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2019 -
Long-Term Fellowships - LTF

An integrated organoid-engineering approach to study human brain development


Eli and Edythe Broad CIRM Center for Regenerative Medicine and Stem Cell Research - University of Southern California - Los Angeles - USA

MORSUT Leonardo (Host supervisor)
QUADRATO Giorgia (Host supervisor)

The generation of human cerebral organoids promises to profoundly change our understanding of human brain development by enabling its detailed study. However, current protocols have several limitations and give rise to organoids characterized by low reproducibility, small size, lack of vascularization, limited neuronal maturation, and reduced cellular diversity. These limit the use of organoids as a model system to study human brain development. Here, I propose to integrate existing 3D culturing protocols with synthetic biology and tissue engineering approaches in order to push the field beyond these limitations and study basic mechanisms of human brain development. In so doing, I expect to both generate cerebral organoids that better resemble the human brain and study basic mechanisms in an “understanding by building” framework. Specifically, I plan to (i) increase neuronal maturity by integrating the brain organoid with a smart functional vasculature system (ii) increase organoid size and cell diversity by modulating the gene expression dynamics of Hes1 (iii) increase organoid reproducibility by building an artificial notochord producing a sonic hedgehog (SHH) gradient in a time regulated fashion in order to pattern brain structures. By integrating live cell imaging, massive single-cell profiling, and electrophysiology, I will be able to study the overall effects of the three specific goals on organoid development.

2019 -
Career Development Awards

Coordination of mitochondria biogenesis and cell growth


Institute of Functional Epigenetics - Helmholtz Zentrum - Munich - GERMANY

Mitochondria are a fundamental component of eukaryotic cells that provide ATP through oxidative phosphorylation and play important roles in the regulation of metabolism and cell death. Accurate homeostasis of mitochondria is important for cell function, and misregulation of mitochondria in humans causes a variety of severe diseases. To achieve homeostasis, cells have to ensure that the amount of mitochondria doubles with each cell doubling period. Because mitochondria maintain their own genome encoding for essential mitochondrial proteins and RNAs, this requires a quantitative coordination of mitochondrial DNA replication, production of mitochondrial proteins, and cell growth. However, how cells couple mitochondrial biogenesis with cell growth remains unknown.
Here, I propose to use budding yeast as a model organism to identify the molecular mechanism underlying the coordination of mitochondria biogenesis with cell growth and cell cycle. I will use an interdisciplinary approach, combining the microscopy based protein concentration and cell size measurements established during my postdoc, a LacO-LacI based live-cell imaging approach, and qPCRs to unravel how concentrations of mitochondrial components as well as their production rates are linked to cell cycle and cell growth. I will then use mathematical modelling to establish a quantitative model for mitochondrial homeostasis that I will test by measuring the dynamic response of mitochondria maintenance to transient perturbations. I expect that our work will then catalyze our knowledge of mitochondria maintenance in human cells and will thereby lead the way to a better understanding of mitochondrial diseases.

2019 -
Grant Awardees - Program Grants

Elucidating the development of biological optical nanostructures


Dept. of Evolution and Optics of Nanostructures - Ghent University - Ghent - BELGIUM


School of Integrated Technology/Nano Convergence Systems Group - Yonsei University - Incheon - KOREA, REPUBLIC OF (SOUTH KOREA)


Center for Interdisciplinary Research in Biology - College de France - Paris - FRANCE

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.

2019 -
Career Development Awards

Revealing the functional roles of cell-specific DNA methylation following implantation


Department of Molecular Cell Biology - Weizmann Institute of Science - Rehovot - ISRAEL

DNA methylation is essential for normal mammalian development. While seminal work has provided tremendous insight into the dynamic regulation of DNA methylation throughout embryogenesis, comprehensive understanding of how cell-specific methylation programs are established and maintained, and how are they involved in defining cell states in vivo through regulation of target genes, remains a formidable task. Revolutionary technologies now offer unprecedented opportunities for understanding the function of DNA methylation in specifying, memorizing and modulating embryonic programs. These powerful tools motivate further development of novel experimental systems, integrating single cell monitoring with flexible engineering of markers, reporters and perturbations. This will enable to precisely target key rare embryonic cell populations for in-depth analysis.
Here, combining cutting-edge methods for single cell mapping of DNA methylation and gene expression, and by developing a novel approach for inferring spatial information from single cell genomic data, we propose to comprehensively chart the post-implantation embryo, at unprecedented resolution. To move to functional studies, we will implement our recently established reporter system that enables monitoring and isolation of cells based on endogenous locus-specific changes in DNA methylation. Specifically, we will study the developmental potential of rare epiblast cells that we identified to exhibit lower-than-expected genome-wide methylation levels. Our combined approach will open new avenues for elucidating the contribution of cell-specific DNA methylation changes to cell-state and function following implantation.

2019 -
Grant Awardees - Program Grants

Enhancing mitochondrial DNA fidelity to improve mammalian lifespan and healthspan


Research Group Stewart - Max Planck Institute for Biology of Ageing - Cologne - GERMANY


Institut Européen de Chimie et Biologie - Univ. Bordeaux, U1212, CNRS UMR5320 - Pessac - FRANCE


Dept. of Ecology, Evolution and Organismal Biology - Iowa State University - Ames - USA

Animal mitochondrial DNA (mtDNA) has a higher substitution rate than nuclear DNA, with the accumulation of mtDNA mutations being one of the hallmarks of ageing. This discrepancy in the rates of evolution is partially due to the lack of mismatch repair activities in the mitochondria. Octocorals – a group of cnidarians – have a reduced rate of mitochondrial evolution and encode a MUTS-like protein (mt-MutS) in their mtDNA. Previous analyses suggested that this enzyme was acquired from a virus and has been universally retained among octocoral taxa. Its function, however, remains unknown. The project will combine comparative, structural, and experimental approaches to investigate the function of mt-MutS and to test whether mt-MutS expression results in lower mutation rates in mtDNA and improve heath in ageing. Comparative analysis of octocoral mtDNA will be used to identify distinct mutation patterns among its lineages and correlate them with the changes in mt-MutS. Partial mt-MutS sequences will be used to identify clades with unusual or accelerated rates of mtDNA evolution for additional sampling. Site- and taxa-specific evolutionary rates in mt-MutS will be analyzed to infer functional and structural constraints and to optimize the choice of the mt-MutS for transgenesis. Ancestral sequences of mt-MutS for the nodes of interest will also be reconstructed and analyzed. A structure-function approach will be utilized for in vitro dissection of mt-MutS functions. The full-length proteins from several species and a reconstructed ancestral sequence will be tested for stability and for amenable structure determination. Isolated domains will also be used for high-resolution structural analysis by diverse biophysical techniques to probe molecular details of binding and nuclease activity for understanding and improving function. Finally, we will generate transgenic mice that express a mitochondrially-targeted version of this optimized mt-MutS enzyme to test its effects on mtDNA mutation rate. The transgene construct will be knocked-in to mice by directed Easi-CRISPR template repair or BAC-transgenesis. mtDNA mutation rate analyses in wildtype and mice with enhanced mitochondrial mutation rates will be undertaken. An ageing study on mice expressing mt-MutS will determine if enhanced mtDNA fidelity can positively affect organismal lifespan and healthspan.

2019 -
Grant Awardees - Program Grants

Communication and the coordination of collective behavior across spatial scales in animal societies


Dept. of Biology - University of Konstanz - Konstanz - GERMANY


Dept. of Evolutionary Biology and Environmental Studies - University of Zurich - Zurich - SWITZERLAND


College of Science and Engineering - James Cook University - Townsville - AUSTRALIA


Dept. of Ecology, Evolutionary Biology and Behavior - Michigan State University - East Lansing - USA

ROCH Marie (USA)

Dept. of Computer Science - San Diego State University - San Diego - USA

We propose to use new tracking technology and computational modeling to determine how vocal communication influences collective behavior in animal societies. Canonical examples of collective movement such as bird flocking and fish schooling involve cohesive groups making short-term decisions in a shared context. However, many animals form stable social groups that coordinate and cooperate over extended time spans, across varying distances, and in diverse contexts. In these stable animal societies, group members must make decisions despite varying access to information and exposure to the costs and benefits of coordinating. Moreover their decisions are likely to be shaped by the long-term social relationships among group members. To achieve coordination in such systems, many species use sophisticated signaling systems, such as vocal communication, that transfer information among group-mates. Animals can flexibly control the vocalizations they produce independent of their movements, resulting in a complex interplay between signaling and movement that ultimately drives group-level outcomes such as collective decisions and coordinated actions.
To understand the mechanisms underlying coordination in animal societies, we will record movements and vocal signals concurrently from all members of wild animal groups at a high resolution, and across varying degrees of spatial dispersion. We will compare three mammal species that face a common set of coordination task, but differ in cohesiveness: meerkats form highly cohesive groups, coatis are moderately cohesive, and spotted hyenas live in fission-fusion societies. In each species, we will 1) fit at least one entire social group in the wild with tags that continuously record fine-scale movements and vocalizations, 2) combine supervised and unsupervised machine learning to identify animal calls and movement states, 3) develop modeling approaches to reveal how animals integrate spatial and acoustic information, how information flows through groups, and how social interactions give rise to collective outcomes, and 4) conduct audio playback experiments to isolate causal factors driving collective dynamics. Combining these approaches with long-term data from field studies will shed light on both unifying features underlying coordination mechanisms across animal societies and differences imposed by distinct constraints.

2019 -
Grant Awardees - Program Grants

How complex behavior is encoded in the genome and wired in the brain

STREELMAN Jeffrey Todd (USA)

School of Biological Sciences - Georgia Institute of Technology - Atlanta - USA


Dept. Genes - Circuits - Behavior - Max Planck Institute of Neurobiology - Martinsried - GERMANY

Despite effort, it remains incredibly difficult to identify the cellular basis, and/or the causative genetic variants, underlying complex behavior. Understanding how behavior is encoded requires solving a dual problem involving both neurodevelopment and circuit function. Genes build nervous systems; nervous systems are activated to produce behavior. Streelman and Baier will collaborate to develop a unique model system to chart the complex path from genome to brain to behavior, in vertebrates from natural populations. In Lake Malawi, male cichlid fishes construct sand ‘bowers’ to attract females for mating. Bower building is an innate, repeatable natural behavior that we quantify in the lab. Males build two bower types: 1) pits, which are depressions in the sand, and 2) castles, which resemble miniature volcanoes. Species that build these two bower types can interbreed in the lab. Remarkably, first-generation hybrids of pit- and castle- species perform both behaviors in sequence, constructing first a pit and then a castle bower, indicating that a single brain containing two genomes can produce each behavior in succession. Moreover, brain gene expression in these hybrids is biased towards pit- alleles during pit digging, and castle- alleles during castle building. This phenomenon of allele-specific expression matched to behavior is compelling and offers the chance to identify the genome regulatory logic and neural circuitry underlying complex behavior. Streelman’s group will use single-cell RNA-sequencing to pinpoint specific cell populations that mediate context-dependent allele-specific expression in male bower builders. Baier’s team will use genome editing and optogenetic tools to manipulate the neurons that matter in the brains of behaving Malawi bower builders. Our collaborative work will thus identify the neurons responsible for biased allelic gene expression matched to behavior, and then manipulate those neurons to modify behavioral output. Achieving our goals will demonstrate how the genome is activated in particular cell types to produce context-dependent natural social behaviors.

2019 -
Long-Term Fellowships - LTF

Survival strategy of anaerobes in human microbiome using radical enzyme-assisted peptide metabolites


Department of Pharmacy - National University of Singapore - Singapore - SINGAPORE

MORINAKA Brandon I. (Host supervisor)

With the increasing plethora of microbial genomes, the importance of complex microbial communities in nature has been recognized. The human microbiome has particularly attracted researchers because of its direct link to health and disease. Despite their great diversity, which is apparent from the available genomic information, little is known about the bioactive small molecules secreted by the human microbiome. Few studies have investigated the presence and ecological roles of these metabolites in the human microbiome at physiological conditions, because of the lack of appropriate methodologies.
This project attempts to address uncharacterized peptide metabolites encoded in the genomes of human-associated anaerobic bacteria, and elucidate how they contribute increasing the fitness of the host microbes. These peptides are likely to have a novel scaffold made by a unique class of oxygen-sensitive enzymes. I intend to perform multidisciplinary research composed of two approaches: A) Production of the peptide metabolites in a heterologous microbe; B) A metagenome-based assay to evaluate the population changes in the cultured microbial community upon peptide treatment. The former approach has the potential to generate natural products independent of the strain; the latter enables us to decipher the biological activities of small molecules in a complex microbial community. Innovation in both concepts will increase our understanding and the availability of microbiome-derived compounds that could improve human health. Furthermore, given the broad distribution of target peptides in anaerobes, this research will also elucidate the breadth of anaerobic chemistry inside humans.

2019 -
Cross Disciplinary Fellowships - CDF

Understanding synaptic diversity using quantitative single-molecule localization microscopy


Department of Synaptic Plasticity - Max Planck Institute for Brain Research - Frankfurt am Main - GERMANY

SCHUMAN Erin M. (Host supervisor)

Synaptic diversity is crucial for neuronal function. The heterogeneity of individual synaptic proteomes in a neuron underlies its input integration, compartmentalization of function, and neuronal plasticity during learning. The canonical classification of synapses based on neurotransmitter systems (e.g. the generic excitatory or inhibitory synapses) has become increasingly inadequate to explain the diverse synaptic responses in electrophysiology and plasticity. Without quantifying the variability of molecular combinations in synapses, our basic understanding of synapses is incomplete. Recently single-molecule localization microscopy has created an opportunity to interrogate individual synapses across a whole neuron. DNA Points Accumulation for imaging in Nanoscale Topography (DNA-PAINT) is ideally suited for localizing and quantifying protein copies of interest compared to conventional stochastic optical reconstruction microscopy (STORM) because of its well-defined blinking kinetics and absence of photobleaching issues. This proposal aims to quantify the proteomic heterogeneity of synapses in individual neurons using DNA-PAINT and to interrogate the relationship between plasticity and synaptic diversification using multiple approaches. It will address the following:
1. The differential localization, variability, and stoichiometry of five representative synaptic proteins in the synapses of a neuron
2. The correlation between synapse activity and their proteomes
3. A potential change in synaptic diversity after single-spine plasticity
4. The effects of global homeostatic scaling on synaptic diversity
5. The remodeling of synapse diversity by newly synthesized proteins

2019 -
Long-Term Fellowships - LTF

Circuit mechanisms for visual stability


- Institute of Molecular and Clinical Ophthalmology - Basel - SWITZERLAND

ROSKA Botond (Host supervisor)

Our eye movements constantly generate motion patterns on our retina, yet our perception of objects in space remains stable. For the brain to recognize self-generated visual motions to construct an image that is spatially invariant of one’s own movement, retinotopic visual information has to be integrated with eye position. Primate studies have shown cortical areas containing neurons responding to different combinations of these two reference frames, and perturbations of these areas suggest a role for space perception. However, little is known about the cellular and circuit mechanisms by which reference frame transformations underlying visual stability occur. I propose to fill that void by bringing this field into mouse research where methods to monitor, perturb and dissect neurons and circuits in much more precise and targeted ways are possible. Using preparations in which visual inputs, eye positions and eye movements are experimentally tracked and controlled will allow me to isolate visual, proprioceptive and motor components. Brain-wide functional ultrasound imaging will reveal areas activated by each component alone or in combination. Calcium imaging will then provide large population responses with single cell resolution. Finally, single-cell-initiated rabies tracing and whole-cell recording will be used to delve into the circuit and synaptic mechanisms underlying the functional properties we find among different neuronal populations. This project will accelerate the field of visual stability, and more generally sensorimotor integration, by providing a new niche where cellular and circuit mechanisms can be thoroughly dissected.

2019 -
Long-Term Fellowships - LTF

A new reversible lesion technique for studying the primate brain in naturalistic environments


Department of Neuroscience - University of Pennsylvania - Philadelphia - USA

PLATT Michael (Host supervisor)

Understanding how the brain works is the next great frontier in biology. One can choose among many available tools and approaches to undertake this challenge. One approach that has proven successful is to investigate the necessary role of a brain area as a first step before trying to understand the neural mechanisms by which it accomplishes its role. To identify the necessary role of a brain area, one needs to manipulate its activity with causal research methods. Optogenetics is a new causal research method that brought about a revolution in brain research conducted with rodents and small animals. Unfortunately, the same cannot be said about optogenetics when applied to animals more similar to humans, such as non-human primates. The larger primate brain poses new challenges to the success of this powerful research method. In this research project, I will develop a new optogenetic-based reversible lesion technique that addresses these challenges. Moreover, I will do so while freeing non-human primates from the physical restraints imposed on them for the practical purpose of accessing their brain. This original method will put together three new technologies: inhibitory step-function opsins, convection-enhanced delivery of opsins, and chronically implanted LED illuminators that are wirelessly activated. With this tool, it will be possible to study for the first time the causal role of brain areas of monkeys that are behaving in naturalistic environments. I will apply this tool to the study of social cognition in monkeys interacting with their peers, and borrow well-studied behavioural tasks from ethology and primatology to develop the emerging field of neuro-ethology.

2019 -
Cross Disciplinary Fellowships - CDF

Modelling the sequence - structure - function relationship in proteins with machine learning


Edmond J. Safra Center for Bioinformatics - Tel Aviv University - Tel Aviv - ISRAEL

WOLFSON Haim (Host supervisor)
ZUK Or (Host supervisor)

Understanding and exploiting the sequence - structure - function relationship is one of the fundamental goals of bioinformatics. Systematic predictions of protein structure, function and interactions could provide a global understanding of the protein interaction networks that underlie cell’s life. Conversely, being able to design molecules, peptides or proteins with a prescribed fold or function is a promising lead for drug design. Unfortunately, current state-of-the-art remains far away from this goal. Indeed, direct approaches based on physical interactions, such as Ab Initio Molecular Dynamics (MD) or Fragment-based folding are often either too computationally expensive or inaccurate. Recently, important progress were achieved in structure, function and protein-protein interactions prediction by the introduction of coevolution-based approaches such as Direct Coupling Analysis (DCA). Though powerful, such models are strongly limited in practice because they require many sequences from the same family in order to accurately predict structure. One natural explanation is that they lack general knowledge of proteins biochemistry, such as amino-acid similarity and stereotypes of interactions. Here, we propose to leverage recent advances in the fields of transfer learning and deep generative models to design new coevolution models that share knowledge between protein families, and thus would be more efficient than naive DCA. Beyond structure prediction, we also propose new general-purpose architectures that aim at learning directly the sequence - structure - function relationship, and discuss applications to protein function prediction and sequence/fold designs.

2019 -
Long-Term Fellowships - LTF

Role of functional connectivity dynamic changes to cognitive control-mood interplay


Department of Psychology - Stanford University - Stanford - USA

POLDRACK Russell (Host supervisor)

This project aims at clarifying how brain connectivity fluctuations over time relate to mood changes and its interplay to cognitive control. Three specific aims are defined. 1) I will clarify how aberrant mood responses contribute to cognitive control abilities. 2) I will identify how mood fluctuations over short timescales of days to months impact cognitive control abilities. 3) I will investigate how dynamic changes in brain connectivity relates to mood changes and its interplay with cognitive control.
I will use reinforcement learning algorithms and computational models of mood to describe cognitive control-mood interplay and determine how aberrant mood reactions relate to impaired behavioural adjustment. To this end, I will investigate cognitive control-mood interplay in a large sample of healthy adolescents and adolescents with clinical and subclinical depression as impaired mood reactivity is mostly pronounced in this population. Behavioural data of cognitive control–mood interplay will be used conjointly with functional resting state brain imaging data. Repeated, longitudinal assessment at the behavioural and brain level will elucidate dynamics of network configuration over time and how they impact on mood changes and its interaction with cognitive control. With this project I aim to reach a mechanistic understanding of cognitive control-mood interplay and associated brain networks. The project has also implications for the understanding of symptoms of depression, which is a major, leading cause of disability.

2019 -
Grant Awardees - Program Grants

Regulation of membrane receptor function in the brain by lipid composition and dietary inputs


Dept. of Physics - University of Helsinki - Helsinki - FINLAND


Dept. of Integrative Biology and Pharmacology - University of Texas Health Science Center at Houston - Houston - USA


Dept. of Chemistry - University of Akron - Akron - USA


Dept. of Molecular Neurobiology - German Center for Neurodegenerative Diseases (DZNE) - Munich - GERMANY

Approximately 30% of mammalian genes code for transmembrane proteins, which comprise the majority of signal receptors and transducers. These functions are not solely encoded in protein structure, but are also regulated by the unique physicochemical environment of mammalian membranes. A key unmet challenge is to understand the interplay between the composition of membranes, their collective physical properties, and their resulting effect on protein function. The knowledge gap is especially apparent for mammalian neural tissue, whose membranes are highly enriched in omega-3 polyunsaturated fatty acids (PUFAs), which our bodies do not synthesize. This composition is central to neural function as evidenced by brain lipid alterations in numerous developmental, psychological, and neurodegenerative disorders; however the mechanistic relationships between the brain’s unique lipid composition and neurological functions are unknown. Major open questions are how neuronal function is influenced by the lipid content of the membranes that host neural signal transduction receptors, and how factors like diet and environment can influence those lipid compositions. Here, we assess the paradigm-shifting hypothesis that alterations of neuronal membrane lipid composition affect the signaling in the brain and contribute to the pathogenesis of neurological disorders. Particular emphasis is placed on the role of dietary lipids in modulating membrane composition, and the functional consequences thereof. Breakthroughs in understanding the central role of lipids will emerge from the project’s interdisciplinary crosstalk between detailed comprehensive lipidomics, molecular computer simulations, quantitative cellular biophysics, and molecular neurobiology. We focus on two parallel research streams: pattern-recognition receptors and G protein-coupled receptors, which are here used as representative systems to explore the regulation of neural receptors by lipids in a pipeline involving computational, synthetic, and natural model systems, as well as cultured cells and in vivo studies. As the influence of lipids on neuronal receptor function has so far been almost completely ignored, these studies will generate significant impact. Further, the modulation of membrane composition by diet may provide important translational insights and drug-free therapeutic strategies.

2019 -
Long-Term Fellowships - LTF

Unraveling regulation of mutagenesis by DNA damage and antibiotic stress responses in single cells


Department of Biochemistry - University of Oxford - Oxford - UK

UPHOFF Stephan (Host supervisor)

Classical ensemble experiments have been used to characterize DNA damage responses that protect bacterial cells against the toxic and mutagenic effects of DNA damage and stress conditions. More recently, examining the underlying gene regulatory mechanisms at the level of single cells and single molecules revealed unexpected stochastic effects that cause cellular heterogeneity in the damage response. These observations provoke the question whether variations in gene expression modulate DNA repair activities and the rates of mutagenesis of individual cells.

I will address this fundamental question using the E. coli adaptive response to DNA alkylation damage as a model. It has been shown that the regulator protein Ada exhibits extreme variation in gene expression between cells in an isogenic population. Using single-cell expression reporters and single-molecule tracking, I will investigate how this variation affects the expression and DNA repair activities of the genes that are regulated by Ada, namely aidB, alkA and alkB. I will then link phenotypic and genetic variation by whole-genome sequencing of single-cell isolates. This will uncover the impact of DNA repair heterogeneity on mutagenesis at the genome level. Finally, I will investigate the role of the adaptive response in antibiotic-induced mutagenesis.

Because the DNA repair pathways are highly conserved, my findings will also impact our understanding of DNA repair and mutagenesis in eukaryotes.

2019 -
Long-Term Fellowships - LTF

Mechanisms controlling development of aggression


Division of Biology - Caltech - Pasadena - USA

ANDERSON David (Host supervisor)

Aggression, manifested in violence and brutality, poses major risks to our society. While our understanding of the neural correlates of aggression progressed substantially, how aggression is developed and why, remains an open question. Part of this gap is due to the view of the neural circuitry of aggression as fixed, making aggressive behaviors inevitable. Yet, recent advances in the anatomical and genetic basis reveal that aggression has an experience dependent aspect, holding great promise for unraveling the mechanisms of aggression. Aimed at revealing the mechanisms that control aggressive behaviors, this study leverages experience dependent plasticity in tandem with a fusion of circuit and cellular level approaches.
Using a combination of advanced technologies including 2-photon imaging in head fixed mice, single cell RNA sequencing, a combination of 2-photon imaging with in-situ hybridization and gene editing with CRISPR/CAS9, three questions are addressed: 1. What are the changes in neuronal activity following experience dependent plasticity? 2. What are the genetic changes that are associated with the alterations found in neuronal activity? 3. What links these genetic changes to neuronal activity and aggression?
Joining cellular level and circuit level approaches to examine how aggression is developed by experience dependent plasticity should yield a new understanding on the relationship between genes, neuronal function and aggressive behavior. By discovering these basic features in the mechanism underlying aggression, the knowledge obtained from this study could open the path to a new era in which prevention of pathological aggression will become feasible.

2019 -
Cross Disciplinary Fellowships - CDF

Cellular resolution neuronal activation and recording in freely moving flies and fish

VO Doan Tat Thang (VIETNAM)

Institute of Biology I - University of Freiburg - Freiburg - GERMANY

STRAW Andrew D. (Host supervisor)

Functional imaging and optogenetic activation of neural circuits during behavior of untethered animals would allow detailed investigation into the closed-loop interaction of sensory inputs, brain, and motor outputs of behaviors in naturalistic conditions. Previous work investigating the role of Drosophila central complex in navigation and the optic tectum of larval zebrafish in prey capture have been informative, but tethering the animals in such experiments has limited the extent to which circuit mechanisms for multi-sensory, closed-loop control could be investigated. I propose to achieve cellular resolution neural activation and recording in freely moving Drosophila and larval zebrafish, by developing a system for aiming a fast volumetric two-photon microscope to precisely follow, with minimal latency, a region in the animal’s brain. The “feedback based lock-on module” provides low latency feedback to aim the imaging volume of the microscope. Integrating the lock-on module to a fast two-photon microscope based on Bessel beam optics along with an optogenetic stimulator will enable closed-loop analysis of neural mechanism of animal behaviors. The system will then be used to study neural activities of the Drosophila central complex during idiothetic path integration and optic tectum of larval zebrafish during prey capture after expressing GECIs and optogenetic channels into the various sets of neurons using different binary expression systems like GAL4 and LexA.

2019 -
Grant Awardees - Young Investigator Grants

Paradoxical responses of immune systems at the tipping point


Dept. of Bioengineering - Stanford Univeristy - Stanford - USA


French Associates Institute for Agriculture and Biotechnology of Drylands - Blaustein Institutes for Desert Research - Midreshet Ben Gurion - ISRAEL

Basic knowledge is lacking to understand immunological tipping points, where the same stimulus can cause paradoxical immune responses. For example, immune activation after injury or infection is required for downstream tissue repair and pathogen removal, but may also cause tissue damage; immune tolerance or rejection may occur after similar type of tissue transplant, but it is difficult to predict the outcome a priori; inflammation is needed to form blood vessels connecting mother and embryo upon implantation, but too much inflammation can lead to immune disorders during pregnancy. In contrast to these critical clinical complications caused by immune overactivation, cancer cells can bias the tipping point towards the opposite direction to suppress the immune response, but the underlying mechanism still remains unclear. Driven by a series of recent breakthroughs made in our laboratories, we propose to establish planarian flatworms as a novel ideal model to delineate these paradoxical effects: with their simple immune system, we will quantify and manipulate immune cell behaviors both directly in live animals and in engineered ex vivo reconstituted cell systems; with their unsurpassed regenerative ability, we will test the response of immune cells to immunological challenges during allogeneic tissue transplantation. Integrating quantitative experiments, genetic manipulation, and mathematical modeling, we will connect gene functions, cell feedback circuits, and organismal phenotypes, to provide for the first time a multi-scale systems-level mechanism by which the immunological tipping point balances the system sensitivity and robustness. This mechanistic and predictive understanding will also allow synthetically rebuilding immune cell circuits ex vivo for bioengineering and therapeutic purposes. Our work will establish a mechanistic footing to understand more complex immune systems, and can help optimize outcomes in cancer immunotherapy, tissue transplantation, autoimmune diseases, and pregnancy disorders. This work is only achievable through a unique international collaboration that bridge several distinct fields: cellular immunology, functional genomics, and quantitative biology.

2019 -
Grant Awardees - Program Grants

Molecular approaches to study individually identified mechanosensory neurons of the leech


Dept. of Molecular and Cell Biology - University of California, Berkeley - Berkeley - USA


Instituto de Fisiologia Celular-Neurociencias - Universidad Nacional Autónoma de México - Ciudad de Mexico - MEXICO


Institute FBMC - University of Buenos Aires - Caba - ARGENTINA


Dept. of Life Science - National Taiwan University - Taipei - CHINA, REPUBLIC OF (TAIWAN)

To study how nervous systems arise and function, scientists use animal models in which it is possible to integrate research on fundamental processes across different levels of organization from genes to behavior, and from the zygote to the adult. The medicinal leech (genus Hirudo) provides one useful model, because its nervous system is much simpler and easier to work with than vertebrate or mammalian nervous sytems, even though it functions in a similar manner--Hirudo has been used to study phenomena of general importance such as: how glial cells function; how neurotransmitter are released; how synapses form and regenerate; and how neural circuits function to control behavior. Another leech (genus Helobdella) is used to study development and how development changes during evolution, giving rise to kinds of animals over hundreds of millions of years. These two leech species exhibit marked similarities of course, but also some differences. Technical considerations (small embryos for Hirudo; small adults for Helobdella) have made it difficult to integrate these two models, e.g. by applying molecular approaches in Hirudo or to study the adult nervous system in Helobdella. The goals of our project are: 1) to enhance the power of the Hirudo model by introducing newly-developed molecular approaches; 2) to implement approaches in Helobdella that will enable us to unite molecular and cellular approaches to developmental and behavioral neurobiology; 3) to develop new optical techniques for stimulating and recording neuronal activity without exogenous dyes or genetic manipulations.
The intellectual significance of the proposed work is twofold. First, it will enhance our abilities to answer fundamental questions regarding how nervous systems function and development by introducing cutting edge technical approaches to the cellularly simple, physiologically accessible leech models. Of equal importance, it will provide a new evolutionary perspective into neurobiology, by allowing us to examine similarities and differences between leech and other models, including arthropods, nematodes, and vertebrates which have all been evolving separately for more than half a billion years.

2019 -
Grant Awardees - Program Grants

3D atomic-scale movies of molecular machines in action

WEISS Shimon (USA)

Dept. of Chemistry and Biochemistry, Dept. of Physiology - University of California, Los Angeles - Los Angeles - USA


Dept. of Biochemistry - University of Zurich - Zurich - SWITZERLAND


Dept. of Biochemistry - University of Washington - Seattle - USA


Dipartimento di Elettronica, Informazione e Bioingegneria - Politecnico di Milano - Milan - ITALY

Capturing the dynamic 3D atomic-scale structure of a macromolecular machine while it performs its biological function remains an outstanding goal of biology. Conventional structural tools (e.g. X-ray crystallography, NMR & cryoEM) only provide ‘snapshots’ of stable states along a reaction pathway. Reaction intermediates, and in particular short-lived intermediates, are hard to capture and characterize with such conventional techniques. Here we propose to combine (prior) information from multiple existing static structures of stable states with dynamic datasets of inter-atomic distances obtained by high-throughput non-equilibrium single-molecule FRET (smFRET) measurements in a microfluidic mixer using novel time-resolved multi-pixel single-photon avalanche diode detector. These measurements will be performed on libraries of molecular constructs, sampling multiple inter-atomic distances as function of reaction time. These measured distance distributions will then serve as multiple intra- and inter-domain distance constraints which, together with prior information (available structures), will enable the Rosetta software to achieve large-scale energy optimization-based refinement of time-resolved ‘snap shots’ of complex structures with improved accuracy. These time-resolved Rosetta structures together with intermediary molecular dynamics simulations will allow solving the 3D atomic-level structure of the macromolecule for each sampled reaction time point, eventually producing a 3D structural dynamic movie of the macromolecule in action. To demonstrate the utility of the proposed method, we will solve the dynamic structure of RNA polymerase during transcription initiation (promoter binding, bubble opening, abortive initiation, promoter clearance) and a pair of intrinsically disordered proteins (IDPs) involved in transcription regulation (ACTR and NCBD) that adopt a fully folded structure during a coupled folding and binding reaction. In addition to elucidating outstanding questions in transcription by combining detector developments, high-throughput and time-resolved out-of-equilibrium single-molecule FRET measurements with new experimentally-constrained molecular structure computational approaches, this multidisciplinary project will result in a new generic toolkit applicable to a large array of enzymes, proteins and molecular machines.