Cell Biology (Affolter)Head of Research Unit Prof. Dr.Markus AffolterOverviewMembersPublicationsProjects & CollaborationsProjects & Collaborations OverviewMembersPublicationsProjects & Collaborations Projects & Collaborations 23 foundShow per page10 10 20 50 Implementing 3Rs in Switzerland: an interdisciplinary in-depth exploration of barriers and facilitators [Implement-3R] Research Project | 10 Project MembersThe 3R principles and regulation of animal research The use of animals in biomedical and other research presents an ethical dilemma: we do not want to lose scientific benefits, nor do we want to cause laboratory animals to suffer". In Switzerland, the majority of animals (64%) is used in fundamental research and a minority (ca. 20%) are used for developing and testing pharmaceutical and chemical products. The 3R-strategies ("replace, reduce, refine") are "today widely accepted by scientists as a moral obligation to treat animals humanely and if possible to use alternative methods in experiments". The national and international regulatory framework concerning the use of animals for research stipulates adherence to the 3R principles[3-5]. In addition, Swiss law requires researchers to demonstrate a favourable harm-benefit ratio to justify animal experiments[6, 7]. There is an ongoing discussion in the fields of ethics, law and science concerning the interpretation of the 3Rs and harm-benefit analysis. Eminent ethicists have recently proposed a more elaborated, argued way how to balance social benefit and animal welfare in this context. Cell Morphogenesis in Organ-Specific Microvasculature Research Project | 1 Project MembersNo Description available Angiogenesis in zebrafish: how are vascular networks formed? Research Project | 1 Project MembersNo Description available Angiogenesis in zebrafish: how do vascular networks form? Research Project | 1 Project Membersto be added ... Drosophila Branching 2.0 Research Project | 4 Project MembersHow animals acquire their distinct and species-specific shapes and three-dimensional morphologies and how such morphologies are encoded in the genome has been a fascinating topic for numerous scientists over many decades. While the question of how animal form and diversity is achieved is fascinating, gaining a cellular and molecular understanding of the phenomenon of morphogenesis requires breaking down the problem into more precise or better defined questions, such as how individual parts of an animal, for example external extremities or internal organs reach their precise shape during development.My laboratory has tried to answer questions related to the overall topic of animal morphogenesis by using genetic approaches, mainly in Drosophila melanogaster, the fruit fly, and more recently also in Danio rerio, the zebrafish. In this grant application, we would like to capitalise on recent developments in our lab and answer two sets of questions that have preoccupied us over a long period. We and others have studied tracheal development in drosophila as a paradigm for a complex morphogenesis process. We have shown that cell rearrangements are key to generate the distinct morphologies of different tracheal branches in the drosophila tracheal system, and that these cell rearrangements are controlled by transcription factors such as Spalt and Knirps and by cellular activities such as cell migration, cell shape changes and adherens junction remodelling. However, we know very little about the targets of these transcriptional regulators and their cellular and molecular functions in regulating cell rearrangements. We propose to undertake transcriptional profiling to unravel the key role of the Spalt transcription factor. Profiting from the efficiency of Crispr-based methods in drosophila and the short generation time, functional studies of candidate genes can then rapidly be performed and the data integrated into the existing molecular networks. While we hope to isolate more genes involved in the control of cell rearrangement starting from transcriptional profiling, it is very likely that many genes or proteins involved in tracheal morphogenesis have not been identified using classical zygotic loss of function mutagenesis, due to maternal contribution. We have recently developed a nanobody-based method allowing for the depletion of maternally provided proteins in specific tissues and study their role in morphogenesis processes. Using this novel approach, we would like to undertake a pioneering pilot screen to search for proteins with hitherto uncharacterized roles in the tracheal system and in the amnioserosa, a central force-producer during the tissue morphogenesis process of dorsal closure. Furthermore, and inspired by the wealth of nanobody-based novel tools we have already designed and used in drosophila, we want to introduce and validate novel protein-directed approaches based on short peptide tags and the corresponding binding proteins reported to be functional in cultured cells. If these tools work well in multicellular organisms where they have not been tested yet, a wealth of novel applications in developmental biology will emerge, which will help us studying candidate genes from our screens.Question 1: How is the process of cell intercalation regulated at the molecular level?Aim 1: Identify Spalt-regulated genes using RNAseq and study their role in cell intercalationQuestion 2: Can we identify many more proteins involved in tracheal branching morphogenesis, whose contribution has not yet been identified? Aim 2: Identify novel genes (proteins) regulating tracheal morphogenesis using deGradFP.Question 3: Can short peptides be used for in vivo tagging and subsequent protein manipulation?Aim 3: Generate/validate novel tools to directly study proteins in morphogenesis processes Anti-angiogenic Effects of BAL078 in Zebra-Fish Research Project | 1 Project MembersNo Description available Investigating organ patterning and growth by the Dpp/BMP morphogen gradient using novel synthetic receptors Research Project | 2 Project MembersCell-cell communication in developing organs is critical to acquire proper cell fate and organ size. It is therefore not surprising that defects in such communications cause birth defects or disease such as cancer. Despite its importance, it remains largely unknown how cells communicate with each other to build our functional organs. A class of molecules that mediates cell-cell communication during development is called "morphogens". Morphogens are diffusible molecules that disperse and form a concentration gradient in developing organs to control patterning and growth. For over 20 years, organ development of Drosophila melanogaster has served as a leading model to study the organ development through morphogens. Decapentaplegic (Dpp), a Bone morphogenetic protein (BMP) type ligand in flies, represents the first validated secreted morphogens to control patterning and growth of wing imaginal disc (wing precursor tissue). Previous studies have identified Dpp signaling components, downstream target genes, factors required for Dpp morphogen gradient formation. Recent studies combine mathematical approaches to model how these identified factors work together as a system to build organs. However, these bottom-up approaches are still far from understanding how morphogen acts. Therefore, in this proposal, I would like to take an alternative top-down approach to directly and acutely manipulate the endogenous Dpp morphogen gradient at the protein level. Keys to achieving this goal are protein binders such as nanobodies or DARPins (Designed Ankyrin Repeat Proteins) with strong binding affinity against their targets like antibodies. Unlike conventional antibodies, protein binders function as single domain proteins. Therefore, protein binders can be expressed and functionalized in the cells by fusing them with a functional domain of other proteins. Recently, I generated (1) a robust genome engineering platform to insert a tag or introduce mutations in the endogenous dpp gene, and (2) novel synthetic trap system consisting of protein binders (scFv, nanobodies, DARPins) fused to the transmembrane domain of CD8 to trap Dpp on the cell surface. By combining these novel tools, it is now possible to visualize and directly manipulate endogenous Dpp morphogen gradient at the protein level.Using these novel tools, I will ask the following specific questions:(1) What is the spatial and temporal requirement of Dpp dispersal for patterning and growth of wing disc?(2) What is the mechanism of Dpp gradient formation (exocytosis, dispersal, and endocytosis)(3) What is the mechanism of scaling of the Dpp morphogen gradient with tissue size?Direct manipulation of morphogen gradient using the synthetic receptors would allow me to distinguish between a variety of models proposed for how Dpp acts as a morphogen to control organ development. Furthermore, I am open to see unexpected results that require changing our current understanding of morphogen function. Since a variety of evolutionarily conserved signaling pathways have been repeatedly used as morphogens during development, my approach would give technological and biological impact to study cell-cell communications in general. Given recent attempts reconstructing functional human organs in vitro, our approaches should also be applicable to investigate how human organs develop and how defects in cell-cell communications lead to malformations and diseases. Manipulating Dpp/BMP morphogen gradient by a novel synthetic receptor system Research Project | 1 Project MembersNo Description available In vivo cell biology of organ morphogenesis Research Project | 1 Project Membersto be added Roche financing employment of Verena Küppers Research Project | 2 Project Membersto be added 123 123 OverviewMembersPublicationsProjects & Collaborations
Projects & Collaborations 23 foundShow per page10 10 20 50 Implementing 3Rs in Switzerland: an interdisciplinary in-depth exploration of barriers and facilitators [Implement-3R] Research Project | 10 Project MembersThe 3R principles and regulation of animal research The use of animals in biomedical and other research presents an ethical dilemma: we do not want to lose scientific benefits, nor do we want to cause laboratory animals to suffer". In Switzerland, the majority of animals (64%) is used in fundamental research and a minority (ca. 20%) are used for developing and testing pharmaceutical and chemical products. The 3R-strategies ("replace, reduce, refine") are "today widely accepted by scientists as a moral obligation to treat animals humanely and if possible to use alternative methods in experiments". The national and international regulatory framework concerning the use of animals for research stipulates adherence to the 3R principles[3-5]. In addition, Swiss law requires researchers to demonstrate a favourable harm-benefit ratio to justify animal experiments[6, 7]. There is an ongoing discussion in the fields of ethics, law and science concerning the interpretation of the 3Rs and harm-benefit analysis. Eminent ethicists have recently proposed a more elaborated, argued way how to balance social benefit and animal welfare in this context. Cell Morphogenesis in Organ-Specific Microvasculature Research Project | 1 Project MembersNo Description available Angiogenesis in zebrafish: how are vascular networks formed? Research Project | 1 Project MembersNo Description available Angiogenesis in zebrafish: how do vascular networks form? Research Project | 1 Project Membersto be added ... Drosophila Branching 2.0 Research Project | 4 Project MembersHow animals acquire their distinct and species-specific shapes and three-dimensional morphologies and how such morphologies are encoded in the genome has been a fascinating topic for numerous scientists over many decades. While the question of how animal form and diversity is achieved is fascinating, gaining a cellular and molecular understanding of the phenomenon of morphogenesis requires breaking down the problem into more precise or better defined questions, such as how individual parts of an animal, for example external extremities or internal organs reach their precise shape during development.My laboratory has tried to answer questions related to the overall topic of animal morphogenesis by using genetic approaches, mainly in Drosophila melanogaster, the fruit fly, and more recently also in Danio rerio, the zebrafish. In this grant application, we would like to capitalise on recent developments in our lab and answer two sets of questions that have preoccupied us over a long period. We and others have studied tracheal development in drosophila as a paradigm for a complex morphogenesis process. We have shown that cell rearrangements are key to generate the distinct morphologies of different tracheal branches in the drosophila tracheal system, and that these cell rearrangements are controlled by transcription factors such as Spalt and Knirps and by cellular activities such as cell migration, cell shape changes and adherens junction remodelling. However, we know very little about the targets of these transcriptional regulators and their cellular and molecular functions in regulating cell rearrangements. We propose to undertake transcriptional profiling to unravel the key role of the Spalt transcription factor. Profiting from the efficiency of Crispr-based methods in drosophila and the short generation time, functional studies of candidate genes can then rapidly be performed and the data integrated into the existing molecular networks. While we hope to isolate more genes involved in the control of cell rearrangement starting from transcriptional profiling, it is very likely that many genes or proteins involved in tracheal morphogenesis have not been identified using classical zygotic loss of function mutagenesis, due to maternal contribution. We have recently developed a nanobody-based method allowing for the depletion of maternally provided proteins in specific tissues and study their role in morphogenesis processes. Using this novel approach, we would like to undertake a pioneering pilot screen to search for proteins with hitherto uncharacterized roles in the tracheal system and in the amnioserosa, a central force-producer during the tissue morphogenesis process of dorsal closure. Furthermore, and inspired by the wealth of nanobody-based novel tools we have already designed and used in drosophila, we want to introduce and validate novel protein-directed approaches based on short peptide tags and the corresponding binding proteins reported to be functional in cultured cells. If these tools work well in multicellular organisms where they have not been tested yet, a wealth of novel applications in developmental biology will emerge, which will help us studying candidate genes from our screens.Question 1: How is the process of cell intercalation regulated at the molecular level?Aim 1: Identify Spalt-regulated genes using RNAseq and study their role in cell intercalationQuestion 2: Can we identify many more proteins involved in tracheal branching morphogenesis, whose contribution has not yet been identified? Aim 2: Identify novel genes (proteins) regulating tracheal morphogenesis using deGradFP.Question 3: Can short peptides be used for in vivo tagging and subsequent protein manipulation?Aim 3: Generate/validate novel tools to directly study proteins in morphogenesis processes Anti-angiogenic Effects of BAL078 in Zebra-Fish Research Project | 1 Project MembersNo Description available Investigating organ patterning and growth by the Dpp/BMP morphogen gradient using novel synthetic receptors Research Project | 2 Project MembersCell-cell communication in developing organs is critical to acquire proper cell fate and organ size. It is therefore not surprising that defects in such communications cause birth defects or disease such as cancer. Despite its importance, it remains largely unknown how cells communicate with each other to build our functional organs. A class of molecules that mediates cell-cell communication during development is called "morphogens". Morphogens are diffusible molecules that disperse and form a concentration gradient in developing organs to control patterning and growth. For over 20 years, organ development of Drosophila melanogaster has served as a leading model to study the organ development through morphogens. Decapentaplegic (Dpp), a Bone morphogenetic protein (BMP) type ligand in flies, represents the first validated secreted morphogens to control patterning and growth of wing imaginal disc (wing precursor tissue). Previous studies have identified Dpp signaling components, downstream target genes, factors required for Dpp morphogen gradient formation. Recent studies combine mathematical approaches to model how these identified factors work together as a system to build organs. However, these bottom-up approaches are still far from understanding how morphogen acts. Therefore, in this proposal, I would like to take an alternative top-down approach to directly and acutely manipulate the endogenous Dpp morphogen gradient at the protein level. Keys to achieving this goal are protein binders such as nanobodies or DARPins (Designed Ankyrin Repeat Proteins) with strong binding affinity against their targets like antibodies. Unlike conventional antibodies, protein binders function as single domain proteins. Therefore, protein binders can be expressed and functionalized in the cells by fusing them with a functional domain of other proteins. Recently, I generated (1) a robust genome engineering platform to insert a tag or introduce mutations in the endogenous dpp gene, and (2) novel synthetic trap system consisting of protein binders (scFv, nanobodies, DARPins) fused to the transmembrane domain of CD8 to trap Dpp on the cell surface. By combining these novel tools, it is now possible to visualize and directly manipulate endogenous Dpp morphogen gradient at the protein level.Using these novel tools, I will ask the following specific questions:(1) What is the spatial and temporal requirement of Dpp dispersal for patterning and growth of wing disc?(2) What is the mechanism of Dpp gradient formation (exocytosis, dispersal, and endocytosis)(3) What is the mechanism of scaling of the Dpp morphogen gradient with tissue size?Direct manipulation of morphogen gradient using the synthetic receptors would allow me to distinguish between a variety of models proposed for how Dpp acts as a morphogen to control organ development. Furthermore, I am open to see unexpected results that require changing our current understanding of morphogen function. Since a variety of evolutionarily conserved signaling pathways have been repeatedly used as morphogens during development, my approach would give technological and biological impact to study cell-cell communications in general. Given recent attempts reconstructing functional human organs in vitro, our approaches should also be applicable to investigate how human organs develop and how defects in cell-cell communications lead to malformations and diseases. Manipulating Dpp/BMP morphogen gradient by a novel synthetic receptor system Research Project | 1 Project MembersNo Description available In vivo cell biology of organ morphogenesis Research Project | 1 Project Membersto be added Roche financing employment of Verena Küppers Research Project | 2 Project Membersto be added 123 123
Implementing 3Rs in Switzerland: an interdisciplinary in-depth exploration of barriers and facilitators [Implement-3R] Research Project | 10 Project MembersThe 3R principles and regulation of animal research The use of animals in biomedical and other research presents an ethical dilemma: we do not want to lose scientific benefits, nor do we want to cause laboratory animals to suffer". In Switzerland, the majority of animals (64%) is used in fundamental research and a minority (ca. 20%) are used for developing and testing pharmaceutical and chemical products. The 3R-strategies ("replace, reduce, refine") are "today widely accepted by scientists as a moral obligation to treat animals humanely and if possible to use alternative methods in experiments". The national and international regulatory framework concerning the use of animals for research stipulates adherence to the 3R principles[3-5]. In addition, Swiss law requires researchers to demonstrate a favourable harm-benefit ratio to justify animal experiments[6, 7]. There is an ongoing discussion in the fields of ethics, law and science concerning the interpretation of the 3Rs and harm-benefit analysis. Eminent ethicists have recently proposed a more elaborated, argued way how to balance social benefit and animal welfare in this context.
Cell Morphogenesis in Organ-Specific Microvasculature Research Project | 1 Project MembersNo Description available
Angiogenesis in zebrafish: how are vascular networks formed? Research Project | 1 Project MembersNo Description available
Angiogenesis in zebrafish: how do vascular networks form? Research Project | 1 Project Membersto be added ...
Drosophila Branching 2.0 Research Project | 4 Project MembersHow animals acquire their distinct and species-specific shapes and three-dimensional morphologies and how such morphologies are encoded in the genome has been a fascinating topic for numerous scientists over many decades. While the question of how animal form and diversity is achieved is fascinating, gaining a cellular and molecular understanding of the phenomenon of morphogenesis requires breaking down the problem into more precise or better defined questions, such as how individual parts of an animal, for example external extremities or internal organs reach their precise shape during development.My laboratory has tried to answer questions related to the overall topic of animal morphogenesis by using genetic approaches, mainly in Drosophila melanogaster, the fruit fly, and more recently also in Danio rerio, the zebrafish. In this grant application, we would like to capitalise on recent developments in our lab and answer two sets of questions that have preoccupied us over a long period. We and others have studied tracheal development in drosophila as a paradigm for a complex morphogenesis process. We have shown that cell rearrangements are key to generate the distinct morphologies of different tracheal branches in the drosophila tracheal system, and that these cell rearrangements are controlled by transcription factors such as Spalt and Knirps and by cellular activities such as cell migration, cell shape changes and adherens junction remodelling. However, we know very little about the targets of these transcriptional regulators and their cellular and molecular functions in regulating cell rearrangements. We propose to undertake transcriptional profiling to unravel the key role of the Spalt transcription factor. Profiting from the efficiency of Crispr-based methods in drosophila and the short generation time, functional studies of candidate genes can then rapidly be performed and the data integrated into the existing molecular networks. While we hope to isolate more genes involved in the control of cell rearrangement starting from transcriptional profiling, it is very likely that many genes or proteins involved in tracheal morphogenesis have not been identified using classical zygotic loss of function mutagenesis, due to maternal contribution. We have recently developed a nanobody-based method allowing for the depletion of maternally provided proteins in specific tissues and study their role in morphogenesis processes. Using this novel approach, we would like to undertake a pioneering pilot screen to search for proteins with hitherto uncharacterized roles in the tracheal system and in the amnioserosa, a central force-producer during the tissue morphogenesis process of dorsal closure. Furthermore, and inspired by the wealth of nanobody-based novel tools we have already designed and used in drosophila, we want to introduce and validate novel protein-directed approaches based on short peptide tags and the corresponding binding proteins reported to be functional in cultured cells. If these tools work well in multicellular organisms where they have not been tested yet, a wealth of novel applications in developmental biology will emerge, which will help us studying candidate genes from our screens.Question 1: How is the process of cell intercalation regulated at the molecular level?Aim 1: Identify Spalt-regulated genes using RNAseq and study their role in cell intercalationQuestion 2: Can we identify many more proteins involved in tracheal branching morphogenesis, whose contribution has not yet been identified? Aim 2: Identify novel genes (proteins) regulating tracheal morphogenesis using deGradFP.Question 3: Can short peptides be used for in vivo tagging and subsequent protein manipulation?Aim 3: Generate/validate novel tools to directly study proteins in morphogenesis processes
Anti-angiogenic Effects of BAL078 in Zebra-Fish Research Project | 1 Project MembersNo Description available
Investigating organ patterning and growth by the Dpp/BMP morphogen gradient using novel synthetic receptors Research Project | 2 Project MembersCell-cell communication in developing organs is critical to acquire proper cell fate and organ size. It is therefore not surprising that defects in such communications cause birth defects or disease such as cancer. Despite its importance, it remains largely unknown how cells communicate with each other to build our functional organs. A class of molecules that mediates cell-cell communication during development is called "morphogens". Morphogens are diffusible molecules that disperse and form a concentration gradient in developing organs to control patterning and growth. For over 20 years, organ development of Drosophila melanogaster has served as a leading model to study the organ development through morphogens. Decapentaplegic (Dpp), a Bone morphogenetic protein (BMP) type ligand in flies, represents the first validated secreted morphogens to control patterning and growth of wing imaginal disc (wing precursor tissue). Previous studies have identified Dpp signaling components, downstream target genes, factors required for Dpp morphogen gradient formation. Recent studies combine mathematical approaches to model how these identified factors work together as a system to build organs. However, these bottom-up approaches are still far from understanding how morphogen acts. Therefore, in this proposal, I would like to take an alternative top-down approach to directly and acutely manipulate the endogenous Dpp morphogen gradient at the protein level. Keys to achieving this goal are protein binders such as nanobodies or DARPins (Designed Ankyrin Repeat Proteins) with strong binding affinity against their targets like antibodies. Unlike conventional antibodies, protein binders function as single domain proteins. Therefore, protein binders can be expressed and functionalized in the cells by fusing them with a functional domain of other proteins. Recently, I generated (1) a robust genome engineering platform to insert a tag or introduce mutations in the endogenous dpp gene, and (2) novel synthetic trap system consisting of protein binders (scFv, nanobodies, DARPins) fused to the transmembrane domain of CD8 to trap Dpp on the cell surface. By combining these novel tools, it is now possible to visualize and directly manipulate endogenous Dpp morphogen gradient at the protein level.Using these novel tools, I will ask the following specific questions:(1) What is the spatial and temporal requirement of Dpp dispersal for patterning and growth of wing disc?(2) What is the mechanism of Dpp gradient formation (exocytosis, dispersal, and endocytosis)(3) What is the mechanism of scaling of the Dpp morphogen gradient with tissue size?Direct manipulation of morphogen gradient using the synthetic receptors would allow me to distinguish between a variety of models proposed for how Dpp acts as a morphogen to control organ development. Furthermore, I am open to see unexpected results that require changing our current understanding of morphogen function. Since a variety of evolutionarily conserved signaling pathways have been repeatedly used as morphogens during development, my approach would give technological and biological impact to study cell-cell communications in general. Given recent attempts reconstructing functional human organs in vitro, our approaches should also be applicable to investigate how human organs develop and how defects in cell-cell communications lead to malformations and diseases.
Manipulating Dpp/BMP morphogen gradient by a novel synthetic receptor system Research Project | 1 Project MembersNo Description available