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2020 -
Long-Term Fellowships - LTF

Recording and programming human retina cell fates

LIN Hsiu-Chuan (CHINA, REPUBLIC OF (TAIWAN))

Human Retina and Organoid Development Group - Institute of Molecular and Clinical Ophthalmology Basel - Basel - SWITZERLAND

CAMP Gray (Host supervisor)
It is a longstanding goal to decipher cell fate decisions during human development. Stem cell-based culture systems, combined with single-cell sequencing and CRISPR-Cas technologies, offer exciting possibilities to track human cell identities at unprecedented resolution and to engineer specific cell types with high precision. In this proposal, I will develop novel methods to record cell histories, trace lineages, and program human cell fates, with a focus on the human retina. In the first aim, I will develop a fate recording system in induced pluripotent stem cells (iPSCs) that unites evolvable barcodes with molecular memory recorders (Cas1/2) that capture expressed RNA into DNA. This will be used to simultaneously record real transcriptome and lineage histories along the path of cell fate acquisition. In the second aim, I will adopt the CRISPR-Cas12a(Cpf1) system that enables multiplexed gene activation to engineer cell fates based on predictions learned from single-cell transcriptome dissection of organ development. I will focus my project on human retina development where we have a well-characterized three-dimensional organoid culture system, and extensive data analyzing cell fates that emerge in these organoids. This focus will provide insights into the molecular mechanisms controlling human retinogenesis, and at the same time identify methods to engineer specific retinal cell types for therapeutic screening. The strategies and technologies developed in this project are applicable beyond the retina and will contribute to our understanding of general cell fate decisions in humans and guide cell type engineering with clinical potential.
2020 -
Long-Term Fellowships - LTF

Deciphering the principles of enhancer cooperativity

LOUBIERE Vincent (FRANCE)

- Research Institute of Molecular Pathology - Vienna - AUSTRIA

STARK Alexander (Host supervisor)
Metazoan development relies on tightly regulated gene expression patterns. Hence, the regulatory landscapes of developmental genes are complex and often contain several simultaneously active enhancers. Recent studies reported that the combined enhancer activities in such loci can either be additive, super-additive or sub-additive compared to the sum of the individual activities. Yet, the principles that might govern enhancer cooperativity remain elusive. To fill this gap, I propose measuring the activity of large collections of systematically paired developmental enhancers using STARR-Seq (500x500 enhancers), which was developed in my host lab to map individual enhancers genome-wide. I expect to identify an unprecedented number of additive, super- and sub-additive enhancer pairs and to reveal the sequence features associated with these distinct behaviors. To test all the genomic elements that could potentiate and/or interfere with developmental enhancer activity, I will use another STARR-Seq variant in which selected developmental enhancers will be individually paired with a comprehensive library of genomic DNA fragments (1xall elements). This will address how housekeeping enhancers, which form a functionally distinct class of enhancers, and other types of genomic elements impact developmental enhancer activities. Finally, I will validate my results in situ using CRISPR/Cas9 genome editing and test whether the genes’ endogenous regulatory landscapes influence enhancer cooperativity. Overall, this original approach will address a key aspect of developmental gene regulation which, despite its relevance for the study of development and pathogenesis, remains elusive.
2020 -
Long-Term Fellowships - LTF

Immune-vascular crosstalk in the postnatal period

MADEIRA Silvia (PORTUGAL)

Department of Oncology - University of Lausanne - Lausanne - SWITZERLAND

PETROVA Tatiana (Host supervisor)
The neonatal period is witness to fast physiological changes. The, up until birth, largely sterile environment becomes colonized by microbes, crucial for imprinting durable and specific immune responses, a period named “window of opportunity”. Postnatal immune imprinting involves extensive inter-organ trafficking of immune cells, and hence their interaction with endothelial cells of blood and lymphatic vessels. Still, to date virtually nothing is known about immune-vascular cross talk during postnatal period and the role of endothelial cells in immune imprinting. By using a mouse model that expresses a photoconvertible fluorescent protein, I will first characterize the immune subsets trafficking from gut, an essential organ for postnatal immune education, to other organs. Second, I will study the molecular changes in endothelial cells in this neonatal period to understand how this affects inter-organ communication and the pre-programming of long-lasting immune responses. This will be done by flow cytometry, high resolution 3D imaging and single cell RNA sequencing. Moreover, while there is a significant knowledge on the impact of environmental factors such as nutrition, exposure to antibiotic and farm animals on future health, much less is known about postnatal education of blood and lymphatic vasculature. Accordingly, I will investigate the impact on diet and microbial colonization in the neonatal period and their effects on both immune and vascular components. Altogether, I wish to unveil how early life events impact the immune-vasculature crosstalk and future health imprinting, thus helping to unlock intricate disease mechanism and their prevention in the future.
2020 -
Long-Term Fellowships - LTF

Interplay between membrane tension and branched actin network dynamics during cell migration

MEHIDI Amine (FRANCE)

Department of Biochemistry - University of Geneva - Geneva - SWITZERLAND

AUMEIER Charlotte (Host supervisor)
Cell motility is an integrated process implicated in many cellular functions such as embryogenesis, immunological response, wound healing, and growth cone path finding. For a long time, the study of cell migration has mainly focused on the observation of molecular components responsible for the formation of cell protrusion: actin networks and actin regulatory proteins. Recent studies propose that membrane biophysics, in particular membrane tension, could control actin network formation and cell migrations. Thus, in this project I propose to study the interplay between membrane tension and actin networks during cell migration. To do so, I will use a tension-sensitive fluorescent probe (FliptR probe) to measure membrane tension, together with a cell permeable fluorogenic F-actin label (SiR-actin) suitable for live labelling of the actin cytoskeleton. Using these probes, I will measure the local membrane tension variations during cell migration and monitor how these affect and/or are affected by actin cytoskeleton. By combining these approaches with the cutting-edge technique of adhesion protein micro-patterning, I will create a controlled and reproducible environment to finely tune cell migration. This will allow us to understand how the cellular environment affects the interplay between membrane tension and actin cytoskeleton. All together, I propose an innovative approach that will enable us for the first time to study the impact of local membrane tension on cell migration. The findings of this project will shed light into the cell migration, actin and lipid membrane fields.
2020 -
Long-Term Fellowships - LTF

Macrophage – intestinal epithelial cell crosstalk in the integration of cell death and tissue repair

MELI Alexandre (CANADA)

Department of Immunobiology - Yale University - New Haven - USA

ROTHLIN Carla (Host supervisor)
The intestinal epithelium is a mucosal tissue reliant on its regenerative capacity as it is perpetually exposed to cell death whether it be during homeostasis or insult. Defective regeneration of the epithelium is associated with multiple diseases such as Intestinal Bowel Disease. Macrophages can act to sense damage and enhance tissue repair in the gut. Whether they can integrate the recognition of dying cells with intestinal epithelial regeneration remains unknown. We previously reported that the recognition of apoptotic cells by macrophages enhances their capacity to promote tissue repair. However, depending on the nature of intestinal injury, intestinal cells can undergo divergent cell death modalities including apoptosis, necroptosis and pyroptosis. Thus, we propose a model wherein macrophages directly participate in the integration of cell death with tissue repair. We aim to characterize cell death modalities, including that of stem cells, in models of intestinal injury. We additionally plan to identify the relevant cell death receptors for macrophage detection of apoptotic, necroptotic or pyroptotic cell death and correlate these findings with regeneration/repair or its failure during disease. Current treatment strategies focus on resolving inflammation during intestinal disease instead of enhancing regenerative responses of the epithelium. Thus, uncovering the nature and consequence of cell death in the gut may provide valuable insight into the treatment of gastrointestinal diseases.
2020 -
Long-Term Fellowships - LTF

Investigation of sleep stage-dependent synaptic plasticity in the living brain

WEINHARD Laetitia (FRANCE)

Department of Neuroscience and Physiology - New York University School of Medicine - New York - USA

GAN Wenbiao (Host supervisor)
Sleep is widely acknowledged to be essential to brain function, including learning and memory. Although learning relies on synaptic plasticity, there is surprisingly no consensus on the nature and function of the synaptic changes sleep would mediate. The controversy in the field partly derives from the complexity of sleep structure, as sleep is composed of REM and non-REM stages suggested to hold different functions that are poorly investigated at the synaptic level. To address this issue, I propose to use a recently-developed 3D two-photon microscope in order to continuous monitor synaptic changes during learning and specific sleep stages. The mechanisms underlying sleep stage-dependent synaptic plasticity will be further investigated by combining Ca2+ imaging, light-induced CaMKII inhibitor and optogenetics. The data obtained will provide new insights into the role of sleep in synaptic plasticity, and might further reveal fundamental mechanism underlying learning and memory.
2020 -
Long-Term Fellowships - LTF

Dissecting the molecular basis of human muscle stem cell heterogeneity and quiescence

YAN Lu (CHINA, PEOPLE'S REPUBLIC OF)

Department of Pathology - Brigham and Women's Hospital - Boston - USA

POURQUIE Olivier (Host supervisor)
The regeneration of the adult skeletal muscle relies on the activity of resident muscle stem cells (satellite cells). These PAX7-positive (PAX7+) cells are predominantly quiescent in uninjured muscles, while being able to rapidly proliferate upon injury. Mouse satellite cells are known to be heterogeneous, with subpopulations exhibiting graded quiescent states and variable regenerative potential. However, the molecular identities and developmental trajectories of satellite cell subsets are not well established. Moreover, due to the limited access to human tissues, little is known about the biology of human PAX7+ cells. The host laboratory has recently developed robust differentiation protocols that enable efficient generation of myofibers and PAX7+ cells from pluripotent stem cells in vitro. In this project, I plan to use single-cell transcriptomics to chart the dynamic cellular landscape of human in vitro myogenic differentiation, as well as mouse muscle development in vivo, with the aim of revealing the identities of emerging cells during myogenesis, developmental roadmap for cellular heterogeneity and molecular signatures of quiescence. In addition, preliminary results implicate the retinoic acid (RA) signaling in controlling satellite cell quiescence. Assisted by single-cell profiling, the role of RA will be characterized in detail. To further uncover in an unbiased fashion the gene networks linked to quiescence, I will also conduct functional genomic screens based on the in vitro platform. Overall, this study will use diverse techniques and model systems to provide new insights into muscle stem cell biology and potentially benefit the treatment for muscle diseases.
2020 -
Long-Term Fellowships - LTF

Dissecting the function of temporal variation of gene expression in limb morphogenesis

ZHU Meng (CHINA, PEOPLE'S REPUBLIC OF)

Department of Genetics - Harvard University - Boston - USA

TABIN Cliff (Host supervisor)
Phenotypic evolution concurs with spatial and temporal gene expression changes. Most work have been focused on spatial gene expression variations, yet little has been known about whether the temporal alteration of gene expression, termed heterochrony, also contributes to morphological variations. In this proposed work, I aim to address this question using forelimb (FL) and hindlimb (HL) development in the mouse embryo as a model, due to the ease of experimental manipulation of this system. Although gene regulatory network underlying limb development is highly conserved, the morphology of FL and HL differs. An important distinction in their development is the relative duration of gene expression time – nearly all limb regulators express for a shorter period in HL, correlates with its curtailed differentiation time-scale comparing to FL. Whether and to what extent do such temporal gene expression differences relate to morphological divergence remains unclear. To this end, I will systematically and quantitatively characterize the kinetics of key limb regulators’ expression during the FL and HL development in the mouse embryo; then combining in vitro culture assays and sequencing methods to investigate the mechanisms causing temporal gene expression differences between the two appendages; finally, I will functionally assess the role of such heterochrony to limb morphology by altering the gene expression timing, first experimentally, in mouse and chick embryos; and then theoretically, by using mathematical modelling. Together, this proposed work can advance our understanding of the connection from genotype to phenotype and will also aid biomedical research on limb defects.
2020 -
Long-Term Fellowships - LTF

Mechanisms of cellular dedifferentiation in regeneration

SALINAS SAAVEDRA Miguel (CHILE)

Centre of Chromosomal Biology - National University of Ireland Galway - Galway - IRELAND

FRANK Uri (Host supervisor)
Some animals can regenerate lost organs and tissues with high efficiency. In many such species, cellular dedifferentiation plays a major role in this process. How dedifferentiation is induced in vivo in the correct context is unknown and represents a major question in regenerative biology. The cnidarian Hydractinia can regenerate any lost body part in a tissue-specific manner. Regeneration of some body parts is mediated by adult, migratory stem cells called i-cells that are absent from the intact head. Surprisingly, host lab found that isolated heads can, nevertheless, regenerate a fully functional animal including i-cells that arise by dedifferentiation. Following amputation, isolated heads transiently display features consistent with cellular senescence. Here, I propose to study the mechanisms accompanying injury-induced senescence in Hydractinia dedifferentiation-dependent whole-body regeneration. My approach includes the mechanistic characterization of the molecular/cellular events that occur between decapitation and the appearance of new i-cells through dedifferentiation. For this, I will perform transcriptomic analyses, gene function experiments, and in vivo imaging. The strength of my proposal rests upon the usage of a unique animal model that is highly regenerative, displays natural dedifferentiation, and allows performing in vivo experiments that are impossible to conduct in other animals.
2020 -
Long-Term Fellowships - LTF

Brain remodeling and deceleration of aging upon caste transition in the ant Harpegnathos saltator

SIERIEBRIENNIKOV Bogdan (UKRAINE)

Department of Biochemistry and Molecular Pharmacology - New York University Langone School of Medicine - New York - USA

REINBERG Danny (Host supervisor)
Identical genomes in social insects generate castes with distinct morphologies and behaviors. The Harpegnathos ants have a remarkable lifestyle, whereby the queen’s death triggers transition of a subset of workers to become her substitutes, called gamergates. This entails minor changes in morphology, yet a pronounced shift in behavior, and remarkably, a fivefold extension of lifespan. This process is reversible if the gamergate is placed in a colony with a real queen. The capability of such a dramatic transition in adult life provides a unique context to study neuronal plasticity and aging. Moreover, social insects challenge major aging theories as their calorie intake and fecundity positively correlate with lifespan, in contrast to most animals. I aim to provide an extensive mechanistic description of the worker-to-gamergate transition in H. saltator, and to characterize the mechanisms of the resultant aging deceleration. I will study the brain as it regulates caste transition and produces hormones implicated in aging, while also being affected by both processes. First, I will perform single-cell RNA-sequencing of the worker and nascent gamergate brains to describe changes in genes controlling synaptic wiring and cellular signaling. Next, I will test whether deceleration of aging in gamergates is accompanied by decreased rates of molecular damage, protein synthesis, transposon activity, and telomere shortening. Finally, I will combine both datasets to link transcriptional changes during the caste transition with aging-related molecular processes. Deciphering the paradoxical aging mechanism in social insects may uncover previously unknown strategies to prolong lifespan.
2020 -
Long-Term Fellowships - LTF

A chemical biology approach to unravel phosphatidylethanolamine transport and metabolism

SIMON Clémence (FRANCE)

Biochemistry Department - University of Geneva - Geneva - SWITZERLAND

RIEZMAN Howard (Host supervisor)
Glycerophospholipids are one of the major lipid species in cellular membranes of eukaryotic cells. Among glycerophospholipids, phosphatidylethanolamine (PE) is a highly conserved lipid, found in all organelles but mainly synthesized in the endoplasmic reticulum (ER) and in mitochondria. If it has been shown to play essential roles in membrane protein structure and function, its distribution and metabolism have not been fully elucidated. Indeed, little is known about lipid transport in terms of quantitative and qualitative contribution of each pathway, because methods to measure lipid transport between organelles in cells are lacking. Unlike proteins which are relatively easily manipulated by genetic techniques, individual lipid species cannot be readily modified in vivo. Here, I propose to develop and use chemical tools to study PE transport and metabolism in living cells with high spatiotemporal resolution. I will synthesize photocaged PE which can be localized with precision to mitochondria (or ER) within cells and then released with irradiation of light. Using isotope-labelling, I will trace their transport and quantify the metabolites inside living cells by lipidomics and mass spectrometry approaches. Moreover, mass spectrometry imaging could allow visualizing the de novo labeled phospholipids. This interdisciplinary approach will give us the spatiotemporal resolution that is needed to create for the first time a 4D map of PE metabolism in cells. It will allow us to elucidate its metabolism related to its dependence on intracellular localization and prove that PE can or cannot be transported, for instance, bidirectionally between ER and mitochondria.
2020 -
Long-Term Fellowships - LTF

Force generation and sensation in rapid plant movement

SLEBODA David (USA)

Department of Physiology - McGill University - Montreal - CANADA

SHARIF NAEINI Reza (Host supervisor)
Touch the sensitive plant Mimosa pudica and it rapidly changes shape, folding its leaves to deter herbivory and dislodge pests. This and other feats of rapid plant motion have fascinated scientists for centuries, but the physiological mechanisms underlying them remain poorly understood. I propose a collaborative, interdisciplinary project to elucidate two key aspects of rapid plant motion: sensation of mechanical stimuli and rapid generation of force. I hypothesize that knowledge of the sensory and motor strategies of motile animals can inform our approach to the study of rapid plant motion, and outline a novel experimental approach that draws tools and theory from the fields of biomechanics, neurophysiology, and materials science to explore rapid motion in Mimosa pudica, a model system for the study of rapid plant movement. I have identified an ideal host laboratory in which to carry out this work situated at a nexus of expertise in mechanotransduction, plant physiology, and mechanical modeling. To this environment I will bring expertise in skeletal muscle physiology and the biomechanics of force production. The proposed work forges a connection between seemingly disparate areas of physiological research, will provide unique training that increases my technical and theoretical breadth, and has great potential to elucidate basic principles underlying sensory and motor strategies in biological systems.
2020 -
Long-Term Fellowships - LTF

Dissection of the relationship between folding stability and biological lifetime of proteins

TSUBOYAMA Kotaro (JAPAN)

Department of Pharmacology and Center for Synthetic Biology - Northwestern University - Chicago - USA

ROCKLIN Gabriel (Host supervisor)
Protein degradation systems maintain protein homeostasis. A failure of these systems causes various diseases, such as neurodegenerative diseases and cancers. In eukaryotic cells, ubiquitin is a general marker for selective degradation and determines protein lifetime in vivo. In selective degradation, E3 ubiquitin ligases determine target proteins. Although there are ~800 E3 ligases in human, only a handful of them have already shown to recognize specific short peptide motifs called “degrons”. Moreover, the feature(s) for unstable or misfolded structures recognized by E3 ligases remain unclear. In part, this is because we lack a comprehensive approach to investigate the global relationship between protein structural stability, ubiquitination status, and lifetime of the protein. To reveal the effect of protein folding stability (and other features) on ubiquitination and biological lifetime in vivo, I propose to measure these parameters for thousands of designed mini-proteins, whose folding stability has been previously characterized in detail. First, I will measure biological lifetime for these mini-proteins by flow cytometry, and monitor their ubiquitination status by using top-down proteomics approach. Then, I will analyze these data by using in silico analysis and decipher what factor(s) determine ubiquitination states and biological lifetime. This highly innovative and comprehensive approach using thousands of designed proteins will allow me to uncover the fundamental principle for protein lifetime in vivo and provide a mechanistic basis for designing better tools to manipulate protein lifetime.
2020 -
Long-Term Fellowships - LTF

Context matters: dissecting metabolic heterogeneity in the tumor niche

VAN DER REEST Jiska (NETHERLANDS)

Department of Cell Biology - Harvard Medical School - Boston - USA

HAIGIS Marcia (Host supervisor)
Metabolic transformation is a key characteristic of cancer cells and considered a promising avenue for therapeutic intervention. Nevertheless, metabolic lethalities identified in laboratories often fail to translate to the clinic. This may be because the complexities of tumor metabolism cannot be recapitulated by studying cancer cell autonomous metabolism alone: tumors are not solid masses of cancer cells, but include host tissue, stroma, and immune cells that together shape the tumor metabolic environment. This insight highlights the need to observe and interrogate tumor metabolism in vivo. Metabolic studies rely heavily on mass spectrometry-based analysis of metabolites extracted from cells or tissues, but this approach lacks spatial resolution to resolve metabolic heterogeneity between cells. To address this unmet need, I will develop a transformative platform for single-cell tissue metabolomics using mass spectrometry imaging and use it to study redox metabolism in liver metastatic cancer to investigate a major unanswered question in cancer research: how does the host tissue support metastatic tumor growth? This work contributes a new strategy to incorporate spatial resolution into metabolomics and provides the first comprehensive analysis of the metabolic symbiosis between host and cancer cells in vivo. It aims to redefine the traditional approach to metabolic studies that can progress our understanding of metabolic heterogeneity. Focusing on liver metastatic cancer, I aim to provide novel mechanistic understanding of redox-metabolic crosstalk between cells in the tumor-host niche and explore strategies to interfere with redox metabolism to combat metastasis.
2020 -
Long-Term Fellowships - LTF

Systems-level circuit implementation of memory-based action selection

VENKATASUBRAMANIAN Lalanti (INDIA)

Department of Zoology - University of Cambridge - Cambridge - UK

ZLATIC Marta (Host supervisor)
Living organisms constantly make important decisions in their dynamic environments in order to survive. Animals make effective decisions in response to sensory stimuli by recruiting both innate and learned information in their nervous systems, thereby guiding them to approach or avoid cues that are appetitive or aversive respectively. Importantly, in order to adapt to a changing environment, animals also need to update their memories over time. Understanding how the nervous system invokes previously stored information to dictate future decisions has been a central and historical goal in neuroscience. Recently, significant progress has been made in identifying the connectivity and functionality of specific circuits involved in distinct aspects of learning and memory. However, a challenging task has been to obtain a comprehensive and systems-level understanding of neural circuit dynamics by which memories are retained, extinguished or expanded and integrated with innate information to eventually drive various behavioural actions. In this proposal I aim to address this problem by using a novel setup to record the activities of multiple individual neurons that function at different levels downstream of the learning and memory centre in the insect brain in freely behaving animals subject to specific tasks over time. Importantly, by combining this data with synaptic-level circuit maps generated by electron microscopy, mathematical modelling to produce reliable circuit motifs, and precise optogenetic manipulations to test the models in vivo, we will obtain both a global and high-resolution understanding of memory-based and context-dependent action-selection.
2020 -
Long-Term Fellowships - LTF

Epigenetic plasticity and imprinting dynamics during development

WEINBERG-SHUKRON Ariella (ISRAEL)

Department of Genetics - University of Cambridge - Cambridge - UK

FERGUSON-SMITH Anne C. (Host supervisor)
Epigenetic modifications regulate development. In particular, genome-wide DNA methylation patterns undergo dynamic changes at cornerstones of mammalian development. While DNA methylation is essential for normal embryogenesis, how it regulates and maintains cell fate and function in vivo remains unclear. A key methylation-regulated developmental process is parental imprinting and perturbations to methylation imprints result in embryonic lethality. We discovered that paternal deletion of an essential imprinting control region in mammals results in a temporal-specific compensatory methylation switch onto the opposite maternal allele, minimising detrimental developmental defects. This suggests a novel paradigm of epigenetic plasticity with wider implications for the epigenetic control of genome function. Here, we will utilize cutting-edge DNA and RNA single-cell sequencing methods and in vivo knockout models to study the functional roles of parental-origin specific epigenetic plasticity during development. We will determine whether loss of imprinting drives compensating changes on the other allele and decipher the associated mechanism of regulation, maintenance and hierarchical interactions. We will investigate the genome-wide plasticity of imprinting by relating imprinted gene dosage dynamics with developmental time points and tissue expression, with specific focus on particular developmental niches. Our approach will elucidate a new avenue of epigenetic control during development and disease, contribute to our understanding of the evolution and flexibility of imprinting and provide novel insights into regenerative medicine, reproduction and epigenetic inheritance.
2020 -
Cross Disciplinary Fellowships - CDF

Characterization of the sorting platform's assembly in bacteria using 4Pi microscopy and DNA-PAINT

SCHUEDER Florian (GERMANY)

Department Cell Biology and Department of Microbial Pathogenesis - Yale University - New Haven - USA

GALAN Jorge E. (Host supervisor)
Many bacteria have evolved protein machineries to inject proteins into eukaryotic or prokaryotic cells. The type III secretion system (T3SS), widely distributed among gram-negative bacteria, is made up of a needle complex, an export apparatus and a sorting platform (SP). Although, the whole injectosome was characterized in a recent study, the assembly process of the SP is poorly understood. It has been shown that SpaO (a core component of the SP) localizes in two spatial populations, one at the bacterial membrane and another one located within the bacterial cytosol. This observation of the presence of SpaO-associated clusters away from the bacterial membrane suggests the presence of fully or partially assembled SPs in the bacterial cytoplasm. Furthermore, there are two existing versions of SpaO, a short (SpaOS) and a long (SpaOL) version. It has been postulated that SpaOS plays a role in chaperoning SpaOL during the assembly process of the SP but is not a structural part of the SP itself. If this holds true, SpaOS should only be localized in the (partly assembled) SPs located in the bacterial cytosol. In order to understand the assembly of the SP and the role of SpaOS in this process, I will image all proteins of the SP using recently developed 4Pi microscopy together with DNA-PAINT super-resolution microscopy. This novel combination, will enable me to image all proteins of the SP with molecular resolution (5 nm isotropic resolution). This will be an ambitious step towards understanding the sequence of events leading to the assembly of the SP in the T3SS. I anticipate that the tools developed for this work will bring new technologies to the field of bacterial physiology.
2020 -
Cross Disciplinary Fellowships - CDF

Biomechanical induction of a primitive streak in a synthetic human embryo

VALET Manon (FRANCE)

Center for Studies in Physics and Biology - The Rockefeller University - New York - USA

BRIVANLOU Ali (Host supervisor)
During gastrulation, embryonic stem cells are primed towards three germ layers: the ectoderm, the mesoderm and the endoderm. While acquiring their fate, the cells re-arrange massively and the embryo polarizes with the appearance of an antero-posterior axis. The signals that are necessary for this patterning and the role of the mechanical forces remain poorly understood, in particular for the human embryo. Synthetic biology has thus developed new models of embryos using human stem cell lines (hESCs). We propose to study the effect of differential mechanical tension on the appearance of a primitive streak in a synthetic human embryo. We will investigate the relationship between the WNT pathway activation and mechanical instability, following recent results on a biomechanical regulation of this pathway. We will first work on the correlation between the substrate induced curvature and WNT expression in 2D cultures of human stem cells. Secondly, we will map the resulting stress field of the cells to this expression pattern. Thirdly, we will study cellular self-organization under mechanical load in a 3D model synthetic embryo. We will eventually focus on the inducer role of a second cell population in the polarization of this synthetic embryo. In the four cases we will correlate the resulting dynamic features with the expression of markers relative to the three germ layers. We hope to provide a first quantitative understanding of the relationship between the force field and the differentiation of stem cells in a model synthetic embryo. This project will be co-supervised by Dr. Eric Siggia and Dr. Ali Brivanlou. The experiments will be performed in the Brivanlou lab.
2020 -
Cross Disciplinary Fellowships - CDF

Mycobacterium tuberculosis modulation of host, elucidated by super-resolution imaging and proteomics

ASHDOWN George (UK)

Division of Infectious Diseases and Immune Defence - Walter and Eliza Hall Institute of Medical Research - Melbourne - AUSTRALIA

COUSSENS Anna (Host supervisor)
Mycobacterium tuberculosis (Mtb) survival and replication within, or escape from, its phagocytic host requires molecular exploitation of host proteins and lipids. However, there are still large gaps in our understanding of bacterial survival and escape, particularly regarding quantitative analysis of interactions between host and Mtb derived proteins or lipids. Super-resolution fluorescence imaging has begun to yield unprecedented insight into biological processes including protein distribution at the nanometer scale and live dynamics of proteins, lipids and the cytoskeleton. However, these techniques have rarely been applied to Mtb research. Here, super-resolution imaging complimented with proteomics will allow the spatiotemporal analysis of membrane associated proteins, including pathogen derived proteins which interact with the intra- and extracellular milieu of primary human macrophages and neutrophils. Work will also utilise live-cell imaging sensitive to membrane composition to elucidate how the pathogen modulates host cell lipids for survival and replication. Additionally, how Mtb escapes its host for transmission is poorly understood. Mtb modulation of the host’s cytoskeleton, membrane and linker proteins using high- and super-resolution imaging, will clarify this process. Quantifying the distribution or disruption of host proteins and lipids through the host-pathogen lifecycle, and investigating whether Mtb proteins themselves present on the host plasma membrane will translate to application of therapeutics. This work will generate novel hypotheses for the Mtb field, and other pathogens, in the context of host immunity and disease equilibrium.
2020 -
Cross Disciplinary Fellowships - CDF

Global Neuronal Network theory vs Integrated Information Theory

BENDTZ Katarina (SWEDEN)

Children's Hospital - Harvard University - Boston - USA

KREIMAN Gabriel (Host supervisor)
Understanding how consciousness arises from neural activity in the human brain remains one of the greatest scientific challenges of modern times. Two of the most influential theories of the neural correlates of consciousness (NCC) are Global Neuronal Workspace (GNW) theory and Integrated Information Theory (IIT). GNW posits that what we experience as a conscious state is the global broadcasting of information in an interconnected neuronal network. IIT instead defines a measure for integrated information, known as phi (f), which can theoretically be used to locate this physical substrate within the brain. The empirical research on these theories have been focused on the visual modality alone, even though the theories make predictions of the full conscious experience, where all sensory modalities necessarily must be included. For example, very little attention has been directed towards the auditory modality. These theories have also been developed separately without much interaction between them and the empirical research has been focused on testing each theory individually, rather than contrasting them to evaluate their explanatory and predictive power. The research question of this project is which of these two theories better explains the NCC, using an adversarial and multi-modal approach designed to cover a maximal space of the testable predictions of these models. It is adversarial in the sense that the strongest advocators for both theories are involved in the project, and it is multi-modal in the sense that all experiments are tested in both the visual and the auditory modality.