the cell-cycle, how metabolic oscillations are regulated, and their benefit to cells in terms of thermodynamic efficiency. Spatial flux analysis will be instrumental for investigating glutaminolysis - a ‘hallmark’ metabolic adaptation in cancer involving shuttling of metabolic intermediates and cofactors between mitochondria and cytoplasm.
On a clinical front, our spatio-temporal fluxomics analysis will enable to disentangle oncogene-induced flux alterations, having an important tumorigenic role, from artefacts originating from population averaging. A comprehensive view of how cells adapt their metabolism due to oncogenic mutations will reveal novel targets for anti-cancer drugs.
Summary
The metabolism of cancer cells is altered to meet cellular requirements for growth, providing novel means to selectively target tumorigenesis. While extensively studied, our current view of cancer cellular metabolism is fundamentally limited by lack of information on variability in metabolic activity between distinct subcellular compartments and cells.
We propose to develop a spatio-temporal fluxomics approach for quantifying metabolic fluxes in the cytoplasm vs. mitochondria as well as their cell-cycle dynamics, combining mass-spectrometry based isotope tracing with cell synchronization, rapid cellular fractionation, and computational metabolic network modelling.
Spatio-temporal fluxomics will be used to revisit and challenge our current understanding of central metabolism and its induced adaptation to oncogenic events – an important endeavour considering that mitochondrial bioenergetics and biosynthesis are required for tumorigenesis and accumulating evidences for metabolic alterations throughout the cell-cycle.
Our preliminary results show intriguing oscillations between oxidative and reductive TCA cycle flux throughout the cell-cycle. We will explore the extent to which cells adapt their metabolism to fulfil the changing energetic and anabolic demands throughout the cell-cycle, how metabolic oscillations are regulated, and their benefit to cells in terms of thermodynamic efficiency. Spatial flux analysis will be instrumental for investigating glutaminolysis - a ‘hallmark’ metabolic adaptation in cancer involving shuttling of metabolic intermediates and cofactors between mitochondria and cytoplasm.
On a clinical front, our spatio-temporal fluxomics analysis will enable to disentangle oncogene-induced flux alterations, having an important tumorigenic role, from artefacts originating from population averaging. A comprehensive view of how cells adapt their metabolism due to oncogenic mutations will reveal novel targets for anti-cancer drugs.
Max ERC Funding
1 481 250 €
Duration
Start date: 2017-02-01, End date: 2022-01-31
Project acronym CHROMATINSYS
Project Systematic Approach to Dissect the Interplay between Chromatin and Transcription
Researcher (PI) Nir Friedman
Host Institution (HI) THE HEBREW UNIVERSITY OF JERUSALEM
Call Details Advanced Grant (AdG), LS2, ERC-2013-ADG
Summary Epigenetic mechanisms play an important role in regulating and maintaining the functionality of cells and have been implicated in a wide range of human diseases. Histone proteins that form the protein core of nucleosomes are subject to a bewildering array of covalent and structural modifications, which can repress, permit, or promote transcription. These modifications can be added and removed by specialized complexes that are recruited by other covalent modifications, by transcription factors, or by the transcriptional machinery. Advances in genomics led to comprehensive mapping of the ``epigenome'' in a range of tissues and organisms. These maps established the tight connection between histone modifications and transcription programs. These static charts, however, are less successful at uncovering the underlying mechanisms, logic, and function of histone modifications in establishing and maintaining transcriptional programs. Our premise is that we can answer these basic questions by observing the effect of genetic perturbations on the dynamics of both chromatin state and transcriptional activity. We aim to dissect the chromatin-transcription system in a systematic manner by building on our extensive experience in modeling and analysis, and a unique high-throughput experimental system we established in my lab.
We plan to use the budding yeast model organism, which allows for
efficient genetic and experimental manipulations. We will combine two technologies: (1) high-throughput measurements of single-cell
transcriptional output using fluorescence reporters; and (2) high-throughput immunoprecipitation sequencing assays to map chromatin state. Measuring with these the dynamics of response to stimuli under different genetic backgrounds and using advanced stochastic network models, we will chart detailed mechanisms that are opaque to current approaches and elucidate the general principles that govern the interplay between chromatin and transcription.
Summary
Epigenetic mechanisms play an important role in regulating and maintaining the functionality of cells and have been implicated in a wide range of human diseases. Histone proteins that form the protein core of nucleosomes are subject to a bewildering array of covalent and structural modifications, which can repress, permit, or promote transcription. These modifications can be added and removed by specialized complexes that are recruited by other covalent modifications, by transcription factors, or by the transcriptional machinery. Advances in genomics led to comprehensive mapping of the ``epigenome'' in a range of tissues and organisms. These maps established the tight connection between histone modifications and transcription programs. These static charts, however, are less successful at uncovering the underlying mechanisms, logic, and function of histone modifications in establishing and maintaining transcriptional programs. Our premise is that we can answer these basic questions by observing the effect of genetic perturbations on the dynamics of both chromatin state and transcriptional activity. We aim to dissect the chromatin-transcription system in a systematic manner by building on our extensive experience in modeling and analysis, and a unique high-throughput experimental system we established in my lab.
We plan to use the budding yeast model organism, which allows for
efficient genetic and experimental manipulations. We will combine two technologies: (1) high-throughput measurements of single-cell
transcriptional output using fluorescence reporters; and (2) high-throughput immunoprecipitation sequencing assays to map chromatin state. Measuring with these the dynamics of response to stimuli under different genetic backgrounds and using advanced stochastic network models, we will chart detailed mechanisms that are opaque to current approaches and elucidate the general principles that govern the interplay between chromatin and transcription.
Max ERC Funding
2 396 450 €
Duration
Start date: 2014-01-01, End date: 2018-12-31
Project acronym CNIDARIAMICRORNA
Project Elucidation of the evolution of post-transcriptional regulation by characterizing the cnidarian microRNA pathway
Researcher (PI) Yehu Moran
Host Institution (HI) THE HEBREW UNIVERSITY OF JERUSALEM
Call Details Starting Grant (StG), LS2, ERC-2014-STG
Summary Over the past decade small RNAs such as microRNAs (miRNAs) and small interfering RNAs (siRNAs) have been shown to carry pivotal roles in post-transcriptional regulation and genome protection and to play an important part in various physiological processes in animals. miRNAs can be found in a very wide range of animals yet their functions were studied almost exclusively in members of the Bilateria such as insects, nematodes and vertebrates. Hence studying their function in representatives of non-bilaterian phyla such as Cnidaria (sea anemones, corals, hydras and jellyfish) is crucial for understanding the evolution of miRNAs in animals and can provide important insights into their roles in the ancient ancestor of Cnidaria and Bilateria. The sea anemone Nematostella vectensis is an excellent model for such a study since it can be grown in large numbers throughout its life cycle in the lab and because well-established genetic manipulation techniques are available for this species. Our preliminary results indicate that miRNAs in Nematostella frequently have a nearly perfect match to their messenger RNA (mRNA) targets, resulting in cleavage of the target. This mode of action is common for plant miRNAs, but is very rare in Bilateria. This finding together with my recent discovery of a Nematostella homolog of HYL1, a protein involved in miRNA biogenesis in plants, raises the exciting possibility that the miRNA pathway existed in the common ancestor of plants and animals. Here I suggest to bring together an array of advanced biochemical and genetic methods such as gene knockdown, transgenesis, high throughput sequencing and immunoprecipitation in order to obtain - for the first time - a deep understanding of the biogenesis and mechanism of action of small RNAs in Cnidaria. This will provide a novel way to understand the evolution of this important molecular pathway and to evaluate its age and ancestral form.
Summary
Over the past decade small RNAs such as microRNAs (miRNAs) and small interfering RNAs (siRNAs) have been shown to carry pivotal roles in post-transcriptional regulation and genome protection and to play an important part in various physiological processes in animals. miRNAs can be found in a very wide range of animals yet their functions were studied almost exclusively in members of the Bilateria such as insects, nematodes and vertebrates. Hence studying their function in representatives of non-bilaterian phyla such as Cnidaria (sea anemones, corals, hydras and jellyfish) is crucial for understanding the evolution of miRNAs in animals and can provide important insights into their roles in the ancient ancestor of Cnidaria and Bilateria. The sea anemone Nematostella vectensis is an excellent model for such a study since it can be grown in large numbers throughout its life cycle in the lab and because well-established genetic manipulation techniques are available for this species. Our preliminary results indicate that miRNAs in Nematostella frequently have a nearly perfect match to their messenger RNA (mRNA) targets, resulting in cleavage of the target. This mode of action is common for plant miRNAs, but is very rare in Bilateria. This finding together with my recent discovery of a Nematostella homolog of HYL1, a protein involved in miRNA biogenesis in plants, raises the exciting possibility that the miRNA pathway existed in the common ancestor of plants and animals. Here I suggest to bring together an array of advanced biochemical and genetic methods such as gene knockdown, transgenesis, high throughput sequencing and immunoprecipitation in order to obtain - for the first time - a deep understanding of the biogenesis and mechanism of action of small RNAs in Cnidaria. This will provide a novel way to understand the evolution of this important molecular pathway and to evaluate its age and ancestral form.
Max ERC Funding
1 499 587 €
Duration
Start date: 2015-05-01, End date: 2020-04-30
Project acronym CrackEpitranscriptom
Project Cracking the epitranscriptome
Researcher (PI) Schraga SCHWARTZ
Host Institution (HI) WEIZMANN INSTITUTE OF SCIENCE
Call Details Starting Grant (StG), LS2, ERC-2016-STG
Summary Over 100 types of distinct modifications are catalyzed on RNA molecules post-transcriptionally. In an analogous manner to well-studied chemical modifications on proteins or DNA, modifications on RNA - and particularly on mRNA - harbor the exciting potential of regulating the complex and interlinked life cycle of these molecules. The most abundant modification in mammalian and yeast mRNA is N6-methyladenosine (m6A). We have pioneered approaches for mapping m6A in a transcriptome wide manner, and we and others have identified factors involved in encoding and decoding m6A. While experimental disruption of these factors is associated with severe phenotypes, the role of m6A remains enigmatic. No single methylated site has been shown to causally underlie any physiological or molecular function. This proposal aims to establish a framework for systematically deciphering the molecular function of a modification and its underlying mechanisms and to uncover the physiological role of the modification in regulation of a cellular response. We will apply this framework to m6A in the context of meiosis in budding yeast, as m6A dynamically accumulates on meiotic mRNAs and as the methyltransferase catalyzing m6A is essential for meiosis. We will (1) aim to elucidate the physiological targets of methylation governing entry into meiosis (2) seek to elucidate the function of m6A at the molecular level, and understand its impact on the various steps of the mRNA life cycle, (3) seek to understand the mechanisms underlying its effects. These aims will provide a comprehensive framework for understanding how the epitranscriptome, an emerging post-transcriptional layer of regulation, fine-tunes gene regulation and impacts cellular decision making in a dynamic response, and will set the stage towards dissecting the roles of m6A and of an expanding set of mRNA modifications in more complex and disease related systems.
Summary
Over 100 types of distinct modifications are catalyzed on RNA molecules post-transcriptionally. In an analogous manner to well-studied chemical modifications on proteins or DNA, modifications on RNA - and particularly on mRNA - harbor the exciting potential of regulating the complex and interlinked life cycle of these molecules. The most abundant modification in mammalian and yeast mRNA is N6-methyladenosine (m6A). We have pioneered approaches for mapping m6A in a transcriptome wide manner, and we and others have identified factors involved in encoding and decoding m6A. While experimental disruption of these factors is associated with severe phenotypes, the role of m6A remains enigmatic. No single methylated site has been shown to causally underlie any physiological or molecular function. This proposal aims to establish a framework for systematically deciphering the molecular function of a modification and its underlying mechanisms and to uncover the physiological role of the modification in regulation of a cellular response. We will apply this framework to m6A in the context of meiosis in budding yeast, as m6A dynamically accumulates on meiotic mRNAs and as the methyltransferase catalyzing m6A is essential for meiosis. We will (1) aim to elucidate the physiological targets of methylation governing entry into meiosis (2) seek to elucidate the function of m6A at the molecular level, and understand its impact on the various steps of the mRNA life cycle, (3) seek to understand the mechanisms underlying its effects. These aims will provide a comprehensive framework for understanding how the epitranscriptome, an emerging post-transcriptional layer of regulation, fine-tunes gene regulation and impacts cellular decision making in a dynamic response, and will set the stage towards dissecting the roles of m6A and of an expanding set of mRNA modifications in more complex and disease related systems.
Max ERC Funding
1 402 666 €
Duration
Start date: 2016-11-01, End date: 2021-10-31
Project acronym CRISPRsition
Project Developing CRISPR adaptation platforms for basic and applied research
Researcher (PI) Ehud Itzhak Qimron
Host Institution (HI) TEL AVIV UNIVERSITY
Call Details Consolidator Grant (CoG), LS2, ERC-2018-COG
Summary The CRISPR-Cas system has been extensively studied for its ability to cleave DNA. In contrast, studies of the ability of the system to acquire and integrate new DNA from invaders as a form of prokaryotic adaptive immunity, have lagged behind. This delay reflects the extreme enthusiasm surrounding the potential of using the system’s cleavage capabilities as a genome editing tool. However, the enormous potential of the adaptation process can and should arouse a similar degree of enthusiasm. My lab has pioneered studies on the CRISPR adaptation process by establishing new methodologies, and applying them to demonstrate the essential role of the proteins and DNA elements, as well as the molecular mechanisms, operating in this process. In this project, I will establish novel platforms for studying adaptation and develop them into biotechnological applications and research tools. These tools will allow me to identify the first natural and synthetic inhibitors of the adaptation process. This, in turn, will provide genetic tools to control adaptation, as well as advance the understanding of the arms race between bacteria and their invaders. I will also harness the adaptation process as a platform for diversifying genetic elements for phage display, and for extending phage recognition of a wide range of hosts. Lastly, I will provide the first evidence for an association between the CRISPR adaptation system and gene repression. This linkage will form the basis of a molecular scanner and recorder platform that I will develop and that can be used to identify crucial genetic elements in phage genomes as well as novel regulatory circuits in the bacterial genome. Together, my findings will represent a considerable leap in the understanding of CRISPR adaptation with respect to the process, potential applications, and the intriguing evolutionary significance.
Summary
The CRISPR-Cas system has been extensively studied for its ability to cleave DNA. In contrast, studies of the ability of the system to acquire and integrate new DNA from invaders as a form of prokaryotic adaptive immunity, have lagged behind. This delay reflects the extreme enthusiasm surrounding the potential of using the system’s cleavage capabilities as a genome editing tool. However, the enormous potential of the adaptation process can and should arouse a similar degree of enthusiasm. My lab has pioneered studies on the CRISPR adaptation process by establishing new methodologies, and applying them to demonstrate the essential role of the proteins and DNA elements, as well as the molecular mechanisms, operating in this process. In this project, I will establish novel platforms for studying adaptation and develop them into biotechnological applications and research tools. These tools will allow me to identify the first natural and synthetic inhibitors of the adaptation process. This, in turn, will provide genetic tools to control adaptation, as well as advance the understanding of the arms race between bacteria and their invaders. I will also harness the adaptation process as a platform for diversifying genetic elements for phage display, and for extending phage recognition of a wide range of hosts. Lastly, I will provide the first evidence for an association between the CRISPR adaptation system and gene repression. This linkage will form the basis of a molecular scanner and recorder platform that I will develop and that can be used to identify crucial genetic elements in phage genomes as well as novel regulatory circuits in the bacterial genome. Together, my findings will represent a considerable leap in the understanding of CRISPR adaptation with respect to the process, potential applications, and the intriguing evolutionary significance.
Max ERC Funding
2 000 000 €
Duration
Start date: 2019-12-01, End date: 2024-11-30
Project acronym DecodeDegRNA
Project Post-transcriptional regulation of RNA degradation in early zebrafish development
Researcher (PI) Michal Rabani
Host Institution (HI) THE HEBREW UNIVERSITY OF JERUSALEM
Call Details Starting Grant (StG), LS2, ERC-2019-STG
Summary Regulation of gene expression lies at the heart of fundamental biological processes, such as the formation of different cell types inside an embryo or responses to environmental stimuli. Living cells ensure that the right genes are expressed at the right time and place by carefully controlling every RNA molecule inside a cell from its ‘birth’ by transcription to its final ‘death’ by degradation. While vast efforts strive to understand the first part of this process – transcription, studies of RNA degradation have been more limited. Current knowledge largely relies on small-scale investigation of key – but anecdotal – cases, while technical and experimental difficulties limit its large-scale analysis. Therefore, we still lack a systematic and predictive understanding of RNA degradation: technologies to globally measure it, the molecular mechanisms involved, its functional and physiological implications and models to decode and predict it. Transcriptional silencing makes early embryos an ideal system to study RNA degradation and uncover its basic concepts, as I propose here. Aim 1 will decipher how genomic information within native RNA sequences determines their degradation in embryos. Aim 2 will develop the technology to investigate RNA degradation at single-cell resolution, and uncover its regulation within arising embryonic cell populations. Aim 3 will reveal the molecular implementation of the regulatory code of RNA degradation and determine its physiological roles that underlie the massive degradation of maternal mRNAs – a key regulatory event and a main developmental transition in early embryos of all animals. This work will uncover new principles of RNA degradation in early development and elicit its mechanisms and functions using the zebrafish as an in vivo model system. The assays and models to be developed will be broadly applicable to study RNA degradation in diverse contexts, ranging from disease mechanisms to engineering of RNA- protein interactions.
Summary
Regulation of gene expression lies at the heart of fundamental biological processes, such as the formation of different cell types inside an embryo or responses to environmental stimuli. Living cells ensure that the right genes are expressed at the right time and place by carefully controlling every RNA molecule inside a cell from its ‘birth’ by transcription to its final ‘death’ by degradation. While vast efforts strive to understand the first part of this process – transcription, studies of RNA degradation have been more limited. Current knowledge largely relies on small-scale investigation of key – but anecdotal – cases, while technical and experimental difficulties limit its large-scale analysis. Therefore, we still lack a systematic and predictive understanding of RNA degradation: technologies to globally measure it, the molecular mechanisms involved, its functional and physiological implications and models to decode and predict it. Transcriptional silencing makes early embryos an ideal system to study RNA degradation and uncover its basic concepts, as I propose here. Aim 1 will decipher how genomic information within native RNA sequences determines their degradation in embryos. Aim 2 will develop the technology to investigate RNA degradation at single-cell resolution, and uncover its regulation within arising embryonic cell populations. Aim 3 will reveal the molecular implementation of the regulatory code of RNA degradation and determine its physiological roles that underlie the massive degradation of maternal mRNAs – a key regulatory event and a main developmental transition in early embryos of all animals. This work will uncover new principles of RNA degradation in early development and elicit its mechanisms and functions using the zebrafish as an in vivo model system. The assays and models to be developed will be broadly applicable to study RNA degradation in diverse contexts, ranging from disease mechanisms to engineering of RNA- protein interactions.
Max ERC Funding
1 500 000 €
Duration
Start date: 2020-03-01, End date: 2025-02-28
Project acronym DEPICT
Project Design principles and controllability of protein circuits
Researcher (PI) Uri Alon
Host Institution (HI) WEIZMANN INSTITUTE OF SCIENCE
Call Details Advanced Grant (AdG), LS2, ERC-2009-AdG
Summary Cells use circuits of interacting proteins to respond to their environment. In the past decades, molecular biology has provided detailed knowledge on the proteins in these circuits and their interactions. To fully understand circuit function requires, in addition to molecular knowledge, new concepts that explain how multiple components work together to perform systems level functions. Our lab has been a leader in defining such concepts, based on combined experimental and theoretical study of well characterized circuits in bacteria and human cells. In this proposal we aim to find novel principles on how circuits resist fluctuations and errors, and how they can be controlled by drugs: (1) Why do key regulatory systems use bifunctional enzymes that catalyze antagonistic reactions (e.g. both kinase and phosphatase)? We will test the role of bifunctional enzymes in making circuits robust to variations in protein levels. (2) Why are some genes regulated by a repressor and others by an activator? We will test this in the context of reduction of errors in transcription control. (3) Are there principles that describe how drugs combine to affect protein dynamics in human cells? We will use a novel dynamic proteomics approach developed in our lab to explore how protein dynamics can be controlled by drug combinations. This research will define principles that unite our understanding of seemingly distinct biological systems, and explain their particular design in terms of systems-level functions. This understanding will help form the basis for a future medicine that rationally controls the state of the cell based on a detailed blueprint of their circuit design, and quantitative principles for the effects of drugs on this circuitry.
Summary
Cells use circuits of interacting proteins to respond to their environment. In the past decades, molecular biology has provided detailed knowledge on the proteins in these circuits and their interactions. To fully understand circuit function requires, in addition to molecular knowledge, new concepts that explain how multiple components work together to perform systems level functions. Our lab has been a leader in defining such concepts, based on combined experimental and theoretical study of well characterized circuits in bacteria and human cells. In this proposal we aim to find novel principles on how circuits resist fluctuations and errors, and how they can be controlled by drugs: (1) Why do key regulatory systems use bifunctional enzymes that catalyze antagonistic reactions (e.g. both kinase and phosphatase)? We will test the role of bifunctional enzymes in making circuits robust to variations in protein levels. (2) Why are some genes regulated by a repressor and others by an activator? We will test this in the context of reduction of errors in transcription control. (3) Are there principles that describe how drugs combine to affect protein dynamics in human cells? We will use a novel dynamic proteomics approach developed in our lab to explore how protein dynamics can be controlled by drug combinations. This research will define principles that unite our understanding of seemingly distinct biological systems, and explain their particular design in terms of systems-level functions. This understanding will help form the basis for a future medicine that rationally controls the state of the cell based on a detailed blueprint of their circuit design, and quantitative principles for the effects of drugs on this circuitry.
Max ERC Funding
2 261 440 €
Duration
Start date: 2010-03-01, End date: 2015-02-28
Project acronym DMR-CODE
Project Decoding the Mammalian transcriptional Regulatory code in development and stimulatory responses
Researcher (PI) Ido Amit
Host Institution (HI) WEIZMANN INSTITUTE OF SCIENCE
Call Details Starting Grant (StG), LS2, ERC-2012-StG_20111109
Summary Transcription factors (TF) regulate genome function by controlling gene expression. Comprehensive characterization of the in vivo binding of TF to the DNA in relevant primary models is a critical step towards a global understanding of the human genome. Recent advances in high-throughput genomic technologies provide an extraordinary opportunity to develop and apply systematic approaches to learn the underline principles and mechanisms of mammalian transcriptional networks. The premise of this proposal is that a tractable set of rules govern how cells commit to a specific cell type or respond to the environment, and that these rules are coded in regulatory elements in the genome. Currently our understanding of the mammalian regulatory code is hampered by the difficulty of directly measuring in vivo binding of large numbers of TFs to DNA across multiple primary cell types and their natural response to physiological stimuli.
Here, we overcome this bottleneck by systematically exploring the genomic binding network of 1. All relevant TFs of key hematopoietic cells in both steady state and under relevant stimuli. 2. Follow the changes in TF networks as cells differentiate 3. Use these models to engineer cell states and responses. To achieve these goals, we developed a new method for automated high throughput ChIP coupled to sequencing (HT-ChIP-Seq). We used this method to measure binding of 40 TFs in 4 time points following stimulation of dendritic cells with pathogen components. We find that TFs vary substantially in their binding dynamics, genomic localization, number of binding events, and degree of interaction with other TFs. The analysis of this data suggests that the TF network is hierarchically organized, and composed of different types of TFs, cell differentiation factors, factors that prime for gene induction, and factors that bind more specifically and dynamically. This proposal revisits and challenges the current understanding of the mammalian regulatory code.
Summary
Transcription factors (TF) regulate genome function by controlling gene expression. Comprehensive characterization of the in vivo binding of TF to the DNA in relevant primary models is a critical step towards a global understanding of the human genome. Recent advances in high-throughput genomic technologies provide an extraordinary opportunity to develop and apply systematic approaches to learn the underline principles and mechanisms of mammalian transcriptional networks. The premise of this proposal is that a tractable set of rules govern how cells commit to a specific cell type or respond to the environment, and that these rules are coded in regulatory elements in the genome. Currently our understanding of the mammalian regulatory code is hampered by the difficulty of directly measuring in vivo binding of large numbers of TFs to DNA across multiple primary cell types and their natural response to physiological stimuli.
Here, we overcome this bottleneck by systematically exploring the genomic binding network of 1. All relevant TFs of key hematopoietic cells in both steady state and under relevant stimuli. 2. Follow the changes in TF networks as cells differentiate 3. Use these models to engineer cell states and responses. To achieve these goals, we developed a new method for automated high throughput ChIP coupled to sequencing (HT-ChIP-Seq). We used this method to measure binding of 40 TFs in 4 time points following stimulation of dendritic cells with pathogen components. We find that TFs vary substantially in their binding dynamics, genomic localization, number of binding events, and degree of interaction with other TFs. The analysis of this data suggests that the TF network is hierarchically organized, and composed of different types of TFs, cell differentiation factors, factors that prime for gene induction, and factors that bind more specifically and dynamically. This proposal revisits and challenges the current understanding of the mammalian regulatory code.
Max ERC Funding
1 500 000 €
Duration
Start date: 2012-10-01, End date: 2017-09-30
