The shape and architecture of every plant depends on how its cells grow. The outer membrane of every plant cell is surrounded by a wall made of cellulose fibres embedded in a matrix, and neighbouring cells are stuck together so they cannot move. Despite these constraints, plants can generate remarkable shapes, from orchid flowers to tree canopies. These forms arise through a dynamic process in which the pressure within each cell causes the walls to stretch irreversibly, a process known as creep. Wall thickness is maintained through synthesis of new layers of fibers at the cell membrane, and partitioning walls are also added, preventing cells from becoming too large. Although the pressure in each cell acts equally in all directions, the wall fibers are not randomly arranged, causing cells to creep more in some orientations than others. The secret of plant shape therefore lies in how the cell walls are structured to yield with specific rates and orientations. Although much progress has been made in understanding how genes control these processes, we still lack a quantitative understanding of how growth of even a single plant cell is controlled. One difficulty is that cell wall properties depend on their history of formation, which is usually unknown. A further problem is that growth is modified by mechanical constraints and signals from neighbouring cells. This project aims to circumvent these problems by exploiting a simplified system in which the formation of a new wall can be followed from scratch. Protoplasts are single plant cells in which the walls have been digested away. Under the right conditions, protoplasts will regenerate their walls and exhibit oriented growth. Using the state-of-the-art techniques, we will quantify and perturb different components of the wall as it is made and measure its mechanical properties as it strengthens and begins to undergo creep. We will also synthesize simplified artificial plant cells and cellulose nanofiber networks to test hypotheses for how walls acquire their mechanical properties and the feedback mechanisms involved. By exploring hypotheses through computational modelling, we will evaluate which best predict experimental results and thus arrive at an integrated quantitative understanding of cell wall synthesis, assembly, mechanics and growth that underpins plant development.

Phyllotaxis, the regular arrangement of leaves around stems, is one of the most striking natural patterns; it has puzzled biologists, physicists and mathematicians for centuries. Phyllotaxis first evolved in simple plants, like the moss Physcomitrium Patens, but has mostly been studied in plants of recent evolutionary origin, like Arabidopsis. In contrast with the multicellular Arabidopsis shoot apex, successive rotating division planes of a single apical cell directly determine moss phyllotaxis, with each apical cell derivative generating directly a leaf. This provides a system to understand how the geometry of a single apical cell and its daughter cells, their resultant physical forces and biochemical cues self-organize 4D patterns of division orientation and ultimately shape a shoot. To explore the fundamental question of how phyllotaxis emerged, at single cell-resolution, we will use our unique inter-disciplinary expertise to combine developmental genetics, optical and physical imaging, single cell genomics, optogenetics and computational modeling in moss. This will generate key insights into the contribution of cell division orientation to the evolution of phyllotaxis.

Bones serve as a mineral reservoir in vertebrates to achieve calcium and phosphate homeostasis. However, the precise cellular regulation of this process is not fully understood. Osteocytes (Oy), the most abundant bone cells, form an interconnected dendritic network embedded in the bone tissue through a pore system called the lacuna-canalicular network (LCN). Oy are believed to orchestrate mineral release and uptake either indirectly, by triggering remodeling via bone resorbing/forming cells, or directly, through a localized process, known as osteocytic osteolysis (OO). Although postulated in the 1960-70s, the relative importance of OO for mineral homeostasis is unclear and its mechanisms still poorly described. Most studies on calcium transport focus on the LCN, although there is growing evidence that a complex sub-cellular mineralization pathway exists. In this proposal, we challenge the classical view that the LCN alone determines calcium transport and hypothesize the existence of an intermediate level of mesoscale porosity that plays a central role in calcium exchange. Our scientific approach uses interdisciplinary expertise in materials science, physics and biomedical engineering, and integrates a series of multiscale and multimodal imaging platforms connected through deep-learning and the application of connectomics. Here, we will: i) decipher the mesoscale porosity and mineral heterogeneity at the LCN, ii) determine the spatial distribution of OO, and iii) produce a connectomics analysis of fluid transport and calcium exchange in bone. We will use lactating and weaning C57BL6 mice that exhibit large reversible mineral depletion/remineralization. We will acquire bone ultrastructure visualization in 3D using an emerging plasma focused ion beam scanning electron microscope. This will be combined with confocal and two-photon excitation fluorescence microscopy using an original deep-learning super-resolution correlative imaging approach. Finally, multiscale network connectivity will be analyzed using connectomic approaches based on graph theory and experimental data will be used to perform numerical simulations of fluid transport. This research will reveal the structural mechanisms and extent of OO demineralization/remineralization at the cellular and sub-cellular scale and identify key parameters affecting fluid transport during OO.

A high level of oxygen in the atmosphere is taken for granted as it is indispensable for human and most animal life. Yet, life (bacteria and unicellular eukaryotes) evolved for over 2 billion years in a low oxygen environment. Oxygen became widely available only in the last 600 million years and was likely the main factor behind the rapid diversification of most complex multicellular life forms. For elaborate multicellular tissues to form, their overall shapes however must still satisfy oxygen demands of individual cells. Oxygen consumption will rapidly reduce the oxygen level within a few cell depth into the tissue. This becomes a problem in early embryonic development before respiratory and circulatory systems are established. Many cells still need to differentiate and migrate for long distances and place themselves at the right destination. It is largely unknown what makes the varying oxygen environment and the execution of their precise differentiation and navigation within a tissue compatible. The project aims at elucidating the common basis of oxygen dependency in morphogenesis by studying how the social amoeba Dictyostelium detects and reacts to oxygen at the molecular level and how this dictates cell migration and morphogenesis. Dictyostelium is an organism evolutionarily close to the branches of multicellular evolution, and presents enormous advantages to be easily cultured in the laboratory and reversibly switched from unicellular to multicellular states in less than a day. Amoeboid cells are able to change their state (by differentiation) and place themselves in more favorable conditions for foraging and respiration (by migration). An emphasis is placed on clarifying this most fundamental and primordial characteristic with a focus on the cross-scale quantitative relationship between oxygen, metabolic states, cytoskeletal machineries and migratory modes. New screening methods based on aerotaxis responses, oxygen sensors and 3D microscopy will be combined with molecular genetics of oxygen response pathways. The role of the new mechanism unearthed will further be tested for their roles in neural crest cell migration in zebrafish. They will be formulated into multi-scale mathematical models to clarify how the coupling between oxygen consumption, sensing and migration of individual cells gives rise to robust and collective morphogenetic movements.

Plants have a remarkable ability to undergo transitions during their life in a robust and synchronized fashion. They are able to achieve this despite the environment they are in being highly variable, and not having sophisticated information processing systems like brains. A striking example of how plants use variable signals such as temperature to time a critical developmental transition is the breaking of bud dormancy in in long-lived trees that grow in boreal and temperate regions of the world. In these trees, growth stops in the autumn and a state of dormancy is established that prevents reactivation of leaf formation which is timed to coincide with the advent of spring. The timing of dormancy break so the growth can be restarted is highly critical decision since a premature release from dormancy exposes plants to damage from sudden frosts in the early spring, whereas restarting growing too late will compromise fitness and productivity due to reduction in the length of their growing season. How trees accurately time the break of bud dormancy so that growth can start again in spring is question that has puzzled biologists for over 100 years, and will be addressed in this project. This will be achieved by exploring parallels between how human engineered computers process information, and the cells in tree buds process temperature. The genes which control bud dormancy have recently been described, yet how they operate within the multicellular context of a bud tissue remains unknown. We propose that controlled communication between cells plays a central role in the release from bud dormancy, and their ability to generate robust responses. A synergistic combination of biological experiments, computer modelling, physics and 3D image analysis, will provide unprecedented insight into how tree buds process temperature information. The project will lead to the identification of algorithms invoked by trees and the molecular and cellular logic used by trees to control bud dormancy, thereby revealing the operating system used by a plant to control its growth over the course of its life cycle.

Sexual imprinting is a process of instinctive learning occurring during early postnatal development, through which many animal species acquire memories of the odor, vocalizations, and other characteristics of their parents (or siblings), and then utilize this information to select their mates as adults. Sexual imprinting has evolved in many taxa, and yet surprisingly almost nothing is known about the neural mechanisms controlling this type of learning. What neural circuits are involved in this process? How is information from multiple sensory modalities integrated, and do they have synergistic effects on female preferences? Laboratory mice have proven to be useful models to investigate sexual imprinting, but do these findings generalize to outbred, wild house mice? To answer these questions and decipher whether sexual imprinting represents a general mechanism guiding mate selection in females, we will take a multidisciplinary approach using behavioral, neuroanatomical and advanced optical imaging techniques. We will conduct cross-fostering experiments, and apply whole brain activity pattern reconstruction and in vivo functional imaging to address several specific aims: i) experimentally test the synergistic effects of acoustic and olfactory cues on sexual imprinting; ii) reconstruct the brain regions activated by sexual imprinting; and iii) functionally trace the circuits that influence female mating preferences. To address these aims we created a novel collaboration between three research groups with very different expertise. The multidisciplinary research capitalizes on recent advances in optical imaging and neuroanatomy, including light-sheet whole-brain imaging and highly innovative in vivo technology for simultaneous imaging on multiple brain regions. We aim to reveal the organization of the circuits that are shaped during postnatal development to form memories of conspecific that are recalled in adults, which enhance inbreeding avoidance and reproductive success.

The state of brain circuits is changed by neuromodulatory signals such as dopamine, serotonin or acetylcholine, which are released in spatially and temporally precise, phasic or tonic patterns depending on the physiological context. These neuromodulators act on multiple receptor classes, exerting diverse physiological effects through distinct signaling pathways. Obtaining a clearer picture of the function of neuromodulatory-driven signals in neural information processing and plasticity requires methods for remote-control of specific neurotransmitter receptors with high temporal precision in defined neurons and in behaving animals. We combine labs with expertise in synthetic chemistry and photochemistry (Ellis-Davies) and behavioral neuropharmacology (Mourot) in order to develop technologies to gain optical control over DA neuromodulation in the freely-moving mouse, for studies of neural circuits and behaviors associated with same-sex social interactions. Synthetic photochemical tools (caged and photoswitchable compounds) are, in principle, able to mimic the timing, amplitude and spread of naturally occurring neuronal signals. However, their in vivo use has been highly restricted, notably because these probes are stimulated by UV or visible light that penetrates tissue very poorly. To overcome this issue, we propose to use lanthanide-doped upconverting nanoparticles (UCNPs), which will be functionalized with caged/photoswitchable ligands for glutamate and dopamine receptors, enabling non-invasive control of the very receptors of the brain in behaving mice. We will further combine near-infrared photocontrol with chemo-genetic strategies to achieve optical control of receptors in defined neuronal targets. Leveraging UCNPs in this way will provide a platform for the interrogation of dopamine-related neuropsychiatric disorders in groups of animals in naturalistic environments, and will represent a milestone in our understanding of how dopamine impacts a variety of personality traits and same-sex interactions. The strategies developed here will be applicable to most receptors, neural circuits and animal models, thus our project bears enormous potential in changing the landscape of our understanding of neuromodulation, both in health and disease.

Directed evolution is a technique to investigate evolutionary dynamics and evolve nucleic acids and proteins for research, diagnostic and therapeutic applications. It uses cycles of mutation and screening or selection to mimic Darwinian Evolution. The aim of these experiments is to screen or select large libraries of molecule variants with different sequences (‘genotypes’) and retain those sequences having the required properties (or required ‘phenotypes’). At the moment one essential limitation of Directed Evolution methods is that the properties of each variant in the library are assessed based on bulk samples, i.e. on the average activity of a large numbers of copies of the same molecule. While these copies are identical at the sequence level, they may still exhibit different phenotypes, for example due to the fact that the same sequence can assume multiple three dimensional structures. These ‘heterogeneous’ phenotypes can play important roles in evolutionary dynamics, but at the moment there is no convenient Directed Evolution tool to address them. Here we combine expertise from the fields of directed evolution and single-molecule microscopy to develop a novel approach that allows us to perform directed evolution while looking at the behaviour of single-molecules. The method can be used to perform accurate phenotypic characterization measuring multiple aspects of the phenotype in parallel, something that can be challenging with conventional approaches. For example it can be used to study how a molecule with a specific sequence can sometimes play a role in different chemical reactions, how to achieve ‘specialization’ and what the consequences of specialization are in terms of efficiency. As discussed above, the method can also reveal the existence of phenotypic heterogeneity in molecules with identical sequence. This approach will allow us to address several biological questions that cannot be addressed with conventional methods. These include, the evolution of heterogeneous phenotypes in populations of RNA molecules under selection for binding, the role of heterogeneity in defining the evolutionary potential of antibodies and the correlation between rate and accuracy in DNA polymerases. It will also provide a potent route to develop biomolecules for biomedical and industrial applications, notably in the field of single-molecule technologies.

Uncontrolled loss of neurons is a hallmark of neurodegenerative diseases (NDs) such as Alzheimer’s disease and amyotrophic lateral sclerosis (ALS). In NDs, an abnormal assembly of proteins and mRNAs known as a biomolecular condensate (BC) is believed to contribute to neurotoxicity and ensuing neuronal death. Despite high mortality and poor quality of patients’ lives, as well as the enormous socio-economic burden of patient care, the exact molecular and cellular mechanisms underlying ND pathogenesis remain largely unknown. This roadblock in ND studies primarily originates from mutual dependency between “molecular assembly” of general ND proteins and “functionality” of individual ND proteins. Molecular assembly is a dynamic process that can take place locally, and changes actively in its physical property by transitioning among soluble phase, liquid-like droplets, hydrogels and insoluble aggregates. To our knowledge, there is currently no technique to disassemble ND-related BCs to distinguish these physical and biochemical characteristics, especially in a physiologically relevant context. By integrating expertise in neurophysiology, chemical biology, and soft-matter biophysics, we propose to develop a conceptually unique methodology termed “dePOLYMER”. Here, light illumination or chemical administration actuates a genetically-encoded molecular probe designed to polymerize actin to generate constrictive force against an intended object such as BCs in living cells. When force density exceeds the surface tension of target BCs, dePOLYMER is expected to trigger their dispersion. We will specifically target stress granules as a model for non-pathogenic BCs, as well as pathogenic ALS-related BCs. Using a model for ALS, we will next aim to demonstrate dispersion of BCs consisting of ALS proteins in a manner specific for constituent proteins and cell types. In summary, the goal of our study is to offer a novel technique to efficiently disperse BCs in a rapidly inducible manner. Implementation of dePOLYMER in ALS model systems may enable future identification of the molecular and cellular foundation of neurotoxicity, thus a potentially new treatment strategy for NDs. Due to the inherently modular design of dePOLYMER probes, the technique should be generalizable to virtually all BCs whose molecular components are known, let alone other ND-related BCs.

How organs control their growth to achieve the correct final size is an enduring mystery of biology. In wildtype Drosophila, for example, left and right wing areas rarely differ by more than 1%. We previously found that the hormone Dilp8 controls this developmental precision, as in absence of the hormone, the fluctuating asymmetry (FA) of bilateral organs is strongly increased. In this proposal, we hypothesize that the hormone Dilp8 and cell death cooperate to limit wing size variability. To test this idea and explore its consequences, we will integrate methods from developmental biology with quantitative data analysis and modeling from physics. In particular, we will show how changes in cell death in dilp8 mutants are quantitatively related to changes in final wing size. Our results will provide new insights into how organ size is controlled and refined in biological systems.

Sporadically, novel and potentially devastating pandemic influenza A viruses (IAVs) are generated through genome reassortment between human and animal co-infecting IAVs. Such pandemic viruses emerge as a consequence of the segmentation of IAVs genome into a bundle made of 8 distinct viral RNAs (vRNAs). However the molecular mechanisms of vRNA intracellular transport and assembly into vRNAs bundles, which are critical for reassortment, remain largely unknown. Our project aims to elucidate these fundamental aspects of IAV life cycle by developing innovative approaches.
We challenge the original model that newly synthesized vRNAs, in the form of viral ribonucleoproteins (vRNPs), are transported across the cytoplasm on Rab11-dependent recycling endosomes. Based on our recent work, we hypothesize that the concomitant transport and assembly of vRNPs is driven by their physical association with remodelled endoplasmic reticulum (ER) membranes and Rab11-dependent transport vesicles distinct from recycling endosomes.
We will set up a cellular system which resembles the natural respiratory tissue targeted by IAVs, while being amenable to simultaneous imaging of the endogenous Rab11 protein and tagged vRNAs. We will develop two cutting-edge and complementary imaging methods: dual-color single molecule fluorescence in situ hybridization (FISH) in live cells for the tracking of distinct vRNAs that diffuse concomitantly, and cryo-Focused Ion Beam combined with electron microscopy in situ hybridization (EMISH) to image individual vRNPs and their transport vesicles at molecular resolution. We will further assess the role of cellular ER-shaping proteins by performing CRISPR/Cas9-mediated knockdowns, and by monitoring changes in the viral-induced remodelling of ER and biogenesis of vRNP transport vesicles by live fluorescence imaging and cellular EM.
The proposed research will require the very close collaboration between three partners with distinct but complementary expertise. The approaches developed jointly are poised to revolutionize our understanding of IAV multi-RNA genome transport and bundling, and thus help in the broader goal of achieving better prevention and treatment of influenza disease. Additionally, the proposed technical developments in live cell FISH and cellular EM, will have impact on other fields of studies well beyond the scope of the proposed project.

Optical nanostructures are highly organized composites of materials with varying refractive indices (e.g. keratin, melanin and air) that produce some of the brightest colors found in nature through coherent light scattering. How these tissues organise themselves at the nanometer scale to produce colors is poorly understood, despite its fundamental significance to developmental and evolutionary biology and potential to spark advances in the biomimetic design and "green" commercial manufacture of self-assembling optical materials.
We thus propose to use both transcriptomic, laser diffraction and microscopy-based tools of developmental biology to elucidate the mechanisms by which these nanostructures self-assemble in a subsample of birds (Class Aves), a group with incredibly diverse structural colors and mechanisms. Our working hypothesis is that iridescent colors form through depletion-attraction, phase separation and other self-assembly mechanisms. Because most developmental biology is done at larger size scales, testing these hypotheses will require the use and development of methods such as wet cell TEM and in situ laser diffraction analysis to adequately resolve nanometer-scale changes in developing tissue. We will then test these proposed mechanisms using biomimetic approaches that replicate natural conditions as closely as possible (e.g. at room temperature,at biological pH) using natural or semi-natural materials. Use of optical techniques including angle-resolved spectrophotometry and microspectrophotometry will enable us to compare these properties between the natural and synthetic versions. This approach will enable us to not only experimentally test modes of development but also generate and test new materials and/or processes to produce them.
There are three highly innovative aspects to this proposal. First, it attempts to unlock the developmental pathways producing nanostructured tissues. This is a long-standing question with few answers thus far. Second, it uses biomimicry in novel ways to test developmental hypotheses and pushes the technical boundaries of developmental biology by focusing on nanometer-scale organisation of tissues. Finally, the use of biologically realistic chemistry in our biomimetic approaches is a huge leap forward in this field where most work is done at high temperature or with non-biocompatible materials. This work will therefore significantly advance both our fundamental understanding of these materials and the tools to study them and other nanoscale materials.

We propose to investigate how dietary restrictions (DR) affect aging from a new angle by using social insects as models. Aging is a hallmark of most bilateria and most animals balance their reproduction rate against lifespan. Intriguingly, this trade-off is inverted in reproductive individuals (queens) of social insects (termites, ants, bees). Whereas most studies on aging directly manipulate the lifespan, e.g. of mice or worms or other lab-bred animals, we here propose a radically new approach by employing easily accessible and natural extremely long-lived termite queens as models. Their metabolism, response to DR and fertility will be gauged against genomically identical bu infertile and short lived workers, as well as shorter lived and less fecund queen of a closely related termite species. We will sample termite colonies directly from the field, keep them, expose them to DR and measure their fitness and fecundity. We will examine the role of DR during colony development by sampling transcriptomes, analyzing their epigenetic status, their metabolome and endocrine status and performing in-depth molecular analyses of key molecular components that are known to be implicated in regulating aging and fecundity. Using multiple OMICS methods, reverse genetics, hormonal and dietary administration we will be able to disentangle pathways involved in development of queens and measure the impact of energetic metabolic reprogramming on fitness and reproduction status. Expression patterns and spatio-temporal changes of genetic networks will be used to develop a simple state model. In this model, the metabolic status can be used to predict an individual's trajectory of aging and fecundity depending on its epigenetically imprinted background such as it's caste. Our project thus establishes a new model system for studying the relationship between DR, aging and fecundity, in which the latter two are decoupled and comparison of our model to other model organisms will help understand which dependencies and molecular components have universally conserved interaction partners or phenotypic effects. The project is possible only due to the four participants from three continents, with expertise in dietary research, energy metabolism, field research and social insect genomics.

Organ development depends on the integration of local cell-to-cell interactions with long range signalling throughout the body. We will investigate mechanisms by which long-range signalling via the cerebrospinal fluid (CSF) regulate body axis formation and spine curvature. The CSF is produced by the choroid plexus in the brain ventricles and flows down the central canal in the spinal cord. The CSF instructs brain development by delivering age-dependent grow-promoting factors to target cells. CSF circulation also contributes to the curvature of body axis (embryo) and to the spine (juveniles). However, the mechanisms regulating the flow and content of CSF remain poorly understood. Progress has been impeded by a historical lack of tools and challenges inherent to studying fluids in small, developing organisms. The advent of innovative tools and approaches now provides an unprecedented opportunity to overcome previous limitations. The transparency of zebrafish and the accessibility to mouse CSF provide powerful models for testing our driving hypothesis that the physical and biochemical properties of CSF impact body axis formation and spine curvature. We propose an entirely new, interdisciplinary collaboration of three leaders in their field: (1) physicist Francois Gallaire (EPFL, Switzerland), expert in the theory & modelling of complex fluid dynamics; (2) developmental neurobiologist Maria Lehtinen (Boston Children’s, HMS, USA), expert in analysis of the choroid plexus-CSF system; (3) biophysicist Claire Wyart (ICM, France), expert in imaging and sensory physiology in the spinal cord. Our multi-tiered approach will unravel the principal parameters driving CSF composition and flow. We will map CSF flow in the developing fourth ventricle and central canal (Aim 1), elucidate mechanisms regulating protein secretion into the CSF (Aim 2), and investigate mechanisms of active transport of instructive signals along the anteroposterior axis controlling organogenesis (Aim 3). The proposed studies will transform this historically understudied area of neuroscience into a robust field spanning CSF-based signalling in brain and spine. As studies of paracrine signalling and fluid dynamics lag far behind cell-intrinsic studies of signalling, our techniques and concepts should provide a roadmap for future studies of fluid niches throughout the vertebrate body.

Organ development depends on the integration of local cell-to-cell interactions with long range signalling throughout the body. We will investigate mechanisms by which long-range signalling via the cerebrospinal fluid (CSF) regulate body axis formation and spine curvature. The CSF is produced by the choroid plexus in the brain ventricles and flows down the central canal in the spinal cord. The CSF instructs brain development by delivering age-dependent grow-promoting factors to target cells. CSF circulation also contributes to the curvature of body axis (embryo) and to the spine (juveniles). However, the mechanisms regulating the flow and content of CSF remain poorly understood. Progress has been impeded by a historical lack of tools and challenges inherent to studying fluids in small, developing organisms. The advent of innovative tools and approaches now provides an unprecedented opportunity to overcome previous limitations. The transparency of zebrafish and the accessibility to mouse CSF provide powerful models for testing our driving hypothesis that the physical and biochemical properties of CSF impact body axis formation and spine curvature. We propose an entirely new, interdisciplinary collaboration of three leaders in their field: (1) physicist Francois Gallaire (EPFL, Switzerland), expert in the theory & modelling of complex fluid dynamics; (2) developmental neurobiologist Maria Lehtinen (Boston Children’s, HMS, USA), expert in analysis of the choroid plexus-CSF system; (3) biophysicist Claire Wyart (ICM, France), expert in imaging and sensory physiology in the spinal cord. Our multi-tiered approach will unravel the principal parameters driving CSF composition and flow. We will map CSF flow in the developing fourth ventricle and central canal (Aim 1), elucidate mechanisms regulating protein secretion into the CSF (Aim 2), and investigate mechanisms of active transport of instructive signals along the anteroposterior axis controlling organogenesis (Aim 3). The proposed studies will transform this historically understudied area of neuroscience into a robust field spanning CSF-based signalling in brain and spine. As studies of paracrine signalling and fluid dynamics lag far behind cell-intrinsic studies of signalling, our techniques and concepts should provide a roadmap for future studies of fluid niches throughout the vertebrate body.

Plant development and growth are linked to cellular shape changes, which are controlled by genetic programs but also by perception of environmental signals, including mechanical cues. While both genetic regulation and mechanical control of morphogenesis were studied independently, there is a need to explore how cellular shape-associated strain and stress can mechanically regulate gene expression during differentiation. In animals, mechanical stimuli are known effectors of differentiation. They involve propagation of mechanical forces through the cytoskeleton to the nucleus, leading to chromatin remodeling and modification of gene expression. Thus, nuclear envelope proteins that control nuclear shape and transmit forces to chromatin play a key role in rapid triggering of gene expression. In plants, less is known about mechanotransduction from cell surface to the nucleus.
Using a systems biology approach and an interdisciplinary network, we propose to investigate how mechanical cues affecting cellular shaping are sensed at the nuclear envelope to drive chromatin remodeling in Arabidopsis. We will sudy a unique cellular model, the single root hair in an epidermal tissue context, with well-defined morphogenetic programs linked to cytoskeleton and nuclear dynamics. We will analyze root hair formation and growth in WT and mutants affected in either root hair development or nuclear shape. Combining in vivo live imaging and micro-mechanical measurements (rheometry), we will evaluate mechanical properties of cells and nuclei during root hair development and their dependence on cytoskeleton and nuclear dynamics in relation to gene expression. We will also determine how these mechanical, structural and biological properties are modified when a controlled mechanical stress is applied to the root hair cell during development. Our data will highlight proteins involved in mechanosensing, and we will evaluate their interaction with the nuclear envelope network. Live imaging and rheometry data will be correlated to finite element modeling to estimate strain and stress in the system for predicting chromatin remodeling following cellular and nuclear shape changes.
Altogether, this will highlight the molecular networks involved in mechanosensing at the nucleo-cytoplamic interface and reveal how gene expression is robustly regulated during cellular morphogenesis in higher plants.

Plant development and growth are linked to cellular shape changes, which are controlled by genetic programs but also by perception of environmental signals, including mechanical cues. While both genetic regulation and mechanical control of morphogenesis were studied independently, there is a need to explore how cellular shape-associated strain and stress can mechanically regulate gene expression during differentiation. In animals, mechanical stimuli are known effectors of differentiation. They involve propagation of mechanical forces through the cytoskeleton to the nucleus, leading to chromatin remodeling and modification of gene expression. Thus, nuclear envelope proteins that control nuclear shape and transmit forces to chromatin play a key role in rapid triggering of gene expression. In plants, less is known about mechanotransduction from cell surface to the nucleus.
Using a systems biology approach and an interdisciplinary network, we propose to investigate how mechanical cues affecting cellular shaping are sensed at the nuclear envelope to drive chromatin remodeling in Arabidopsis. We will sudy a unique cellular model, the single root hair in an epidermal tissue context, with well-defined morphogenetic programs linked to cytoskeleton and nuclear dynamics. We will analyze root hair formation and growth in WT and mutants affected in either root hair development or nuclear shape. Combining in vivo live imaging and micro-mechanical measurements (rheometry), we will evaluate mechanical properties of cells and nuclei during root hair development and their dependence on cytoskeleton and nuclear dynamics in relation to gene expression. We will also determine how these mechanical, structural and biological properties are modified when a controlled mechanical stress is applied to the root hair cell during development. Our data will highlight proteins involved in mechanosensing, and we will evaluate their interaction with the nuclear envelope network. Live imaging and rheometry data will be correlated to finite element modeling to estimate strain and stress in the system for predicting chromatin remodeling following cellular and nuclear shape changes.
Altogether, this will highlight the molecular networks involved in mechanosensing at the nucleo-cytoplamic interface and reveal how gene expression is robustly regulated during cellular morphogenesis in higher plants.

Plant development and growth are linked to cellular shape changes, which are controlled by genetic programs but also by perception of environmental signals, including mechanical cues. While both genetic regulation and mechanical control of morphogenesis were studied independently, there is a need to explore how cellular shape-associated strain and stress can mechanically regulate gene expression during differentiation. In animals, mechanical stimuli are known effectors of differentiation. They involve propagation of mechanical forces through the cytoskeleton to the nucleus, leading to chromatin remodeling and modification of gene expression. Thus, nuclear envelope proteins that control nuclear shape and transmit forces to chromatin play a key role in rapid triggering of gene expression. In plants, less is known about mechanotransduction from cell surface to the nucleus.
Using a systems biology approach and an interdisciplinary network, we propose to investigate how mechanical cues affecting cellular shaping are sensed at the nuclear envelope to drive chromatin remodeling in Arabidopsis. We will sudy a unique cellular model, the single root hair in an epidermal tissue context, with well-defined morphogenetic programs linked to cytoskeleton and nuclear dynamics. We will analyze root hair formation and growth in WT and mutants affected in either root hair development or nuclear shape. Combining in vivo live imaging and micro-mechanical measurements (rheometry), we will evaluate mechanical properties of cells and nuclei during root hair development and their dependence on cytoskeleton and nuclear dynamics in relation to gene expression. We will also determine how these mechanical, structural and biological properties are modified when a controlled mechanical stress is applied to the root hair cell during development. Our data will highlight proteins involved in mechanosensing, and we will evaluate their interaction with the nuclear envelope network. Live imaging and rheometry data will be correlated to finite element modeling to estimate strain and stress in the system for predicting chromatin remodeling following cellular and nuclear shape changes.
Altogether, this will highlight the molecular networks involved in mechanosensing at the nucleo-cytoplamic interface and reveal how gene expression is robustly regulated during cellular morphogenesis in higher plants.

Plant development and growth are linked to cellular shape changes, which are controlled by genetic programs but also by perception of environmental signals, including mechanical cues. While both genetic regulation and mechanical control of morphogenesis were studied independently, there is a need to explore how cellular shape-associated strain and stress can mechanically regulate gene expression during differentiation. In animals, mechanical stimuli are known effectors of differentiation. They involve propagation of mechanical forces through the cytoskeleton to the nucleus, leading to chromatin remodeling and modification of gene expression. Thus, nuclear envelope proteins that control nuclear shape and transmit forces to chromatin play a key role in rapid triggering of gene expression. In plants, less is known about mechanotransduction from cell surface to the nucleus.
Using a systems biology approach and an interdisciplinary network, we propose to investigate how mechanical cues affecting cellular shaping are sensed at the nuclear envelope to drive chromatin remodeling in Arabidopsis. We will sudy a unique cellular model, the single root hair in an epidermal tissue context, with well-defined morphogenetic programs linked to cytoskeleton and nuclear dynamics. We will analyze root hair formation and growth in WT and mutants affected in either root hair development or nuclear shape. Combining in vivo live imaging and micro-mechanical measurements (rheometry), we will evaluate mechanical properties of cells and nuclei during root hair development and their dependence on cytoskeleton and nuclear dynamics in relation to gene expression. We will also determine how these mechanical, structural and biological properties are modified when a controlled mechanical stress is applied to the root hair cell during development. Our data will highlight proteins involved in mechanosensing, and we will evaluate their interaction with the nuclear envelope network. Live imaging and rheometry data will be correlated to finite element modeling to estimate strain and stress in the system for predicting chromatin remodeling following cellular and nuclear shape changes.
Altogether, this will highlight the molecular networks involved in mechanosensing at the nucleo-cytoplamic interface and reveal how gene expression is robustly regulated during cellular morphogenesis in higher plants.

The human brain gains much of its computational abilities from the trillions of connections made between cells by synapses. Molecular changes in synapses (a process called synaptic plasticity) are considered to underlie learning and memory. An important component of the synapse is the postsynaptic density (PSD). This specialized structure has been studied extensively to understand its function, in particular because >100 neurologic disorders (such as autism spectrum disorder and schizophrenia) have been associated with PSD dysfunction. Information regarding the overall molecular architecture of the PSD, however, is largely incomplete, in part because the PSD is extremely complex, containing hundreds of individual components connected in a dense network. The PSD is also highly dynamic and asymmetrical—two properties that render protein structural analyses challenging. While solving the structure of the entire PSD seems insurmountable, half of the PSD’s mass is composed of only ten classes of linker proteins. We hypothesize that these highly-abundant proteins form a molecular scaffold, a network we term the ‘postsynaptic scaffold’ (PSS). We aim to develop an approach we call the ‘Thermophile-Assisted Postsynaptic Architecture Strategy’ (TAPAS) to solve the PSS structure. Given the delicate nature of proteins, structural biologists like to work with resilient, temperature-resistant proteins that can be obtained from thermophilic organisms. For brain-derived proteins, there is one animal known to be comfortable at temperatures >50 °C: a worm that lives on hydrothermal vents in the Pacific Ocean. We plan to sequence this worm’s genome (Alvinella pompejana) to find the PSS proteins that it shares with humans. We will determine the structures of the thermophilic proteins by X-ray crystallography. As much of the PSS architecture consists of filamentous structures that do not crystallize, we will use cryo-electron microscopy to define these larger structures. Finally, we will integrate several other methods (biochemical, proteomic, bioinformatic and advanced electron tomography methods) to build a model of the overall PSS. Achieving this ambitious goal will inform studies into learning and memory, lead to new treatments for devastating brain disorders, and help explain how the synapse contributes to human cognition.