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2019 -
Long-Term Fellowships - LTF

The role of neuro-immunological memory in response to enteric infections

AHRENDS Tomasz (POLAND)

Laboratory of Mucosal Immunology - Rockefeller University - New York - USA

MUCIDA Daniel (Host supervisor)

The gastrointestinal (GI) tract is efficiently organized to protect the host from potential dangerous stimuli and to tolerate commensal microbiota and food antigens. It hosts as many neurons as the spinal cord and more immune cells than all other compartments together. The innervation of the digestive tract is involved in determining the patterns of its movements, control of gastric acid secretion, release of gut hormones, modifying nutrient handling and interacting with the gut immune system. Infections of the GI tract can result in neuronal damage and dysregulation of these functions. The host laboratory has recently demonstrated that upon infection, in response to specific neurotransmitters, macrophages – a subset of innate immune cells – can acquire tissue-protective phenotype and minimize neuronal loss. Both immune and nervous systems are equipped with mechanisms allowing to store and recall information on previously encountered events. On this basis, I aim to examine whether GI infections trigger a neuro-immunological memory supporting tissue repair or response to subsequent infections. This is of fundamental and clinical interest, as interactions between immune and neuronal cells are proposed to be part of several disease processes, ranging from multiple sclerosis to irritable bowel syndrome (IBS).

2019 -
Long-Term Fellowships - LTF

The role of tanycytes in temperature and metabolic regulation

AMBROZIAK Wojciech (POLAND)

Institute of Pharmacology - University of Heidelberg - Heidelberg - GERMANY

SIEMENS Jan (Host supervisor)

Hypothalamic tanycytes are sensory glia-like cells lining the walls of the 3rd ventricle (3V) that were proposed to respond to a variety of metabolic and environmental stimuli. The preoptic area (POA) is a hypothalamic region involved in thermoregulation whose nuclei are distributed around the 3V. The POA receives input from peripheral temperature receptors but also contains neurons that respond to deep-brain thermal changes, although the exact mechanism of this phenomenon is not clear. Due to the privileged position of tanycytes, with their somata directly contacting the cerebrospinal fluid and processes reaching portal blood vessels, we propose that tanycytes are part of the central temperature-sensing circuitry. This hypothesis is supported by the fact that they express two receptors, TRPM5 and TRPM3, which are known to be temperature-sensitive. In this project I will use transgenic mouse lines and employ a combination of electrophysiological, chemogenetic, optogenetic and imaging techniques to explore the sensory roles of tanycytes, mostly focusing on thermodetection. I will start with characterising sensory capabilities of tanycytes including their responsiveness to temperature changes and analysing their functional diversity. Then I will test whether tanycytes communicate with neurons in the POA and if so, I will attempt to identify the phenotype of these neurons. Lastly, I will perform in vivo experiments to test the effect of chemogenetic activation and inhibition of temperature-sensitive tanycytes on thermal effector responses as well as eating behaviour. Overall, I expect my study to shed light on the role tanycytes play in temperature detection and homeostasis.

2019 -
Grant Awardees - Program Grants

The repeatability of the genetic mechanisms underlying behavioral evolution

ANDERSEN Erik C. (USA)

Dept. of Molecular Biosciences - Northwestern University - Evanston - USA

BROWN Andre (CANADA)

MRC London Institute of Medical Sciences - Imperial College London - London - UK

HODGINS Kathryn (CANADA)

School of Biological Sciences - Monash University - Clayton - AUSTRALIA

Keen observers of nature have often wondered why diverse species seem to behave similarly. For example, different species of Hawaiian spiders spin similar web architectures, diverse anoles lizard species bob their heads with the same styles and speeds, and distinct species of damselflies avoid predators using the same techniques. These and many other striking examples are thought to represent the convergent evolution of behaviors. Does this convergence reflect changes in the same genes or does evolution act through many genetic routes to create the same behaviors? Genetic differences clearly play a role in behavioral variation, but it remains challenging to identify the genes that underlie evolution of behaviors. However, convergence in the genetic basis of developmental or physiological traits has been discovered with many specific examples of genes and mechanisms, so it is possible to use studies of convergence to discover how behaviors evolve. Therefore, we will use a powerful comparative system to discover the genes and molecular mechanisms that underlie convergent evolution of behaviors for the first time across divergent animals.
The Caenorhabditis nematodes offer a unique experimental platform to connect behavioral differences to genetic differences. Starting with the keystone model organism, C. elegans, and existing data, we will characterize and classify genetic differences across wild isolates from three species of Caenorhabditis - C. briggsae, C. elegans, and C. tropicalis. Whole-genome genotype data combined with high-throughput, high-content imaging of behaviors from these same wild isolates will be input into unsupervised machine learning algorithms to create a high-resolution genotype-phenotype map for a range of natural behaviors and examples of convergence. This map will be queried for signatures of shared genetic changes at orthologous genes to identify which variants are most important evolutionarily. The result will provide the first systematic glimpse into the genomic “knobs” that control behaviors at single-variant resolution across species and insights into the repeatability of the evolution of behaviors.

2019 -
Long-Term Fellowships - LTF

Investigating the role of muscles in morphological plasticity of sea anemone Nematostella vectensis

ANZO Marie (JAPAN)

Developmental Biology Unit - EMBL, Heidelberg - Heidelberg - GERMANY

IKMI Aissam (Host supervisor)

Morphological plasticity is a key adaptive process allowing organisms to cope with changes in their environment. In animals, this process has been described in ephemeral organisms (e.g. flies, worms and mice) that typically experience a specific period in which development will respond to environmental signals producing long-lasting changes in animal body. In contrast, animals with extreme longevity are constantly patterning such that they must continuously adjust their developmental and physiological behaviours with the unpredictable fluctuations of food supply. To decipher the logic of such enhanced morphological plasticity, I am using the sea anemone Nematostella vectensis, a cnidarian laboratory model that exhibits a striking ability to adjust developmental patterns to the nutritional status of the environment while having a long lifespan. Based on preliminary data from the host lab, muscle is emerging as a central tissue that mediates the nutrient-dependent development of Nematostella tentacles. I will define this novel property of muscle cells in response to feeding using transgenic reporter lines. In parallel, I will also establish novel genetic tools to manipulate different muscle types and determine their specific contribution to post-embryonic development. I will also perform single-cell sequencing experiments to characterize the potential metabolic and developmental properties of muscles. Altogether, this proposed work will provide new insight into how morphological plasticity is encoded at the cellular and molecular levels in a long-lived animal.

2019 -
Grant Awardees - Program Grants

Studying sea urchin dermal photoreception to unravel principles of decentralized spherical vision

ARNONE Maria Ina (ITALY)

Dept. of Biology and Evolution of Marine Organisms - Stazione Zoologica Anton Dohrn - Napoli - ITALY

NILSSON Dan-Eric (SWEDEN)

Lund Vision Group, Dept. of Biology - Lund University - Lund - SWEDEN

LUETER Carsten (GERMANY)

Dept. of Evolutionary Morphology (FB1) - Museum fuer Naturkunde - Berlin - GERMANY

LA CAMERA Giancarlo (ITALY)

Dept. of Neurobiology and Behavior - Stony Brook University - Stony Brook - USA

Sea urchins are marine animals genetically close to the vertebrate lineage. Being eye-less and lacking a central nervous system (NS), these animals instead feature dermal photoreceptors dispersed over their spherical body surface and feeding into a decentralized NS. However, sea urchins can visually resolve objects and move towards them, and they can detect looming visual stimuli from any direction and accurately point their spines towards them. Such performance is normally associated with proper eyes feeding information into a brain. Sea urchins thus offer access to a unique visual system of a type that to date has not been studied in terms of its information processing. This alternative solution to vision may also have potential biomimetic applications for robotic miniaturization, smart probes, and intelligent materials where dispersed light detectors control the properties of the material.
The core of the proposed project is to investigate and model the neural mechanisms of information processing, which enables sea urchins to perform spherical vision by deploying an obviously very different mechanism from today's technology, and also very different from visually guided behavior in most other animals. Our study includes molecular and morphological identification of cell types, measurements of behavioral responses and electrophysiological photoreceptor responses, mapping the connectome of sea urchin photoreceptors and NS, and theoretical modelling of the information processing underlying visually guided behavior. We will map the connectomics of the NS and record the activity from key positions in the processing of visual information and generation of locomotory responses. The data will be used for computational modelling of the entire process from visual input to motor control. Special focus will be given to behavioral decisions where small changes in stimuli cause behavioral switches. We will also use genetic approaches to test the agreement between theoretical models and actual behavior.

2019 -
Grant Awardees - Program Grants

nFlare: an innovative light approach to study and modulate neuronal activity in vitro and in vivo

BALLERINI Laura (ITALY)

Dept. of Neurobiology- neuron physiology and technology lab - International School for Advanced Studies SISSA-ISAS - Trieste - ITALY

TIAN Bozhi (CHINA, PEOPLE'S REPUBLIC OF)

Dept. of Chemistry - The University of Chicago - Chicago - USA

FRUK Ljiljana (CROATIA)

Dept. of Chemical Engineering and Biotechnology - University of Cambridge - Cambridge - UK

To understand how the brain represents the world is a central theme in neuroscience. Neural circuits encode information in terms of rate, timing and synchrony of action potentials arising from the activity of a complex spatial organization of excitatory/inhibitory neurons. The identity of the active neurons at any given time has a profound effect onto the final outcome and regulates fundamental human psychophysical behavior. Attempts to decode the complex behaviors in neurons has led to development of several neuromodulation and sensing tools, by which neural activity can be controlled by microelectrodes or through genetically altering specific neural circuits, ultimately to deliver electrical impulses. Unfortunately, these techniques introduce strong perturbations to the observed system, without reaching the desired spatio-temporal resolution. Experimental approaches that allow non-invasive activation of specific phenotypic groups of neurons could introduce systematic variation of the timing of signals revealing how neural ensembles encode information. Here we propose a novel (nano)tool to alter membrane potential in specific neuronal phenotypes in a spatiotemporally controlled manner.
The nFlare project will develop a novel research paradigm in neuroscience based on a new class of injectable nanodevices delivered to neurons and anchored to their membranes. These nanotools have the ability to depolarize cell membranes or to deliver active biomolecules. Electric or chemical stimulation of single cells will be achieved using deep-penetration near infrared excitation light, to reduce invasiveness.
nFlare core aims are: i) the development of nanodevice components able to generate photoelectrochemical current upon illumination with light; ii) state-of-the-art biochemical functionalization of the nanodevice to target specific neurons and to reduce inflammatory response; iii) applicability of such nanoscale devices as high spatio-temporal neuronal activity modulators in 2D and 3D in vitro neuronal networks followed by in vivo validation.
Selective modulation of neural circuits by artificial stimulation of neuronal membrane or controlled delivery of environmental factors could help answering fundamental neurobiological questions.

2019 -
Career Development Awards

Probing the molecular mechanism of SNARE-complex disassembly by NSF

BAO Huan (CHINA, PEOPLE'S REPUBLIC OF)

School of Life Science - University of Science and Technology of China - Hefei - CHINA, PEOPLE'S REPUBLIC OF

Soluble N-ethylmaleimide-sensitive factor (NSF) attachment protein receptor (SNARE) proteins are the essential molecular machinery to catalyze membrane fusion in all the trafficking steps of the secretory pathway. Cognate vesicle SNAREs (v-SNAREs) and target SNAREs (t-SNAREs) progressively zipper together from their N-terminal domains (NTDs) toward their C-terminal domains (CTDs) into a four-helical bundle, which pulls the bilayers together and provides the energy for fusion. Since this helical bundle is inactive for subsequent rounds of fusion, NSF and a-SNAP (alpha-soluble NSF attachment protein) are recruited to disassemble it into individual, reactivated SNAREs, using the energy from ATP hydrolysis. Despite its crucial role in maintaining the fusion competence of the secretory pathway, surprisingly little is known regarding when, where and how NSF disassembles the SNARE complex. In Aim 1, I will examine when the SNARE complex is disassembled by NSF, and address how trans-SNARE complexes, formed in the early stage of membrane fusion, are protected from disassembly by NSF to avoid futile cycling. In Aim 2, I will engineer a new optical sensor to monitor the conformational state of the SNARE complex, thus allowing to visualize where the action of NSF occurs inside living cells. Finally, I will examine how ATP hydrolysis by NSF is converted to disassembly of the SNARE complex using novel reconstitution and single-molecule approaches that afford µsec time solution (Aim 3). Together, the proposed research will reveal unprecedented insights into the molecular mechanism of SNARE-complex disassembly by NSF, and shed new light on fundamental questions in membrane fusion.

2019 -
Long-Term Fellowships - LTF

Cell-cell contacts in tissue patterning and evolution

BARONE Vanessa (ITALY)

Scripps Institution of Oceanography - UC San Diego - La Jolla - USA

LYONS Deirdre (Host supervisor)

How cell differentiation and morphogenesis are coordinated during embryonic development is an open question in developmental biology. The accepted theory sees morphogenesis as a consequence of the cell differentiation events initiated by biochemical signals. However, recent evidence shows that embryonic cells can adjust their differentiation program according to the size and duration of their cell-cell contacts, suggesting that the interplay between biochemical signals and mechanical stimuli determines cell fate. Yet, how these stimuli are integrated in the gene regulatory networks of cell differentiation is largely unknown. Here I will pioneer an evolutionary approach to find conserved mechanisms by which cell-cell contacts regulate gene expression, by comparing two echinoderm species, sea urchin and sea star. In both species, mesendoderm differentiation depends on Wnt/ß-catenin signalling. However, they present different patterns of Wnt/ß-catenin activity and different cell-cell adhesion properties, with sea star cells being far less adhesive than sea urchin cells. Using a combination of live imaging, scRNAseq and theoretical modelling, I will determine how cell-cell contacts regulate mesendoderm cell differentiation in the two species and if variation in cell-cell contact formation underlies the divergence of tissue patterns between sea urchin and star embryos. This study will uncover basic principles of cell differentiation and tissue patterning, broadening our understanding of embryonic development and evolution.

2019 -
Career Development Awards

T-lymphocyte exhaustion in the 3D tumor microenvironment

BAVA Felice Alessio (ITALY)

Department of Genotoxic Stress and Cancer - Curie Institute - Paris-Orsay - FRANCE

Tumors are complex tissues in which cancer cells establish a plethora of interactions with the surrounding microenvironment. In certain tumors, some of these interactions lead to the progressive loss of function of T-lymphocyte sub-populations, which become “exhausted” (Tex). Therapies aimed at restoring Tex activity have proved promising, but the molecular mechanisms underlying exhaustion and their relation to the Tumor MicroEnvironment (TME) organization are poorly understood. To date, much progress has been made using single-cell technologies to define populations based on shared gene-expression patterns. But assigning the spatial distribution of distinct cells in intact tissues is still a hurdle. This is crucial to determine the exact niches in which Tex reside and understand the underpinnings of exhaustion. Here we will employ STARmap [1] – a novel 3D intact-tissue in situ transcriptomics method that I helped develop – to determine the spatial distribution of Tex and the transcriptional programs associated with Tex progression towards dysfunctional states (Aim 1). Based on our preliminary results, we will also explore the involvement of the DNA-damage response (DDR) pathway in T-lymphocyte exhaustion (Aim 2). We will then determine the spatial organization of the TME, in which Tex subpopulations reside, and define 3D Tex-niches (Aim 3). Overall, we will link 3D imaging-based molecular information with functional significance, to uncover novel mechanisms involved in T-lymphocyte exhaustion and in tumorigenesis.

2019 -
Long-Term Fellowships - LTF

Exploring the role of the tumor vasculature in brain metastasis

BEJARANO Leire (SPAIN)

Department of Oncology - University of Lausanne - Lausanne - SWITZERLAND

JOYCE Johanna (Host supervisor)

Brain metastasis (BrM) represents the most common brain malignancy, predominantly arising from non-small cell lung cancer, breast cancer and melanoma. We hypothesize that the tumor microenvironment (TME) plays an essential role in the establishment and progression of BrM. The brain is constituted by a unique TME, which includes tissue-resident cell types such as astrocytes and microglia, and critically, the blood-brain barrier (BBB). Cancer cells are able to exploit the brain vasculature for their own benefit, by forming the blood-tumor barrier. This specialized vasculature is an essential TME compartment, as it may suppress anti-tumor responses by blocking immune cell infiltration, specifically of cytotoxic T cells. Indeed, a new perspective to target the tumor vasculature in combination with immunotherapy has recently emerged from studying other cancers; however, this concept has not been explored in BrM to date. Therefore, in the proposed project, I will investigate the biology of two major cellular components of the BBB: endothelial cells (ECs) and pericytes in BrM arising from distinct primary tumors. For this purpose, I will perform transcriptomic analysis of these cell types in patient BrM samples originating from lung, breast or melanoma primary tumors. I will combine this analysis with different experimental approaches available in the host lab, including immunocompetent mouse BrM models and in vitro and ex vivo cell cultures that have been optimized to model the brain TME. This integrated and complementary experimental strategy will facilitate the identification of innovative therapies for BrM to be used alone or in combination with immunotherapies.

2019 -
Long-Term Fellowships - LTF

Defining how activation of skin and lymph node sensory neurons controls immunity

BEN-SHAANAN Tamar (ISRAEL)

Microbiology and Immunology - University of California - San Francisco - USA

CYSTER Jason G. (Host supervisor)

Pain sensation, though unpleasant, is considered protective as it provokes aversive behavior from noxious stimuli. Studies show that sensory neurons, which generate the perception of pain and touch, interact with another important protection mechanism, the immune system. Ablation of sensory neurons can alter the skin-resident immune cell response. However, many aspects of this interaction remain uncharacterized.
Cells and antigens from barrier sites, like the skin, are drained and sampled by the lymph nodes (LN). Interestingly, LN are innervated by fibers expressing the sensory neuron derived-peptides Substance P and Calcitonin gene-related peptide. Some of these fibers are found in close proximity to leukocytes. However, their direct impact on immune cell activity is unknown.
This project aims to characterize the effects of sensory neurons in the skin and LN on immune cell behavior. To this aim, we propose to combine chemogenetics to locally activate sensory neurons, with a broad analysis of immune cell responses. Mice expressing Cre under the control of sensory neuronal markers, including the broadly expressed gene Advillin and genes selective for subsets of sensory neurons, will be injected intradermally or in the LN with a viral vector encoding the Cre-dependent expression of the hM3D(Gq) activating receptor. Once validated, this technique will be used to locally activate sensory neurons and characterize subsequent immunological changes using flow cytometry, histological and live imaging techniques. This project can provide new insights into how sensory innervations, which regulate behavior by creating the sensation of touch and pain, shape local immune responses.

2019 -
Long-Term Fellowships - LTF

Synaptic basis of temporal learning

BENDER Franziska (GERMANY)

Department of Neuroscience - Institut Pasteur - Paris - FRANCE

DIGREGORIO David (Host supervisor)

A key brain region involved in accurate temporal refinement of motor and also cognitive behaviours is the cerebellum. Cerebellar granule cells (GCs) receive multisensory information from outside the cerebellum via mossy fibres (MF). The prevailing hypothesis is that, in order to learn temporal sequences, the cerebellum requires a rich diversity of GC activity patterns that as a population represent a distributed temporal representation of sensory inputs. But this has never been directly demonstrated. Unpublished network models from the hosting laboratory strongly suggest that their previous findings (input specific diversity of MF-GC synaptic dynamics) are sufficient to generate a diverse GC firing patterns that act as a temporal basis for cerebellar learning. We will use novel high speed two-photon in vivo imaging of GC firing patterns in behaving animals, and examine whether the diversity of GC firing patterns requires functional diversity of MF-GC synapses, and whether this mechanism is specific for different sensory modalities, using optogenetic activation of specific input types. Finally, we will optogenetically inhibit specific MF types to show their influence over specific timescales of eyeblink conditioning, then image the temporal representation in the GC layer before, during and after a temporal learning task. Expected results will included identification of the underlying cellular and circuit mechanism of temporal learning required for fine tuning motor and cognitive processes.

2019 -
Long-Term Fellowships - LTF

Epigenetic normalization through engineering of S-adenosyl methionine metabolism

BOON Ruben (BELGIUM)

Cancer Center - MGH Boston - Boston - USA

MOSTOSLAVSKY Raul (Host supervisor)

Pancreatic ductal adenocarcinoma (PDAC) remains an incurable disease with a high death rate. PDACs are exceptionally difficult to treat because of high genetic plasticity and encapsulation by cancer-associated fibroblasts (CAFs). Either are due to an altered epigenetic landscape where PDACs and CAFs show a switch towards global chromatin hypo-methylation, and concomitant hyper-methylation of tumor-suppressor genes. As CAFs do not show any mutagenesis, the molecular cause must be found in the tumor microenvironment.

The nutrient microenvironment represents a novel but important aspects of tumor biology. Nutrients are converted into the basic building blocks that fuel cellular proliferation, but also regulate epigenetic modifications. S-Adenosyl-Methionine (SAM) represents a nodal point in one carbon metabolism and is required for methylation of histones and DNA. We therefore propose that metabolism cannot sustain the high SAM requirement of both PDAC and CAF, thus shunting away carbons from epigenetic maintenance. This project therefore aims to map the link between nutrient availability, one carbon metabolism, availability of SAM, and the methylation profile of chromatin. Furthermore, we will establish a genetic sensor and CRISPR-based screening system to visualize the regulation of SAM compartmentalization, and attempt to normalize the epigenome of PDACs and CAFs through engineering of SAM metabolism (EpiSAMe). These studies will shed new insight into the crosstalk between metabolism and epigenetics in PDAC and provide novel molecular for the treatment of this devastating disease.

2019 -
Long-Term Fellowships - LTF

Generating a global view of cell lineage trees during early mouse embryo development

BOWLING Sarah (UK)

Stem Cell Program - Boston Children's Hospital - Boston - USA

CAMARGO Fernando (Host supervisor)

Understanding the routes through which a single cell populates the adult organism is one of the most fundamental yet elusive areas of biology. This project aims to use recent innovations in single-cell technology to generate a blueprint of cell fate decisions during early mouse development, and to examine the mechanisms behind the regulative nature of embryogenesis. Firstly, I will employ CRISPR-Cas9 technology to induce thousands of unique barcodes in cells, enabling the interrogation of cellular clonal history. This analysis will shed light on outstanding but fundamental areas in the field of developmental biology, such as pinpointing the earliest clonal groupings and differentiation decisions, and quantifying the number of progenitor cells present at each stage of early mouse development. Secondly I will combine the lineage tracing analysis with single-cell RNA sequencing (scRNA-seq) which will provide the molecular resolution to precisely classify cell types and identify mechanistic players in fate decisions. Finally, I will perform these analyses in a chimeric embryo which contains cells unable to contribute to certain lineages. In this final aim I will analyse how the differentiation of wild-type cells is skewed by the presence of the mutant cells. Together these analyses will generate a detailed chart of the clonal history and expression profiles of every cell during gastrulation and thereby provide an important reference point toward understanding a wealth of cellular processes. Furthermore, the chimera experiments will enable me to shed light on the largely unexplored area of compensatory growth during embryogenesis.

2019 -
Long-Term Fellowships - LTF

Dissecting the role of metabolism in cancer genomic instability

BURGOS BARRAGAN Guillermo (SPAIN)

Meyer Cancer Center - Weill Cornell Medical College - New York - USA

CANTLEY Lewis (Host supervisor)

Genomic instability is a common feature of cancers and can have an important impact on cancer progression and response to treatment. However, the mechanisms underlying cancer genomic instability are not completely understood. While DNA repair defects are thought to be a major cause of this trait, other factors seem to be involved. In the last decade, metabolism has received a lot of attention within the cancer research community. Most of the work has focused on understanding the metabolic requirements of cancer cells for survival and proliferation. However, metabolism does not only provide building blocks for biosynthesis, but it also generates reactive by-products that can damage cellular components such as DNA. Thus, the metabolic alterations characteristic of cancer cells might not only support cancer development by supporting biosynthesis but also by promoting genomic instability.

The aim of this research proposal is to identify changes in the metabolism of endogenous genotoxins in cancer and to elucidate the role of these alterations, particularly in the context of genomic instability. In addition, the potential exploitation of this aspect of metabolism for cancer therapy will be explored. To achieve these goals a diverse range of techniques and study systems will be used, including transcriptomics, metabolomics, CRISPR-based genetic screens, cancer cell lines and mouse models.

2019 -
Long-Term Fellowships - LTF

Prochlorococcus cyanophage: lysogenic potential and development of a genetic system

CAPOVILLA Giovanna (ITALY)

Departments of Civil and Environmental Engineering and Biology - MIT - Cambridge - USA

CHISHOLM Sallie W. (Host supervisor)

Prochlorococcus, the smallest photosynthetic organism, has diversified into many ‘ecotypes’ with finely tuned distinct ecological niches. The small and variable genomes of these cyanobacteria encode genes unique to particular habitats. A driving force for gene transfer and evolution in cyanobacteria is phage-mediated. While numerous reports describe lytic cyanophages that infect Prochlorococcus, surprisingly there is only indirect evidence that lysogenic phages – those that spend a portion of their life cycle incorporated into the host genome – exist. The central goals of this proposal are i) to analyze the occurrence of lysogens in wild populations of Prochlorococcus, and ii) to establish a robust protocol for gene editing allowing precise manipulation of the Prochlorococcus genome. The second goal will exploit, but is not dependent upon, advances from the first.
This will be the first study to assess lysogeny across natural Prochlorococcus populations and to develop phage-mediated genetic systems for engineering cyanobacteria. The development of efficient tools for genome editing would be extremely beneficial for understanding Prochlorococcus metabolism. It would allow functional studies for thousands of unannotated genes that influence the relative fitness of different ecotypes in different environments. A unique aspect of my proposal is the employment of a novel synthetic biology strategy to build engineered phages with broad host specificity, and to use them as vehicles for genome editing systems such as CRISPR/Cas9. This technique will be useful for several applications and will further establish Prochlorococcus as a model organism in environmental microbiology.

2019 -
Long-Term Fellowships - LTF

Structural and functional characterization of P-Rex 1/2 in cell signaling and cancer

CHAKER-MARGOT Malik (CANADA)

Biozentrum - University of Basel - Basel - SWITZERLAND

MAIER Timm (Host supervisor)

Phosphatidyl-inositol 3,4,5 triphosphoshate-dependent Rac exchanger (P-Rex) are a group of proteins involved in the activation of the small GTPase Rac, by acting as a guanidine exchange factor (GEF). Both Rac and P-Rex proteins are known to be overexpressed and altered in several cancers, consistent with their role in both cell mobility and their numerous interactions with both the mTOR axis and the PI3K/PTEN pathway. P-Rex1/2 specifically, are known to interact with mTOR and PTEN, among others, placing them at the nexus of regulatory pathways for cell metabolism, proliferation and mobility.
Despite their central role in cell signaling, very little is known about the interplay between such interactions. No structural information is available for the full-length P-Rex proteins and how different interaction partners affect its structure and function. I will therefore set out to better characterize the emerging properties of the multidomain P-Rex proteins by purifying both P-Rex1/2 and determining their structure using cryo-EM and orthogonal biophysical characterization methods. I then aim to define the interaction between P-Rex proteins and its regulatory partners, PTEN and the mTOR complexes 1 and 2. I will define their domains of interactions and determine the structure of the complexes, using a combination of cryo-EM and cross-linking and mass-spectrometry.
I expect to provide new insights into the functions of P-Rex proteins and their crosstalk with core cellular signaling pathways. Structures of full-lengths P-Rex alone or in complex with its interaction partners will serve as a platform for drug discovery, thereby providing new cancer therapeutic avenues.

2019 -
Cross Disciplinary Fellowships - CDF

How do populations of identical cells break symmetry? A novel data-driven approach using gastruloids

CHEESEMAN Bevan (NEW ZEALAND)

Department of Genetics - University of Cambridge - Cambridge - UK

MARTINEZ-ARIAS Alfonso (Host supervisor)
TRIVEDI Vikas (Host supervisor)

A fundamental question in biology is how robust developmental processes can result from the dynamic and variable interaction, division, and movement of individual cells. In vertebrates, a critical process that provides a blueprint for development is gastrulation. During gastrulation, a population of initially identical cells breaks symmetry, robustly establishing the anteroposterior (AP) axis. In this project, I will investigate how this critical symmetry breaking event arises through space and time from initially identical cells using the gastruloid experimental system and a novel data-driven computational approach. Gastruloids are mouse embryonic stem cell aggregates and have been shown to robustly recapitulate AP symmetry breaking in the absence of external spatial cues. Also, gastruloids provide a tractable high-throughput experimental system amenable to imaging using light-sheet fluorescence microscopy (LSFM). My computational approach will overcome current methodological bottlenecks and allow: tracking of all cells through time in gastruloids using LSFM in high-throughput, uncovering of key cell dynamics during symmetry breaking and their variations at the individual cell level, and identification of gene expression patterns.

This project represents essential first steps towards a mechanistic understanding of how robust symmetry breaking arises out of the individual, variable actions of an initially equal population of cells. In addition to this, the approach and tools are both general and extendable and hold the potential to enable a similar approach across a range of developmental processes and experimental systems.

2019 -
Grant Awardees - Young Investigator Grants

Conversations between brain and vasculature: studying and mimicking their intertwined development

CHEUNG Christine (SINGAPORE)

Lee Kong Chian School of Medicine - Nanyang Technological University - Singapore - SINGAPORE

LOH Kyle (USA)

Dept. of Developmental Biology - Institute for Stem Cell Biology & Regenerative Medicine - Stanford - USA

The brain comprises billions of neural cells interwoven in a highly precise way and it is ultimately responsible for how we perceive, react to, and remember the world around us. We are fascinated by how the intricate structure of the brain is assembled step-by-step at an early stage of life (“embryonic development”), starting as a relatively shapeless mass within the nascent embryo and culminating in a highly complex organ in an adult. Specifically, we hypothesize that blood vessels that permeate the early brain have important roles in fostering brain assembly, and we further predict that they produce important cues that actively shape a specific early step of brain development. This is of great significance, because blood vessels were once thought only to passively supply oxygen and nutrients to various organs. We instead suggest an active role in which blood vessels function as a ‘signaling center’ to actively control the arrangement of neural cells during brain assembly.
We have assembled an intercontinental team to tackle this question via a three-part, integrated effort. First, we will create a map of where and when arteries and veins (two major types of blood vessels) first enter the nascent brain. Second, we will intermingle neural and blood vessel cells in a Petri dish and study their interactions. Third, we will block specific functions exerted by blood vessel cells and propose that this will have a commensurate impact on the spatial arrangement of nearby neural cells. These activities are innately highly collaborative and will involve interactions between the partner laboratories in each nation.
Taken together, we propose to explore how the brain is assembled during embryonic development and we will test whether blood vessels act not as passive support for, but rather as active executors of, this intricate process at early steps. If true, this has important ramifications. Not only will it shed light on how the early brain is assembled, but it adds to the emerging idea that neural cells and blood vessels share an intimate functional and spatial relationship, starting from the embryo and lasting into the adult — an idea pertinent to early brain assembly; our efforts to engineer this process in a Petri dish for regenerative medicine; and finally, the origin of certain neurological disorders that are caused in part by blood vessel dysfunction.

2019 -
Grant Awardees - Program Grants

An integrative approach to decipher flowering time dynamics under drought stress

CONTI Lucio (ITALY)

Dept. of Biosciences - Università degli studi di Milano - Milano - ITALY

JUENGER Thomas (USA)

Dept. of Integrative Biology - University of Texas at Austin - Austin - USA

IZAWA Takeshi (JAPAN)

Dept. of Agricultural and Environmental Biology/Lab of Plant Breeding and Genetics - The University of Tokyo - Tokyo - JAPAN

Plants live in an ever-changing environment which is not always compatible with their survival. A major life threatening condition is drought stress. While most plants can deploy an array of physiological countermeasures to endure remarkable levels of stress, there is a huge variability in the different strategies that plants choose to adopt to deal with drought stress. Many plants evade detrimental stress conditions by activating their reproductive development (flowering) earlier compared to non-stress conditions, a strategy known as drought escape (DE). Notably, even in the most water-rich environments plants face unpredictable dry periods, yet how this information affects the floral network at the molecular levels is unknown.
We will address this question with a blend of molecular and genetics-based approaches. We will leverage known mutants with altered DE to identify candidate mechanisms that are drought sensitive and can act as molecular switch to activate flowering. To comprehensively define DE mechanisms utilized under natural conditions we will assess the variability of the DE response in natural plant populations and in crops, which were selected by human intervention for the different field scenarios. Our targeted large-scale screens will allow us to identify naturally occurring variants in the DE process and decipher the molecular mechanism responsible for DE activation. This information will provide breakthroughs in our understanding of novel regulatory mechanisms that play a role in driving developmental adaptations across extremely variable environmental conditions, their natural genetic variation and the selective forces that maintain such variation in populations, an important aspect for predicting and dealing with the effects of climate change. Finally, because flowering time is a major component of yield potential in crops, the defined mechanisms will help us develop breeding strategies targeted for sub-optimal irrigation scenarios to produce crops with ameliorated performances under drought conditions.