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 cis-CONTROL
Project Decoding and controlling cell-state switching: A bottom-up approach based on enhancer logic
Researcher (PI) Stein Luc AERTS
Host Institution (HI) VIB
Call Details Consolidator Grant (CoG), LS2, ERC-2016-COG
Summary Cell-state switching in cancer allows cells to transition from a proliferative to an invasive and drug-resistant phenotype. This plasticity plays an important role in cancer progression and tumour heterogeneity. We have made a striking observation that cancer cells of different origin can switch to a common survival state. During this epigenomic reprogramming, cancer cells re-activate genomic enhancers from specific regulatory programs, such as wound repair and epithelial-to-mesenchymal transition.
The goal of my project is to decipher the enhancer logic underlying this canalization effect towards a common survival state. We will then employ this new understanding of enhancer logic to engineer synthetic enhancers that are able to monitor and manipulate cell-state switching in real time. Furthermore, we will use enhancer models to identify cis-regulatory mutations that have an impact on cell-state switching and drug resistance. Such applications are currently hampered because there is a significant gap in our understanding of how enhancers work.
To tackle this problem we will use a combination of in vivo massively parallel enhancer-reporter assays, single-cell genomics on microfluidic devices, computational modelling, and synthetic enhancer design. Using these approaches we will pursue the following aims: (1) to identify functional enhancers regulating cell-state switching by performing in vivo genetic screens in mice; (2) to elucidate the dynamic trajectories whereby cells of different cancer types switch to a common survival cell-state, at single-cell resolution; (3) to create synthetic enhancer circuits that specifically kill cancer cells undergoing cell-state switching.
Our findings will have an impact on genome research, characterizing how cellular decision making is implemented by the cis-regulatory code; and on cancer research, employing enhancer logic in the context of cancer therapy.
Summary
Cell-state switching in cancer allows cells to transition from a proliferative to an invasive and drug-resistant phenotype. This plasticity plays an important role in cancer progression and tumour heterogeneity. We have made a striking observation that cancer cells of different origin can switch to a common survival state. During this epigenomic reprogramming, cancer cells re-activate genomic enhancers from specific regulatory programs, such as wound repair and epithelial-to-mesenchymal transition.
The goal of my project is to decipher the enhancer logic underlying this canalization effect towards a common survival state. We will then employ this new understanding of enhancer logic to engineer synthetic enhancers that are able to monitor and manipulate cell-state switching in real time. Furthermore, we will use enhancer models to identify cis-regulatory mutations that have an impact on cell-state switching and drug resistance. Such applications are currently hampered because there is a significant gap in our understanding of how enhancers work.
To tackle this problem we will use a combination of in vivo massively parallel enhancer-reporter assays, single-cell genomics on microfluidic devices, computational modelling, and synthetic enhancer design. Using these approaches we will pursue the following aims: (1) to identify functional enhancers regulating cell-state switching by performing in vivo genetic screens in mice; (2) to elucidate the dynamic trajectories whereby cells of different cancer types switch to a common survival cell-state, at single-cell resolution; (3) to create synthetic enhancer circuits that specifically kill cancer cells undergoing cell-state switching.
Our findings will have an impact on genome research, characterizing how cellular decision making is implemented by the cis-regulatory code; and on cancer research, employing enhancer logic in the context of cancer therapy.
Max ERC Funding
1 999 660 €
Duration
Start date: 2017-06-01, End date: 2022-05-31
Project acronym ComplexAssembly
Project The birth of protein complexes
Researcher (PI) Martin BECK
Host Institution (HI) EUROPEAN MOLECULAR BIOLOGY LABORATORY
Call Details Consolidator Grant (CoG), LS2, ERC-2016-COG
Summary Protein complexes are central to many cellular functions but our knowledge of how cells assemble protein complexes remains very sparse. Biophysical and structural data of assembly intermediates are extremely rare. Particularly in higher eukaryotes, it has become clear that complex assembly by random collision of subunits cannot cope with the spatial and temporal complexity of the intricate architecture of many cellular machines. Here I propose to combine systems biology approaches with in situ structural biology methods to visualize protein complex assembly. I want to investigate experimentally in which order the interfaces of protein complexes are formed and to which extent structures of assembly intermediates resemble those observed in fully assembled complexes. I want develop methods to systematically screen for additional factors involved in assembly pathways. I furthermore want to test the hypothesis that mechanisms must exist in eukaryotes that coordinate local mRNA translation with the ordered formation of protein complex interfaces. I believe that in order to understand assembly pathways, these processes, that so far are often studied autonomously, need to be considered jointly and in a protein complex centric manner. The research proposed here will bridge across these different scientific disciplines. In the long term, a better mechanistic understanding of protein complex assembly and the structural characterization of critical intermediates will be of high relevance for scenarios under which a cell’s protein quality control system has to cope with stress, such as aging and neurodegenerative diseases. It might also facilitate the more efficient industrial production of therapeutically relevant proteins.
Summary
Protein complexes are central to many cellular functions but our knowledge of how cells assemble protein complexes remains very sparse. Biophysical and structural data of assembly intermediates are extremely rare. Particularly in higher eukaryotes, it has become clear that complex assembly by random collision of subunits cannot cope with the spatial and temporal complexity of the intricate architecture of many cellular machines. Here I propose to combine systems biology approaches with in situ structural biology methods to visualize protein complex assembly. I want to investigate experimentally in which order the interfaces of protein complexes are formed and to which extent structures of assembly intermediates resemble those observed in fully assembled complexes. I want develop methods to systematically screen for additional factors involved in assembly pathways. I furthermore want to test the hypothesis that mechanisms must exist in eukaryotes that coordinate local mRNA translation with the ordered formation of protein complex interfaces. I believe that in order to understand assembly pathways, these processes, that so far are often studied autonomously, need to be considered jointly and in a protein complex centric manner. The research proposed here will bridge across these different scientific disciplines. In the long term, a better mechanistic understanding of protein complex assembly and the structural characterization of critical intermediates will be of high relevance for scenarios under which a cell’s protein quality control system has to cope with stress, such as aging and neurodegenerative diseases. It might also facilitate the more efficient industrial production of therapeutically relevant proteins.
Max ERC Funding
1 957 717 €
Duration
Start date: 2018-02-01, End date: 2023-01-31
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 Epiherigans
Project Writing, reading and managing stress with H3K9me
Researcher (PI) Susan GASSER
Host Institution (HI) FRIEDRICH MIESCHER INSTITUTE FOR BIOMEDICAL RESEARCH FONDATION
Call Details Advanced Grant (AdG), LS2, ERC-2016-ADG
Summary Epigenetic inheritance is the transmission of information, generally in the form of DNA methylation or post-translational modifications on histones that regulate the availability of underlying genetic information for transcription. RNA itself feeds back to contribute to histone modification. Sequence accessibility is both a matter of folding the chromatin fibre to alter access to recognition motifs, and the local concentration of factors needed for efficient transcriptional initiation, elongation, termination or mRNA stability. In heterochromatin we find a subset of regulatory factors in carefully balanced concentrations that are maintained in part by the segregation of active and inactive domains. Histone H3 K9 methylation is key to this compartmentation.
C. elegans provides an ideal system in which to study chromatin-based gene repression. We have demonstrated that histone H3 K9 methylation is the essential signal for the sequestration of heterochromatin at the nuclear envelope in C. elegans. The recognition of H3K9me1/2/3 by an inner nuclear envelope-bound chromodomain protein, CEC-4, actively sequesters heterochromatin in embryos, and contributes redundantly in adult tissues.
Epiherigans has the ambitious goal to determine definitively what targets H3K9 methylation, and identify its physiological roles. We will examine how this mark contributes to the epigenetic recognition of repeat vs non-repeat sequence, and mediates a stress-induced response to oxidative damage. We will examine the link between these and the spatial clustering of heterochromatic domains. Epiherigans will develop an integrated approach to identify in vivo the factors that distinguish repeats from non-repeats, self from non-self within genomes and will examine how H3K9me contributes to a persistent ROS or DNA damage stress response. It represents a crucial step towards understanding of how our genomes use heterochromatin to modulate, stabilize and transmit chromatin organization.
Summary
Epigenetic inheritance is the transmission of information, generally in the form of DNA methylation or post-translational modifications on histones that regulate the availability of underlying genetic information for transcription. RNA itself feeds back to contribute to histone modification. Sequence accessibility is both a matter of folding the chromatin fibre to alter access to recognition motifs, and the local concentration of factors needed for efficient transcriptional initiation, elongation, termination or mRNA stability. In heterochromatin we find a subset of regulatory factors in carefully balanced concentrations that are maintained in part by the segregation of active and inactive domains. Histone H3 K9 methylation is key to this compartmentation.
C. elegans provides an ideal system in which to study chromatin-based gene repression. We have demonstrated that histone H3 K9 methylation is the essential signal for the sequestration of heterochromatin at the nuclear envelope in C. elegans. The recognition of H3K9me1/2/3 by an inner nuclear envelope-bound chromodomain protein, CEC-4, actively sequesters heterochromatin in embryos, and contributes redundantly in adult tissues.
Epiherigans has the ambitious goal to determine definitively what targets H3K9 methylation, and identify its physiological roles. We will examine how this mark contributes to the epigenetic recognition of repeat vs non-repeat sequence, and mediates a stress-induced response to oxidative damage. We will examine the link between these and the spatial clustering of heterochromatic domains. Epiherigans will develop an integrated approach to identify in vivo the factors that distinguish repeats from non-repeats, self from non-self within genomes and will examine how H3K9me contributes to a persistent ROS or DNA damage stress response. It represents a crucial step towards understanding of how our genomes use heterochromatin to modulate, stabilize and transmit chromatin organization.
Max ERC Funding
2 500 000 €
Duration
Start date: 2017-06-01, End date: 2022-05-31
Project acronym EpiScope
Project Epigenomics and chromosome architecture one cell at a time
Researcher (PI) Marcelo NOLLMANN MARTINEZ
Host Institution (HI) CENTRE NATIONAL DE LA RECHERCHE SCIENTIFIQUE CNRS
Call Details Consolidator Grant (CoG), LS2, ERC-2016-COG
Summary In Eukaryotes, cellular identity and tissue-specific functions are linked to the epigenetic landscape and the multi-scale architecture of the genome. The packing of DNA into nucleosomes at the ~100 bp scale and the organization of whole chromosomes into functional territories within the nucleus are well documented. At an intermediate scale, chromosomes are organised in megabase to sub-megabase structures called Topologically Associating Domains (TADs). Critically, TADs are highly correlated to patterns of epigenetic marks determining the transcriptional state of the genes they encompass. Until now, the lack of efficient technologies to map chromosome architecture and epigenetic marks at the single-cell level have limited our understanding of the molecular actors and mechanisms implicated in the establishment and maintenance of the multi-scale architecture of chromosomes and epigenetic states, and the interplay between this architecture and other nuclear functions such as transcription.
The overall aim of EpiScope is to unveil the functional, multi-scale, 3D architecture of chromatin at the single-cell level while preserving cellular context, with a toolbox of groundbreaking high-performance microscopies (Hi-M). Hi-M will use unique combinations of multi-focus and single-molecule localization microscopies with novel DNA labeling methods and microfluidics. Hi-M will enable the study of structure-function relationships within TADs of different chromatin types and correlate single-cell variations in epigenomic patterns to 3D conformations with genomic specificity and at the nanoscale. Finally, Hi-M will be used to develop a novel high-throughput, high-content method to unveil the full pairwise distance distribution between thousands of genomic loci at the single cell level and at multiple length-scales. Our findings and technologies will shed new light into the mechanisms responsible for cellular memory, identity and differentiation.
Summary
In Eukaryotes, cellular identity and tissue-specific functions are linked to the epigenetic landscape and the multi-scale architecture of the genome. The packing of DNA into nucleosomes at the ~100 bp scale and the organization of whole chromosomes into functional territories within the nucleus are well documented. At an intermediate scale, chromosomes are organised in megabase to sub-megabase structures called Topologically Associating Domains (TADs). Critically, TADs are highly correlated to patterns of epigenetic marks determining the transcriptional state of the genes they encompass. Until now, the lack of efficient technologies to map chromosome architecture and epigenetic marks at the single-cell level have limited our understanding of the molecular actors and mechanisms implicated in the establishment and maintenance of the multi-scale architecture of chromosomes and epigenetic states, and the interplay between this architecture and other nuclear functions such as transcription.
The overall aim of EpiScope is to unveil the functional, multi-scale, 3D architecture of chromatin at the single-cell level while preserving cellular context, with a toolbox of groundbreaking high-performance microscopies (Hi-M). Hi-M will use unique combinations of multi-focus and single-molecule localization microscopies with novel DNA labeling methods and microfluidics. Hi-M will enable the study of structure-function relationships within TADs of different chromatin types and correlate single-cell variations in epigenomic patterns to 3D conformations with genomic specificity and at the nanoscale. Finally, Hi-M will be used to develop a novel high-throughput, high-content method to unveil the full pairwise distance distribution between thousands of genomic loci at the single cell level and at multiple length-scales. Our findings and technologies will shed new light into the mechanisms responsible for cellular memory, identity and differentiation.
Max ERC Funding
1 999 780 €
Duration
Start date: 2017-09-01, End date: 2022-08-31
Project acronym FastBio
Project A genomics and systems biology approach to explore the molecular signature and functional consequences of long-term, structured fasting in humans
Researcher (PI) Antigoni DIMA
Host Institution (HI) BIOMEDICAL SCIENCES RESEARCH CENTER ALEXANDER FLEMING
Call Details Starting Grant (StG), LS2, ERC-2016-STG
Summary Dietary intake has an enormous impact on aspects of human health, yet scientific consensus about how what we eat affects our biology remains elusive. To address the complex biological impact of diet, I propose to apply an unconventional, ‘humans-as-model-organisms’ approach to compare the molecular and functional effects of a highly structured dietary regime, specified by the Eastern Orthodox Christian Church (EOCC), to the unstructured diet followed by the general population. Individuals who follow the EOCC regime abstain from meat, dairy products and eggs for 180-200 days annually, in a temporally-structured manner initiated in childhood. I aim to explore the biological signatures of structured vs. unstructured diet by addressing three objectives. First I will investigate the effects of the two regimes, and of genetic variation, on higher-level phenotypes including anthropometric, physiological and biomarker traits. Second, I will carry out a comprehensive set of omics assays (metabolomics, transcriptomics, epigenomics and investigation of the gut microbiome), will associate omics phenotypes with genetic variation, and will integrate data across biological levels to uncover complex molecular signatures. Third, I will interrogate the functional consequences of dietary regimes at the cellular level through primary cell culture. Acute and long-term effects of dietary intake will be explored for all objectives through a two timepoint sampling strategy. This proposal therefore comprises a unique opportunity to study a specific perturbation (EOCC structured diet) introduced to a steady-state system (unstructured diet followed by the general population) in a ground-breaking human systems biology type of study. This approach brings together expertise from genomics, computational biology, statistics, medicine and epidemiology. It will lead to novel insights regarding the potent signalling nature of nutrients and is likely to yield results of high translational value.
Summary
Dietary intake has an enormous impact on aspects of human health, yet scientific consensus about how what we eat affects our biology remains elusive. To address the complex biological impact of diet, I propose to apply an unconventional, ‘humans-as-model-organisms’ approach to compare the molecular and functional effects of a highly structured dietary regime, specified by the Eastern Orthodox Christian Church (EOCC), to the unstructured diet followed by the general population. Individuals who follow the EOCC regime abstain from meat, dairy products and eggs for 180-200 days annually, in a temporally-structured manner initiated in childhood. I aim to explore the biological signatures of structured vs. unstructured diet by addressing three objectives. First I will investigate the effects of the two regimes, and of genetic variation, on higher-level phenotypes including anthropometric, physiological and biomarker traits. Second, I will carry out a comprehensive set of omics assays (metabolomics, transcriptomics, epigenomics and investigation of the gut microbiome), will associate omics phenotypes with genetic variation, and will integrate data across biological levels to uncover complex molecular signatures. Third, I will interrogate the functional consequences of dietary regimes at the cellular level through primary cell culture. Acute and long-term effects of dietary intake will be explored for all objectives through a two timepoint sampling strategy. This proposal therefore comprises a unique opportunity to study a specific perturbation (EOCC structured diet) introduced to a steady-state system (unstructured diet followed by the general population) in a ground-breaking human systems biology type of study. This approach brings together expertise from genomics, computational biology, statistics, medicine and epidemiology. It will lead to novel insights regarding the potent signalling nature of nutrients and is likely to yield results of high translational value.
Max ERC Funding
1 500 000 €
Duration
Start date: 2017-06-01, End date: 2023-03-31
Project acronym GENOMIS
Project Illuminating GENome Organization through integrated MIcroscopy and Sequencing
Researcher (PI) Marzena Magda BIENKO
Host Institution (HI) KAROLINSKA INSTITUTET
Call Details Starting Grant (StG), LS2, ERC-2016-STG
Summary In human cells, two meters of DNA sequence are compressed into a nucleus whose linear size is five orders of magnitude smaller. Deciphering how this amazing structural organization is achieved and how DNA functions can ensue in the environment of a cell’s nucleus represent central questions for contemporary biology.
Here, I embrace this challenge by establishing a comprehensive framework of microscopy and sequencing technologies coupled with advanced analytical approaches, aimed at addressing three fundamental highly-interconnected questions: 1) What are the design principles that govern DNA compaction? 2) How does genome structure vary between different cell types as well as among cells of the same type? 3) What is the link between genome structure and function? In preliminary experiments, we have devised a powerful method for Genomic loci Positioning by Sequencing (GPSeq) in fixed cells with optimally preserved nuclear morphology. In parallel, we are developing high-end microscopy tools for simultaneous localization of dozens of genomic locations at high resolution in thousands of single cells.
We will obtain first-ever genome-wide maps of radial positioning of DNA loci in the nucleus, and combine them with available DNA contact probability maps in order to build 3D models of the human genome structure in different cell types. Using microscopy, we will visualize chromosomal shapes at unprecedented resolution, and use these rich datasets to discover general DNA folding principles. Finally, by combining high-resolution chromosome visualization with gene expression profiling in single cells, we will explore the link between DNA structure and function. Our study shall illuminate the design principles that dictate how genetic information is packed and read in the human nucleus, while providing a comprehensive repertoire of tools for studying genome organization.
Summary
In human cells, two meters of DNA sequence are compressed into a nucleus whose linear size is five orders of magnitude smaller. Deciphering how this amazing structural organization is achieved and how DNA functions can ensue in the environment of a cell’s nucleus represent central questions for contemporary biology.
Here, I embrace this challenge by establishing a comprehensive framework of microscopy and sequencing technologies coupled with advanced analytical approaches, aimed at addressing three fundamental highly-interconnected questions: 1) What are the design principles that govern DNA compaction? 2) How does genome structure vary between different cell types as well as among cells of the same type? 3) What is the link between genome structure and function? In preliminary experiments, we have devised a powerful method for Genomic loci Positioning by Sequencing (GPSeq) in fixed cells with optimally preserved nuclear morphology. In parallel, we are developing high-end microscopy tools for simultaneous localization of dozens of genomic locations at high resolution in thousands of single cells.
We will obtain first-ever genome-wide maps of radial positioning of DNA loci in the nucleus, and combine them with available DNA contact probability maps in order to build 3D models of the human genome structure in different cell types. Using microscopy, we will visualize chromosomal shapes at unprecedented resolution, and use these rich datasets to discover general DNA folding principles. Finally, by combining high-resolution chromosome visualization with gene expression profiling in single cells, we will explore the link between DNA structure and function. Our study shall illuminate the design principles that dictate how genetic information is packed and read in the human nucleus, while providing a comprehensive repertoire of tools for studying genome organization.
Max ERC Funding
1 499 808 €
Duration
Start date: 2018-01-01, End date: 2022-12-31
Project acronym GENSURGE
Project Designer recombinases for efficient and safe genome surgery
Researcher (PI) Frank Buchholz
Host Institution (HI) TECHNISCHE UNIVERSITAET DRESDEN
Call Details Advanced Grant (AdG), LS2, ERC-2016-ADG
Summary Recent breakthroughs in the field of genome editing provide a genuine opportunity to establish innovative
approaches to repair DNA mutations to replace, engineer or regenerate malfunctioning cells in vitro or in
vivo. However, most of the recently developed technologies introduce double-strand DNA breaks at a target
locus as the first step to gene correction. These breaks are subsequently repaired by one of the cell intrinsic
DNA repair pathways, typically inducing an abundance of insertions and deletions (indels). Ideally, for many
applications genome editing should, however, be efficient and specific, without the introduction of indels.
Site-specific recombinases (SSRs) allow precise genome editing without triggering endogenous DNA repair
pathways and possess the unique ability to fulfill both cleavage and immediate resealing of the processed
DNA in vivo. However, customizing the DNA binding specificity of SSRs is not straightforward. With this
project, we propose to solve this shortcoming. We have already demonstrated that by applying substrate-linked
directed evolution, SSRs can be generated that specifically recognize therapeutic targets. The
objective of this project is the development of a universal genome editing platform that allows flexible,
efficient and safe gene corrections in cells of any origin without triggering cell intrinsic DNA repair.
GenSurge aims to: i) sequence an unprecedented, comprehensive compendium of evolved SSRs to
understand the directed molecular evolution process at nucleotide resolution; ii) integrate the knowledge
obtained in i) to develop a unique SSR-based approach to correct genomic inversions; iii) develop a
universal SSR-based strategy that allows flawless, precise and safe genome editing to correct any gene defect
in human, animal or plant cells. The successful implementation of this project will deliver a comprehensive,
safe and efficient platform from which genome surgery-based cure strategies can be initiated.
Summary
Recent breakthroughs in the field of genome editing provide a genuine opportunity to establish innovative
approaches to repair DNA mutations to replace, engineer or regenerate malfunctioning cells in vitro or in
vivo. However, most of the recently developed technologies introduce double-strand DNA breaks at a target
locus as the first step to gene correction. These breaks are subsequently repaired by one of the cell intrinsic
DNA repair pathways, typically inducing an abundance of insertions and deletions (indels). Ideally, for many
applications genome editing should, however, be efficient and specific, without the introduction of indels.
Site-specific recombinases (SSRs) allow precise genome editing without triggering endogenous DNA repair
pathways and possess the unique ability to fulfill both cleavage and immediate resealing of the processed
DNA in vivo. However, customizing the DNA binding specificity of SSRs is not straightforward. With this
project, we propose to solve this shortcoming. We have already demonstrated that by applying substrate-linked
directed evolution, SSRs can be generated that specifically recognize therapeutic targets. The
objective of this project is the development of a universal genome editing platform that allows flexible,
efficient and safe gene corrections in cells of any origin without triggering cell intrinsic DNA repair.
GenSurge aims to: i) sequence an unprecedented, comprehensive compendium of evolved SSRs to
understand the directed molecular evolution process at nucleotide resolution; ii) integrate the knowledge
obtained in i) to develop a unique SSR-based approach to correct genomic inversions; iii) develop a
universal SSR-based strategy that allows flawless, precise and safe genome editing to correct any gene defect
in human, animal or plant cells. The successful implementation of this project will deliver a comprehensive,
safe and efficient platform from which genome surgery-based cure strategies can be initiated.
Max ERC Funding
2 380 425 €
Duration
Start date: 2018-01-01, End date: 2022-12-31
Project acronym GeroProtect
Project Developing Geroprotectors to Prevent Polymorbidity
Researcher (PI) Linda PARTRIDGE
Host Institution (HI) MAX-PLANCK-GESELLSCHAFT ZUR FORDERUNG DER WISSENSCHAFTEN EV
Call Details Advanced Grant (AdG), LS2, ERC-2016-ADG
Summary Advancing age is the major risk factor for disability and illness, including cardiovascular, metabolic and neu-rodegenerative disease and cancer. The increasing incidence of older people in European countries is posing major medical, social and economic challenges, and there is an urgent need to find ways of compressing late-life morbidity. Ageing has proved malleable to genetic and pharmacological interventions in laboratory animals, and at least some of the mechanisms are conserved over large evolutionary distances. Reduced activity of the nutrient-sensing insulin/insulin-like growth factor/TOR signalling network can increase health and combat ageing-related disease in laboratory animals, with increasing evidence of its importance in human ageing. There is thus a prospect for pharmacological intervention to prevent more than one ageing-related condition, rather than tackling diseases one by one and as they arise. The aim of this research programme is to evaluate the potential for pharmacological prevention of ageing-related decline in humans with a polypill targeting the nutrient-sensing network. We find that three licensed drugs, lithium, rapamycin and trametinib, act independently, at different nodes in the network, to increase lifespan in the fruitfly Drosophila, implying that the network controls more than one underlying mechanism of ageing, and that a polypill of these drugs could be particularly effective. We shall test this idea in mice, and assess the underlying mechanisms in Drosophila and mice. We have found that suppression of the Ras signalling branch of the network, which has a well known role in human cancer, can extend lifespan in both the fruitfly Drosophila and mice, and we shall assess its role in humans. Interventions that ameliorate ageing often have sex-specific effects, and we shall investigate the mechanisms leading to these for the nutrient-sensing network. The outputs of the work will inform future clinical trails in humans.
Summary
Advancing age is the major risk factor for disability and illness, including cardiovascular, metabolic and neu-rodegenerative disease and cancer. The increasing incidence of older people in European countries is posing major medical, social and economic challenges, and there is an urgent need to find ways of compressing late-life morbidity. Ageing has proved malleable to genetic and pharmacological interventions in laboratory animals, and at least some of the mechanisms are conserved over large evolutionary distances. Reduced activity of the nutrient-sensing insulin/insulin-like growth factor/TOR signalling network can increase health and combat ageing-related disease in laboratory animals, with increasing evidence of its importance in human ageing. There is thus a prospect for pharmacological intervention to prevent more than one ageing-related condition, rather than tackling diseases one by one and as they arise. The aim of this research programme is to evaluate the potential for pharmacological prevention of ageing-related decline in humans with a polypill targeting the nutrient-sensing network. We find that three licensed drugs, lithium, rapamycin and trametinib, act independently, at different nodes in the network, to increase lifespan in the fruitfly Drosophila, implying that the network controls more than one underlying mechanism of ageing, and that a polypill of these drugs could be particularly effective. We shall test this idea in mice, and assess the underlying mechanisms in Drosophila and mice. We have found that suppression of the Ras signalling branch of the network, which has a well known role in human cancer, can extend lifespan in both the fruitfly Drosophila and mice, and we shall assess its role in humans. Interventions that ameliorate ageing often have sex-specific effects, and we shall investigate the mechanisms leading to these for the nutrient-sensing network. The outputs of the work will inform future clinical trails in humans.
Max ERC Funding
2 500 000 €
Duration
Start date: 2017-12-01, End date: 2022-11-30
