Project acronym VitASTEM
Project Regulation of Single Hematopoietic Stem Cells by Intake of Vitamin A
Researcher (PI) Nina CABEZAS WALLSCHEID
Host Institution (HI) MAX-PLANCK-GESELLSCHAFT ZUR FORDERUNG DER WISSENSCHAFTEN EV
Call Details Starting Grant (StG), LS3, ERC-2017-STG
Summary Quiescence preserves the self-renewal capacity and the long-term function of hematopoietic stem cells (HSCs). The regulators of this dormant state include intrinsic pathways and soluble components in the bone marrow niche. Dysregulation of this process is poorly defined and might cause aberrant hematopoiesis. In my previous work, we defined the molecular landscape of HSCs by applying state of the art DNA-methylome, RNA-seq and proteome analyses, and found vitamin A/retinoic acid (RA)-induced signaling predominantly enriched in HSCs (Cabezas-Wallscheid et al., Cell Stem Cell 2014). Intriguingly, we observed that mice fed with a vitamin A-free diet exhibited a robust loss of HSCs (Cabezas-Wallscheid et al., Cell 2017). Treatment of mice with a RA agonist preserved HSC quiescence in stress-activated conditions, indicating that the balance between HSC maintenance and differentiation is tightly regulated by vitamin A signaling.
However, we are only beginning to understand the mechanisms how vitamin A regulates HSC fate. Since treatment of vitamin A deficiency currently shows extremely low therapeutic success, novel insights into the role of HSCs in the development of the disease will be of enormous therapeutic value.
Summary
Quiescence preserves the self-renewal capacity and the long-term function of hematopoietic stem cells (HSCs). The regulators of this dormant state include intrinsic pathways and soluble components in the bone marrow niche. Dysregulation of this process is poorly defined and might cause aberrant hematopoiesis. In my previous work, we defined the molecular landscape of HSCs by applying state of the art DNA-methylome, RNA-seq and proteome analyses, and found vitamin A/retinoic acid (RA)-induced signaling predominantly enriched in HSCs (Cabezas-Wallscheid et al., Cell Stem Cell 2014). Intriguingly, we observed that mice fed with a vitamin A-free diet exhibited a robust loss of HSCs (Cabezas-Wallscheid et al., Cell 2017). Treatment of mice with a RA agonist preserved HSC quiescence in stress-activated conditions, indicating that the balance between HSC maintenance and differentiation is tightly regulated by vitamin A signaling.
However, we are only beginning to understand the mechanisms how vitamin A regulates HSC fate. Since treatment of vitamin A deficiency currently shows extremely low therapeutic success, novel insights into the role of HSCs in the development of the disease will be of enormous therapeutic value.
Max ERC Funding
1 500 000 €
Duration
Start date: 2018-05-01, End date: 2023-04-30
Project acronym VitruVius
Project The Genetics and Physiology of Growth and Size Determination
Researcher (PI) Pierre LEOPOLD
Host Institution (HI) INSTITUT NATIONAL DE LA SANTE ET DE LA RECHERCHE MEDICALE
Call Details Advanced Grant (AdG), LS3, ERC-2015-AdG
Summary Body size is an intrinsic property of living organisms that is intimately linked to the developmental program to produce fit individuals with proper proportions. Final size is the result of both genetic determinants and sophisticated mechanisms adapting size to available resources. Classical regeneration and transplantation experiments have established that different body parts grow according to autonomous programs, challenging the concept of systemic and harmonious growth. Therefore, coordination mechanisms must ensure that all parts have reached appropriate final size before animals stop growing. The main objective of this program is to tackle the mechanisms ensuring growth coordination and size determination. To tackle these questions, we propose to bring together physiology, genetics, advanced imaging and modelling approaches in an unprecedented integrated project. We will develop these approaches on the Drosophila model where we gained expertise, and will develop the vertebrate model Ambystoma mexicanum (axolotl) well adapted for limb growth and regeneration studies. Three major aims will be addressed:
1- We will explore the mechanisms coupling the developmental clock with organ growth.
2- We will dissect the mechanisms allowing inter-organ growth coordination both in response to abnormal growth (injury/neoplasm) and in the course of normal development (control of developmental instability).
3- We will use our knowledge acquired on insect models to develop Ambystoma mexicanum (axolotl) as a vertebrate model for organ growth coordination.
We believe this project brings together novel and complementary expertise that will contribute to broader understanding of the fascinating question of body growth and final size assessment.
Summary
Body size is an intrinsic property of living organisms that is intimately linked to the developmental program to produce fit individuals with proper proportions. Final size is the result of both genetic determinants and sophisticated mechanisms adapting size to available resources. Classical regeneration and transplantation experiments have established that different body parts grow according to autonomous programs, challenging the concept of systemic and harmonious growth. Therefore, coordination mechanisms must ensure that all parts have reached appropriate final size before animals stop growing. The main objective of this program is to tackle the mechanisms ensuring growth coordination and size determination. To tackle these questions, we propose to bring together physiology, genetics, advanced imaging and modelling approaches in an unprecedented integrated project. We will develop these approaches on the Drosophila model where we gained expertise, and will develop the vertebrate model Ambystoma mexicanum (axolotl) well adapted for limb growth and regeneration studies. Three major aims will be addressed:
1- We will explore the mechanisms coupling the developmental clock with organ growth.
2- We will dissect the mechanisms allowing inter-organ growth coordination both in response to abnormal growth (injury/neoplasm) and in the course of normal development (control of developmental instability).
3- We will use our knowledge acquired on insect models to develop Ambystoma mexicanum (axolotl) as a vertebrate model for organ growth coordination.
We believe this project brings together novel and complementary expertise that will contribute to broader understanding of the fascinating question of body growth and final size assessment.
Max ERC Funding
2 499 144 €
Duration
Start date: 2017-01-01, End date: 2021-12-31
Project acronym WING MORPHOGENESIS
Project A physical basis for wing morphogenesis and planar cell polarity
Researcher (PI) Suzanne Eaton
Host Institution (HI) MAX-PLANCK-GESELLSCHAFT ZUR FORDERUNG DER WISSENSCHAFTEN EV
Call Details Advanced Grant (AdG), LS3, ERC-2009-AdG
Summary During development, physical forces are generated in precise patterns and produce elegant choreography of cell movements that determine tissue shape. The function of many tissues depends not only on their shape, but on the correct alignment of planar cell polarity within the tissue. Remarkably, recent evidence from my lab has suggested that physical forces not only shape the wing, but also align the planar polarity of its constituent cells with the proximal distal wing axis. The wing blade is remodeled at pupal stages by proximal-distal stretching caused by contraction of the wing hinge. Hinge contraction produces precise patterns of oriented cell rearrangements and cell divisions in the wing blade that lengthen it proximo-distally and refine its shape. The polarity of cell rearrangements also re-orients intracellularly polarized complexes of Planar Cell Polarity (PCP) proteins to face the distal side of the wing. This occurs because these complexes turn over very slowly, compared with the rate of cell rearrangement. We will investigate three problems defined by this work. First, how does polarized cell stretching cause epithelial remodeling? The pupal wing is the first in vivo example of this process in a genetically and physically accessible model. Second, what are the genetic, cellular, and physical mechanisms that specify the pattern of cellular flow occuring in the wing blade? Third, what signals orient PCP during early wing development? This previously undescribed early polarity is oriented roughly perpendicular to the final direction, is a critical starting point for the later development of proximal-distal polarity. This work will provide important insight into genetic, cellular and physical mechansisms that shape and polarize tissues.
Summary
During development, physical forces are generated in precise patterns and produce elegant choreography of cell movements that determine tissue shape. The function of many tissues depends not only on their shape, but on the correct alignment of planar cell polarity within the tissue. Remarkably, recent evidence from my lab has suggested that physical forces not only shape the wing, but also align the planar polarity of its constituent cells with the proximal distal wing axis. The wing blade is remodeled at pupal stages by proximal-distal stretching caused by contraction of the wing hinge. Hinge contraction produces precise patterns of oriented cell rearrangements and cell divisions in the wing blade that lengthen it proximo-distally and refine its shape. The polarity of cell rearrangements also re-orients intracellularly polarized complexes of Planar Cell Polarity (PCP) proteins to face the distal side of the wing. This occurs because these complexes turn over very slowly, compared with the rate of cell rearrangement. We will investigate three problems defined by this work. First, how does polarized cell stretching cause epithelial remodeling? The pupal wing is the first in vivo example of this process in a genetically and physically accessible model. Second, what are the genetic, cellular, and physical mechanisms that specify the pattern of cellular flow occuring in the wing blade? Third, what signals orient PCP during early wing development? This previously undescribed early polarity is oriented roughly perpendicular to the final direction, is a critical starting point for the later development of proximal-distal polarity. This work will provide important insight into genetic, cellular and physical mechansisms that shape and polarize tissues.
Max ERC Funding
1 531 200 €
Duration
Start date: 2010-03-01, End date: 2015-05-31
Project acronym WNTEXPORT
Project Sorting processes that ensure short and long-range action of Wnts in developing epithelia
Researcher (PI) Jean-Paul B.B. Vincent
Host Institution (HI) THE FRANCIS CRICK INSTITUTE LIMITED
Call Details Advanced Grant (AdG), LS3, ERC-2011-ADG_20110310
Summary Wnts are signaling proteins that act both at short and long range in developing tissues. Several proteins, such as Wntless, are specifically devoted to Wnt secretion, indicating that Wnts may follow a distinct secretory route. Moreover, Wnts carry two lipid modifications, which are likely to interfere with diffusion in the extracellular space. Much of our work will focus on the trafficking of Wingless (the main Drosophila Wnt), which forms a concentration gradient in wing imaginal discs. To chart the route taken by Wingless from the ER to responding cells, we will devise techniques (e.g. BirA-dependent in vivo biotinylation) to pulse label endogenously expressed Wingless in the secretory pathway and at the cell surface. Wingless routing will also be investigated in conditions that alter Evi/Wntless trafficking. We will capitalize on our observation that Wingless and Wntless are present on exosomes in conditioned medium. These exosomes will be purified and characterized by mass spectrometry and the resulting information will be used to devise rigorous functional assays. Similar approaches will be used to identify and characterize proteins that associate with soluble Wingless, which is also present in conditioned medium. Our proposed approaches will also enable us to assess, for the first time, the function of exosomes in an intact animal. Once secreted, Wingless and associated proteins spread in the extracellular space while remaining associated with the epithelial surface. We will use single molecule imaging in a reconstituted system along with mathematical modeling to test the hypothesis that the glypican-Wnt interaction is sufficiently strong to ensure surface retention while allowing diffusion in two dimensions. Finally we will use biochemical approaches and molecular genetics in Drosophila and mice to investigate the mode of action of Notum, a glypican-modifying enzyme that could be relevant to the progression of Wnt signaling dependent cancers.
Summary
Wnts are signaling proteins that act both at short and long range in developing tissues. Several proteins, such as Wntless, are specifically devoted to Wnt secretion, indicating that Wnts may follow a distinct secretory route. Moreover, Wnts carry two lipid modifications, which are likely to interfere with diffusion in the extracellular space. Much of our work will focus on the trafficking of Wingless (the main Drosophila Wnt), which forms a concentration gradient in wing imaginal discs. To chart the route taken by Wingless from the ER to responding cells, we will devise techniques (e.g. BirA-dependent in vivo biotinylation) to pulse label endogenously expressed Wingless in the secretory pathway and at the cell surface. Wingless routing will also be investigated in conditions that alter Evi/Wntless trafficking. We will capitalize on our observation that Wingless and Wntless are present on exosomes in conditioned medium. These exosomes will be purified and characterized by mass spectrometry and the resulting information will be used to devise rigorous functional assays. Similar approaches will be used to identify and characterize proteins that associate with soluble Wingless, which is also present in conditioned medium. Our proposed approaches will also enable us to assess, for the first time, the function of exosomes in an intact animal. Once secreted, Wingless and associated proteins spread in the extracellular space while remaining associated with the epithelial surface. We will use single molecule imaging in a reconstituted system along with mathematical modeling to test the hypothesis that the glypican-Wnt interaction is sufficiently strong to ensure surface retention while allowing diffusion in two dimensions. Finally we will use biochemical approaches and molecular genetics in Drosophila and mice to investigate the mode of action of Notum, a glypican-modifying enzyme that could be relevant to the progression of Wnt signaling dependent cancers.
Max ERC Funding
1 938 846 €
Duration
Start date: 2012-07-01, End date: 2017-06-30
Project acronym XABA
Project Xenophagy and bacterial avoidance
Researcher (PI) Hans-Christian Friedrich-Wilhelm Behrends
Host Institution (HI) JOHANN WOLFGANG GOETHE-UNIVERSITATFRANKFURT AM MAIN
Call Details Starting Grant (StG), LS3, ERC-2011-StG_20101109
Summary "Autophagy is an essential process that enables cells to engulf and digest portions of their cytoplasm, thereby accomplishing quality and quantity control of organelles, proteins and pathogens. These homeostatic and adaptive function intricately link autophagy to diverse health and disease states including innate immunity. Microbial pathogens that successfully parasitize eukaryotic cells have evolved to evade autophagic microbial defenses (xenophagy) and subvert the host autophagic responses for their own survival and/or growth. Central to xenophagy is cargo recognition and dynamic rearrangements of membrane-bound compartments to sequester and deliver pathogen load for lysosomal degradation. Microbial adaptation strategies identified to date have targeted both of these crucial and intertwining functions. However, the precise molecular mechanisms underpinning pathogen avoidance of host-cell autophagy and immune responses are only poorly understood. This raised specific questions about host-pathogen interactions: (1) What is the dynamic response of the core autophagy system to bacterial pathogens? (2) How does the autophagy pathway impinge on the endocytic system and how do pathogens subvert intracellular vesicle trafficking systems? (3) What is the molecular machinery involved in detecting cargo and how do pathogens counter this response? This proposal seeks to elucidate pathways and functions involved in xenophagy and to identify mechanisms by which pathogens usurpate the anti-bacterial cell-autonomous defence system. To address these questions, we will employ a highly complementary multidisciplinary platform, which combines biochemical, cell and infection biological approaches with emerging proteomics and high-content imaging techniques. Together, this proposal has the potential to uncover novel molecular mechanisms that define the signaling machinery driving xenophagy of pathogens, thereby contributing to advancements in the fields of cell and infection biology."
Summary
"Autophagy is an essential process that enables cells to engulf and digest portions of their cytoplasm, thereby accomplishing quality and quantity control of organelles, proteins and pathogens. These homeostatic and adaptive function intricately link autophagy to diverse health and disease states including innate immunity. Microbial pathogens that successfully parasitize eukaryotic cells have evolved to evade autophagic microbial defenses (xenophagy) and subvert the host autophagic responses for their own survival and/or growth. Central to xenophagy is cargo recognition and dynamic rearrangements of membrane-bound compartments to sequester and deliver pathogen load for lysosomal degradation. Microbial adaptation strategies identified to date have targeted both of these crucial and intertwining functions. However, the precise molecular mechanisms underpinning pathogen avoidance of host-cell autophagy and immune responses are only poorly understood. This raised specific questions about host-pathogen interactions: (1) What is the dynamic response of the core autophagy system to bacterial pathogens? (2) How does the autophagy pathway impinge on the endocytic system and how do pathogens subvert intracellular vesicle trafficking systems? (3) What is the molecular machinery involved in detecting cargo and how do pathogens counter this response? This proposal seeks to elucidate pathways and functions involved in xenophagy and to identify mechanisms by which pathogens usurpate the anti-bacterial cell-autonomous defence system. To address these questions, we will employ a highly complementary multidisciplinary platform, which combines biochemical, cell and infection biological approaches with emerging proteomics and high-content imaging techniques. Together, this proposal has the potential to uncover novel molecular mechanisms that define the signaling machinery driving xenophagy of pathogens, thereby contributing to advancements in the fields of cell and infection biology."
Max ERC Funding
1 600 000 €
Duration
Start date: 2012-01-01, End date: 2016-12-31
Project acronym XChromosome
Project Functions of the X chromosome in the mammalian germ line
Researcher (PI) James Turner
Host Institution (HI) THE FRANCIS CRICK INSTITUTE LIMITED
Call Details Consolidator Grant (CoG), LS3, ERC-2014-CoG
Summary Male and female mammals differ in their sex chromosome complement: males have one X chromosome and one Y chromosome (XY), while females have two X chromosomes (XX). Decades of research have demonstrated that the Y chromosome is crucial for male fertility, regulating both sex differentiation and spermatogenesis. However, the role of the X chromosome in these critical stages of germ line development remains poorly understood.
In this proposal, we will explore the functions of the X chromosome in the mammalian germ line. In eutherian mammals, sex determination is dependent on the Y-gene Sry, but in the other major mammalian class, the metatherians, it is controlled not only by a Y-locus, but also by the complement of X chromosomes. In Aim 1 we will apply CRISPR-mediated genome editing to metatherian mammals in order to identify these critical X and Y sex-determining genes. Our findings will provide insight into mammalian sex chromosome evolution and the aetiology of human sex differentiation disorders.
In addition to its role in sex determination, the X chromosome is also predicted to become evolutionarily specialised for spermatogenesis. Consistent with this theory, we discovered that the mouse X chromosome is enriched in genes expressed exclusively in developing sperm. These genes are present in multiple copies, and together comprise an unprecedented 18% of the mouse X chromosome gene content. Many could be important candidates for unexplained male infertility. However, the functions of these genes have not been determined, because they cannot be manipulated using conventional gene targeting strategies. In Aim 2, we will use CRISPR to establish the roles of X-multicopy genes in male reproduction.
Summary
Male and female mammals differ in their sex chromosome complement: males have one X chromosome and one Y chromosome (XY), while females have two X chromosomes (XX). Decades of research have demonstrated that the Y chromosome is crucial for male fertility, regulating both sex differentiation and spermatogenesis. However, the role of the X chromosome in these critical stages of germ line development remains poorly understood.
In this proposal, we will explore the functions of the X chromosome in the mammalian germ line. In eutherian mammals, sex determination is dependent on the Y-gene Sry, but in the other major mammalian class, the metatherians, it is controlled not only by a Y-locus, but also by the complement of X chromosomes. In Aim 1 we will apply CRISPR-mediated genome editing to metatherian mammals in order to identify these critical X and Y sex-determining genes. Our findings will provide insight into mammalian sex chromosome evolution and the aetiology of human sex differentiation disorders.
In addition to its role in sex determination, the X chromosome is also predicted to become evolutionarily specialised for spermatogenesis. Consistent with this theory, we discovered that the mouse X chromosome is enriched in genes expressed exclusively in developing sperm. These genes are present in multiple copies, and together comprise an unprecedented 18% of the mouse X chromosome gene content. Many could be important candidates for unexplained male infertility. However, the functions of these genes have not been determined, because they cannot be manipulated using conventional gene targeting strategies. In Aim 2, we will use CRISPR to establish the roles of X-multicopy genes in male reproduction.
Max ERC Funding
1 912 369 €
Duration
Start date: 2015-11-01, End date: 2021-10-31
Project acronym ZEBRAFISHANGIO
Project Analysis of flow regulated angiogenesis during zebrafish development
Researcher (PI) Arndt Friedrich Siekmann
Host Institution (HI) MAX-PLANCK-GESELLSCHAFT ZUR FORDERUNG DER WISSENSCHAFTEN EV
Call Details Starting Grant (StG), LS3, ERC-2010-StG_20091118
Summary Angiogenesis, or the sprouting of blood vessels from pre-existing ones is a key process in shaping the vasculature. So far, angiogenesis research has mainly focused on identifying genetic players. My proposed research takes a completely novel approach towards understanding angiogenesis by addressing the question how physiological factors, such as changes in blood flow dynamics, influence endothelial biology. I will furthermore address the question if there is an interaction between genetic and physiological influences during angiogenesis. My proposed research will tackle these outstanding questions in endothelial biology using several cutting edge technologies. First, I will perform long term time lapse imaging of the growing vasculature using 2-Photon microscopy. Second, I will use an innovative methodology employing zinc finger nuclease mediated gene targeting to rapidly generate mutant zebrafish in genes implicated in mediating the effects of hemodynamics on angiogenesis. Third, I will laser ablate individual blood vessels in living zebrafish embryos to alter blood flow patterns and study the influence of those changes on angiogenesis. These challenging technological advancements will make it possible for the first time to understand how physiological factors aid in shaping the forming vascular system. These findings will have broad applicability to angiogenic processes in many different vascular beds and developmental time points. Importantly, the mechanisms at play during vascular development in the embryo are reactivated in the adult in situations of pathological angiogenesis, such as tumor progression or ischemia. Therefore, applying this knowledge to angiogenesis occuring in pathological settings will provide us with new avenues for improved disease treatments.
Summary
Angiogenesis, or the sprouting of blood vessels from pre-existing ones is a key process in shaping the vasculature. So far, angiogenesis research has mainly focused on identifying genetic players. My proposed research takes a completely novel approach towards understanding angiogenesis by addressing the question how physiological factors, such as changes in blood flow dynamics, influence endothelial biology. I will furthermore address the question if there is an interaction between genetic and physiological influences during angiogenesis. My proposed research will tackle these outstanding questions in endothelial biology using several cutting edge technologies. First, I will perform long term time lapse imaging of the growing vasculature using 2-Photon microscopy. Second, I will use an innovative methodology employing zinc finger nuclease mediated gene targeting to rapidly generate mutant zebrafish in genes implicated in mediating the effects of hemodynamics on angiogenesis. Third, I will laser ablate individual blood vessels in living zebrafish embryos to alter blood flow patterns and study the influence of those changes on angiogenesis. These challenging technological advancements will make it possible for the first time to understand how physiological factors aid in shaping the forming vascular system. These findings will have broad applicability to angiogenic processes in many different vascular beds and developmental time points. Importantly, the mechanisms at play during vascular development in the embryo are reactivated in the adult in situations of pathological angiogenesis, such as tumor progression or ischemia. Therefore, applying this knowledge to angiogenesis occuring in pathological settings will provide us with new avenues for improved disease treatments.
Max ERC Funding
1 423 000 €
Duration
Start date: 2010-10-01, End date: 2015-09-30
Project acronym ZF-BrainReg
Project Regeneration of the adult zebrafish brain
Researcher (PI) Michael Brand
Host Institution (HI) TECHNISCHE UNIVERSITAET DRESDEN
Call Details Advanced Grant (AdG), LS3, ERC-2014-ADG
Summary The goal of this research proposal is to unravel the cellular and molecular mechanisms for the ability of the adult zebrafish brain to regenerate itself after a lesion, and to compare these mechanisms in the non-regenerating mammalian brain. The corresponding mechanisms, if reactivated, may rekindle regeneration also in mammalian brains. Specifically, we focus on identifying the endogeneous stem and progenitor cells that contribute to neural regeneration in zebrafish, by genetic lineage tracing experiments under conditions of regeneration. We have begun to identify the genes and mechanisms controlling regeneration ability in these cells by transcriptome analysis of specific cell types isolated by FACS sorting and transcriptome analysis, which has revealed a key positive role for inflammation as a trigger in regeneration. The resulting candidate genes are functionally tested in adult zebrafish brains for their requirement and sufficiency to elicit or contribute to brain regeneration. If confirmed, we will test for the function of such genes and mechanism in mammalian tissue culture models of regeneration, and determine in adult mouse brain in vivo whether they are candidates to be tested in mammalian brain regeneration. Functional knock-out, knock-in and viral expression tests of such genes and mechanisms in vivo in mice will determine their ability to rekindle regeneration in the lesioned mammalian brain.
This research proposal will provide fundamental insights into the cellular and molecular mechanisms controlling the process of brain regeneration in vertebrates, and will thus suggest avenues for future progenitor cell-based therapies of the injured or diseased human brain.
Summary
The goal of this research proposal is to unravel the cellular and molecular mechanisms for the ability of the adult zebrafish brain to regenerate itself after a lesion, and to compare these mechanisms in the non-regenerating mammalian brain. The corresponding mechanisms, if reactivated, may rekindle regeneration also in mammalian brains. Specifically, we focus on identifying the endogeneous stem and progenitor cells that contribute to neural regeneration in zebrafish, by genetic lineage tracing experiments under conditions of regeneration. We have begun to identify the genes and mechanisms controlling regeneration ability in these cells by transcriptome analysis of specific cell types isolated by FACS sorting and transcriptome analysis, which has revealed a key positive role for inflammation as a trigger in regeneration. The resulting candidate genes are functionally tested in adult zebrafish brains for their requirement and sufficiency to elicit or contribute to brain regeneration. If confirmed, we will test for the function of such genes and mechanism in mammalian tissue culture models of regeneration, and determine in adult mouse brain in vivo whether they are candidates to be tested in mammalian brain regeneration. Functional knock-out, knock-in and viral expression tests of such genes and mechanisms in vivo in mice will determine their ability to rekindle regeneration in the lesioned mammalian brain.
This research proposal will provide fundamental insights into the cellular and molecular mechanisms controlling the process of brain regeneration in vertebrates, and will thus suggest avenues for future progenitor cell-based therapies of the injured or diseased human brain.
Max ERC Funding
2 500 000 €
Duration
Start date: 2016-01-01, End date: 2020-12-31
Project acronym ZF_Blood
Project Less is more: Single Cell Analysis of Zebrafish Blood Development
Researcher (PI) Ana Cvejic
Host Institution (HI) THE CHANCELLOR MASTERS AND SCHOLARSOF THE UNIVERSITY OF CAMBRIDGE
Call Details Starting Grant (StG), LS3, ERC-2015-STG
Summary Blood stem cells need to both perpetuate themselves (self-renew) and differentiate into all mature blood cells to maintain blood formation throughout life. However, it is unclear how the underlying gene regulatory network maintains this population of self-renewing and differentiating stem cells, and how it accommodates the transition from a stem cell to a mature blood cell. Our current knowledge of transcriptomes of various blood cell types has mainly been advanced by population-level analysis. However, the population of seemingly homogenous blood cells may include many distinct cell types with substantially different transcriptomes and abilities to make diverse fate decisions. To overcome these limitations, I will use single-cell transcriptome sequencing of zebrafish blood cells. I will apply an integrative strategy, combining genetic perturbation with computational sequence and network analysis methods, to reconstruct the regulatory networks that maintain the dynamic balance between different blood cell types. This will be achieved by pursuing two main aims:
1) I will create a comprehensive atlas of single cell gene expression in adult zebrafish blood cells and computationally reconstruct the blood lineage tree. I will order cells according to their most likely developmental chronology and identify genes and gene regulatory networks that define distinct cell types. The completion of the first aim will be followed by a more ambitious long-term one that is based on:
2) The in-depth functional characterisation of a subset of novel key regulators of blood formation and identified cell types in vivo. To achieve this I will generate a number of loss-of-function and transgenic zebrafish lines.
By sequencing thousands of single cells, this study is poised to go beyond traditional approaches in examining the complex relationships between the continuous spectra of blood cells, and will provide unprecedented insight into the regulation of blood cell formation.
Summary
Blood stem cells need to both perpetuate themselves (self-renew) and differentiate into all mature blood cells to maintain blood formation throughout life. However, it is unclear how the underlying gene regulatory network maintains this population of self-renewing and differentiating stem cells, and how it accommodates the transition from a stem cell to a mature blood cell. Our current knowledge of transcriptomes of various blood cell types has mainly been advanced by population-level analysis. However, the population of seemingly homogenous blood cells may include many distinct cell types with substantially different transcriptomes and abilities to make diverse fate decisions. To overcome these limitations, I will use single-cell transcriptome sequencing of zebrafish blood cells. I will apply an integrative strategy, combining genetic perturbation with computational sequence and network analysis methods, to reconstruct the regulatory networks that maintain the dynamic balance between different blood cell types. This will be achieved by pursuing two main aims:
1) I will create a comprehensive atlas of single cell gene expression in adult zebrafish blood cells and computationally reconstruct the blood lineage tree. I will order cells according to their most likely developmental chronology and identify genes and gene regulatory networks that define distinct cell types. The completion of the first aim will be followed by a more ambitious long-term one that is based on:
2) The in-depth functional characterisation of a subset of novel key regulators of blood formation and identified cell types in vivo. To achieve this I will generate a number of loss-of-function and transgenic zebrafish lines.
By sequencing thousands of single cells, this study is poised to go beyond traditional approaches in examining the complex relationships between the continuous spectra of blood cells, and will provide unprecedented insight into the regulation of blood cell formation.
Max ERC Funding
1 500 000 €
Duration
Start date: 2016-05-01, End date: 2021-04-30
Project acronym ZMOD
Project Blood Vessel Development and Homeostasis: Identification and Functional Analysis of Genetic Modifiers
Researcher (PI) Didier STAINIER
Host Institution (HI) MAX-PLANCK-GESELLSCHAFT ZUR FORDERUNG DER WISSENSCHAFTEN EV
Call Details Advanced Grant (AdG), LS3, ERC-2015-AdG
Summary The vascular system is a complex network of blood vessels that transports gases, nutrients and hormones throughout the organism. Most blood vessels that form during development and growth arise by the sprouting of new capillaries from pre-existing vessels, a process termed angiogenesis. An imbalance in angiogenesis contributes to the pathogenesis of numerous disease states: insufficient angiogenesis limits tissue recovery in ischemic disease, whereas stimulation of angiogenesis by cancer cells promotes tumor vascularization and growth. Angiogenesis inhibitors are already in clinical use for anti-tumor therapy; however, multiple reports of resistance are calling for the identification of additional targets. Furthermore, vascular malformations are a significant cause of morbidity and mortality. While the genetic basis for some vascular malformations is known, many genetic factors, including modifiers that affect the age-of-onset and severity of phenotypes, remain to be identified. Identifying modifier genes is important not only to fully assess genetic risk, but also to provide novel targets for therapy; however, identifying modifier genes has proven challenging. We recently uncovered a novel and simple way to identify modifier genes. By investigating gene and protein expression differences between knockout (mutant) and knockdown (antisense treated) zebrafish embryos, we found that mutations in specific genes, including some encoding angiogenic factors, lead to the upregulation of compensating (i.e., modifier) genes while knocking down these same genes does not. We hypothesize that the modifier genes identified through this approach in zebrafish also play important roles in humans. Thus, we will use this simple strategy to identify new genes that regulate vascular formation and homeostasis, and subsequently analyze their function in zebrafish as well as in mammalian models, as they are likely to play key roles in vascular development and disease.
Summary
The vascular system is a complex network of blood vessels that transports gases, nutrients and hormones throughout the organism. Most blood vessels that form during development and growth arise by the sprouting of new capillaries from pre-existing vessels, a process termed angiogenesis. An imbalance in angiogenesis contributes to the pathogenesis of numerous disease states: insufficient angiogenesis limits tissue recovery in ischemic disease, whereas stimulation of angiogenesis by cancer cells promotes tumor vascularization and growth. Angiogenesis inhibitors are already in clinical use for anti-tumor therapy; however, multiple reports of resistance are calling for the identification of additional targets. Furthermore, vascular malformations are a significant cause of morbidity and mortality. While the genetic basis for some vascular malformations is known, many genetic factors, including modifiers that affect the age-of-onset and severity of phenotypes, remain to be identified. Identifying modifier genes is important not only to fully assess genetic risk, but also to provide novel targets for therapy; however, identifying modifier genes has proven challenging. We recently uncovered a novel and simple way to identify modifier genes. By investigating gene and protein expression differences between knockout (mutant) and knockdown (antisense treated) zebrafish embryos, we found that mutations in specific genes, including some encoding angiogenic factors, lead to the upregulation of compensating (i.e., modifier) genes while knocking down these same genes does not. We hypothesize that the modifier genes identified through this approach in zebrafish also play important roles in humans. Thus, we will use this simple strategy to identify new genes that regulate vascular formation and homeostasis, and subsequently analyze their function in zebrafish as well as in mammalian models, as they are likely to play key roles in vascular development and disease.
Max ERC Funding
2 500 000 €
Duration
Start date: 2016-10-01, End date: 2021-09-30
