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.

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.

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.

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.

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.

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.

Transposable elements (TEs) comprise a substantial proportion of all vertebrate genomes, including more than half of our own. They are particularly active in the germ line and during development, and impose an significant evolutionary burden on organisms. As such, numerous mechanisms have evolved to control their spread, including the KRAB zinc finger (KZF) family - the largest and most diverse group of transcription factors in mammals. Strong evidence points to an arms race between TEs and KZFs, and their genome frequencies are highly correlated across diverse vertebrate species. However, KZFs are involved in numerous biological processes beyond TE repression, and we do not yet understand how coevolution with TEs has produced this functional diversity in vertebrates. In addition, whilst the KZFs are mostly limited to tetrapods, other large ZF subfamilies exist outside of this lineage, and the roles of many of these are completely unknown.

Using a comparative and functional genomics approach, I will investigate the dynamics of TE-ZF coevolution, in order to better understand how this phenomenon has shaped the architecture and regulation of vertebrate genomes. Specifically, I will test the hypothesis that TE insertion near ZFs leads to co-regulation of their activity, and furthermore, that this process drives the genomic clustering of ZFs. In parallel, I will use functional techniques and long read re-sequencing to undertake a comprehensive study of of a large, un-described and mysterious family of ZFs in zebrafish. In doing so, I hope to open new lines of research in this essential model organism of embryogenesis and development.

The genome editing field is currently limited in its control over how double-strand breaks can be selectively repaired by non-homologous end joining (NHEJ) or homology-directed repair (HDR) pathways. By providing a donor DNA template, HDR offers potential for corrective therapies targeting a much larger number of genetic diseases which cannot be cured by either single nucleotide substitutions or gene inactivation alone. Yet the efficiency of HDR remains considerably lower than that of NHEJ, hampering its usefulness for therapeutic applications. The use of linear plasmids for enhanced homologous recombination in this proposal directly aims to tackle this challenge and should drive efficient genome editing towards clinical translation.

Unlike circular plasmids which are a ubiquitous tool used throughout modern molecular biology, linear plasmids are an underappreciated biological phenomenon despite being widespread in nature. What constitutes their biological function and what genes they encode is largely unknown. The ability to manipulate linear plasmids in vivo will provide a powerful tool to study and understand their biological significance for the first time. Linear plasmids are double-stranded DNA genetic elements characterised by terminal proteins covalently attached to the 5'-end of each linear DNA strand. In particular, the feature of a terminal protein covalently linked to DNA is a strong prospect for biotechnology applications: I will exploit such linear plasmid systems in mammalian cells for enhanced homologous recombination.

The overall aim of the proposed work is to develop new physical chemistry/bioelectrochemistry techniques in order to better understand the electron-transfer mechanisms of conductive biofilms at the electrode surface. This will have immediate benefits in the development of the promising renewable biophotovoltaic (BPV) technology as well as long-term significance for improvements in understanding the role of key biological electron-transfer species across a broad range of systems.

BPVs use photosynthetic biofilms to harvest solar energy and then convert this into electrical current. They are, however, limited in power output due to unexpectedly slow electron-transfer at the biofilm/electrode interface, and it remains unclear why this is. There is an urgent need to improve fundamental understanding of these electron-transfer mechanisms in order to direct future development of BPVs and remove this bottleneck for their maximum efficiency.

Here, we propose the combination of a number of sophisticated and highly-specialised surface-study techniques in combination with the bioelectrochemistry, synthetic biology and electrode design already in place in the host group, to study the key species (such as the individual redox proteins and complexes) both individually and in situ within model lipid bilayers and finally within the biofilms themselves. Importantly, this will allow us to build up a mechanistic picture of each aspect of these complex buried biointerfaces and hence distinguish between different electron-transfer mechanisms (e.g. via pili aromatic amino acids, outer-membrane redox proteins or diffusional shuttles) that are currently proposed.

Most odontocete cetaceans (toothed whales, dolphins, and porpoises) produce echolocation and communication sounds using bilateral pairs of sound-generating phonic lips. The production of high-frequency echolocation clicks and analysis of the returning echoes allow foraging odontocetes to detect, localize, and capture prey. Communication between odontocete conspecifics is mediated by family-specific sounds, with dolphins producing low-frequency whistles and porpoises emitting high-frequency social clicks. Previous research on the lateralization of sound production in dolphins suggested a left hemisphere bias for echolocation and a right hemisphere bias for conspecific communication; however, similar to other behavioral studies of cetacean lateralization, hemispheric specialization was inferred, rather than measured. Here, I aim to perform the first assessment of hemispheric specialization for the production and perception of echolocation and communication sounds in the bottlenose dolphin (Tursiops truncatus) and harbor porpoise (Phocoena phocoena). Using bioacoustics approaches, novel species-customized electroencephalography-functional near-infrared spectroscopy (EEG-fNIRS) arrays, and functional magnetic resonance imaging (fMRI), I will directly and indirectly measure neural activity underlying motor control and auditory sensation in these odontocetes. In addition to providing unprecedented insights into cetacean brain function, this project has the potential to redefine the limits of neuroscience and animal behavior research through the introduction of portable, non-invasive, and customizable EEG-fNIRS equipment that can be adapted for taxonomically diverse species.

Cytokines act as essential regulators of immunity by eliciting signaling outputs from cell surface receptors. There have been extensive efforts to use cytokines for a wide range of immune disease, including cancer, but their functional pleiotropy has limited their clinical utility. Precedent studies in the Garcia lab, demonstrated that surrogate cytokine ligands can alter signaling output by modifying receptor dimer geometry. For more precise topological control of receptor-ligand interactions and signaling, we hope to create a new class of modular binding scaffold proteins which are applicable to diverse receptor targets. The new scaffolds will be computationally designed and engineered to have three major characteristics: (i) high binding specificity to a target receptor, (ii) self-assembly to form dimers or trimers, (iii) drug-like properties including minimal immunogenicity. These scaffolds will be used for manipulating the geometry of cell surface receptors to fine-tune of signaling outputs. These novel scaffolds can also be utilized to drive the formation of non-natural receptor pairings to elicit completely new functional outputs. Proof-of-concept experiments from Garcia lab have shown that non-natural receptor dimers of receptor tyrosine kinases (RTK) can induce different signaling outputs from each of their parental receptors. We believe that the new scaffolds can efficiently combine any cytokine receptor and RTK to form diverse combinations of dimeric or trimeric receptor pairs. In this way, it will help us to broaden our knowledge on kinase-linked receptor signaling and also have a chance of finding new agonistic and therapeutically meaningful protein ligands.

Intestinal stem cells, which reside in crypts, fuel self-renewal, maintenance, and repair of the epithelial lining of the gut. The regulatory mechanisms that govern maintenance and differentiation of intestinal stem cells, as well as structural and functional differences along regions of the intestine are poorly understood. DNA methylation (DNAme) holds potential in deciphering these mechanisms by revealing gene regulation and clonal structure of cell populations.

The goal is to investigate the dynamics, diversity, and function of DNAme patterns regulating intestinal epithelial homeostasis at single-cell resolution. This proposal will survey single-cell DNAme of intestinal crypts at three anatomical scales: dynamics within individual crypts, diversity between neighboring crypts, and region-specific diversity from proximal to distal sections of the small intestine. Our hypothesis, which builds on preliminary data, is that DNAme patterns of intestinal cells are largely determined by the clonal history of these cells and therefore hold promise as an endogenous lineage tracing tool. To test this hypothesis, I will use a range of single-cell sequencing techniques and develop computational methods to pursue three specific aims:

Aim 1: Determine the single-cell DNAme dynamics within intestinal crypts.

Aim 2: Analyze the diversity of DNAme patterns between crypts along different regions of the intestine.

Aim 3: Disentangle the contribution of cell-type and clonal history in DNAme patterns.

This proposal combines computational with experimental approaches, and leverages existing collaborations between the van Oudenaarden and Clevers labs at the Hubrecht Institute.

The brain has a remarkable capacity for combining long-term memory with exquisite plasticity allowing for adaptation to the environment. A growing aspect of neural plasticity is modulation of myelination. How can myelination be altered? Myelination can be altered by the generation of new myelinating oligodendrocytes or alternatively by mature myelinating oligodendrocytes rendering their existing myelin. The current dogma is that continuous generation of oligodendrocytes underlies myelin remodeling—even though the contribution of mature oligodendrocytes remains poorly understood. Here I propose a multidisciplinary approach, combining my expertise in human cell biology with Dr. Chan’s extensive proficiency in oligodendrocyte biology to study the cellular and molecular mechanisms underlying myelin alterations upon induced neuronal activity and after injury. I plan to explore the myelinogenic potential of mature oligodendrocytes and their contribution to myelin remodeling. This strategy combines genetic fate-mapping and manipulation of oligodendrocyte lineage cells, in vivo pharmacogenetic neuronal stimulation (DREADDs) with single cell, temporal-resolved gene expression analysis and behavioral assessment. More specifically, genetically altered progenitors unable to generate new oligodendrocytes and mature oligodendrocytes will be labeled and profiled using single cell RNA-seq. This will allow for the longitudinal assessment of genes involved in activity-dependent myelination and the consequence of this manipulation. Knowledge of myelin plasticity is essential for the development of new treatments for demyelinating disease such as multiple sclerosis.

Neurons are the most diverse class of nervous system cell types, based on thorough characterization of single cells into molecular taxonomies. Individually neurons are fascinating and complicated, but at the core of computation are their interactions. The unique arrangement of circuits between specialized neuronal types results in a wide range of sophisticated functions and complex behaviors. While female and male brains largely perform the same sophisticated functions, the neurobiological basis for gender differences in some behaviors has fascinated researchers over decades. Sexual dimorphism in the amygdala (specifically the posterodorsal medial amygdala, MePD) shapes gender differences in reproductive behavior, aggression or social recognition, with well-established contribution of gonadal steroid hormones. At the core of this dimorphism are neuroanatomical differences of the MePD, such as regional and cell volume, cell number and cell circuitry. While the phenomenon is well established using relatively simple methods, a more global, systematic characterization of differences in circuits in the context of cell types and gene expression has not been technically possible. To this end, I propose to develop and apply an integrated approach for simultaneous connectomics and transcriptomics measurements of the medial amygdala at single cell resolution. This study will reveal the contribution of circuits, cell types and gene expression to gender-based amygdala dimorphism. Further, the proposed framework to understanding circuit implementations in the context of cell types will advance other studies of nervous system function and behavioral neuroscience.

The goal of this proposal is to understand how heterochromatin functions impose major roadblocks to cell fate changes in development and iPS cell reprogramming. Reprogramming somatic cells into pluripotency by the Yamanaka factors has enormous potential in the regenerative field, although the efficiency remains low. Heterochromatin functions as a major epigenetic barrier to cell fate changes and the Zaret lab showed that its erasure on pluripotent genes at the late stage of reprogramming is a rate-limiting step. However, so far little is known about how heterochromatin domains are established, maintained and reset during development and reprogramming. To fill in this gap, we propose to map the dynamics of heterochromatin formation during mouse peri-implantation, when heterochromatin domains are first established, and to map heterochromatin dynamic resetting in reprogramming intermediates. Taking both natural and reprogramming contexts together, we will construct lineage relationships of heterochromatin dynamics at pluripotency genes as they related to transcription. Finally, to gain temporal and mechanistic insights into heterochromatin regulations of pluripotent genes, we will use a live imaging system to assess directly the consequences of systematic knockdown heterochromatin-associated proteins on the transitions between chromatin topologies and transcriptional activation of pluripotent genes and reprogramming efficiency. Our novel approach of combining datasets and analysis of early development and late reprogramming will contribute to designing robust reprogramming strategies with higher efficiencies and realizing full the potential of iPS cells.

The OXPHOS system is the only process in animal cells with components encoded by two genomes, maternally transmitted mitochondrial DNA (mtDNA) and biparentally transmitted nuclear DNA (nDNA). The protein products of both genomes have to physically assemble with their counterparts to build functional respiratory complexes. Therefore, variability in the OXPHOS encoded genes is limited by a physical match constraint. This imposes a close-fitting co-evolution of both genomes challenged by the very different mechanism to generate variability for nDNA (by sexual reproduction, mutation and co-existence of two alleles) and mtDNA encoded OXPHOS genes (by mutation, polyploidy and segregation). Since the simultaneous co-existence of alternative mtDNA encoded alleles for OXPHOS proteins has been shown to be detrimental for the organism, we postulate that the co-expression of nuclear encoded alternative alleles may have similar adverse consequences, and that specific regulatory mechanism prevent them. Indeed, random mono-allelic expression is not rare and was suggested for about 30 OXPHOS genes in mice. We postulate that is part of a sophisticated and multi-level system of quality control and functional testing to select for the best combination for providing metabolic plasticity.
Potential regulatory mechanisms are: selective transcription of one allele per cell or selection of the expressed alleles at the import or assembly of the OXPHOS complexes. For testing this hypothesis, we integrate different skills for the analysis of mitochondrial functional and genetics profiling (Enriquez), single cell transcriptomic (Eberwine) and functional and dynamic analysis (Busch) in mouse (Enriquez) and zebrafish (Mercader). If confirmed, we will set the ground for a novel theory of genetic interaction for OXPHOS function. If discarded, the existence of mtDNA and its particular way of inheritance would be necessary to group those genes for which allelic variability is detrimental.

One of the most basic aspects of sleep is that it happens at a particular time of day. Neuroscientists have known for almost half a century that this consolidation requires the suprachiasmatic nuclei of the hypothalamus (SCN); beyond this point, the circuit remains untraced. Equally mysterious, healthy young humans sleep in a single consolidated bout, while infants and older individuals, as well as laboratory mice, can have highly fragmented sleep. Informed by a combined cellular and circuit-based model for slow-wave or “deep” sleep (Forger Group), we propose to trace the signals leading from SCN to cortical slow waves at both physical and molecular levels, and then manipulate them to artificially consolidate sleep. Starting from cortex, aided by a novel “triple-CRISPR” approach to generate knockout mice efficiently in a single generation, we shall examine roles of individual cortical ion channels and the signaling pathways they regulate in creating these slow waves both in vivo and in newly developed whole-brain culture (Ueda Group). This information will be incorporated into a cellular and then a thalamocortical model of slow-wave sleep (Forger Group). In parallel, synaptic tracing in cleared brain, as well as calcium imaging correlation analysis using miniature skull-mounted microscopes, will establish physical and functional connectivity from SCN to cortex (Brown Group), which can also be tested in whole-brain culture (Ueda group). Modeling these data comprehensively and then optimizing this whole-brain model (Forger group), we can predict and test molecular sources of homeostatic and circadian influence upon sleep, as well as combined transgenic and optogenetic strategies to create consolidated bouts of sleep from the normally fragmented sleep of the mouse (Brown and Ueda groups). In this way, by using the power of large-scale quantitative modeling to explore synergies in sleep-dependent signaling, we hope to provide a starting point for novel multimodal therapies with the capacity to fundamentally alter the sleep-wake landscape.

Enhancers are relatively short DNA sequence elements (<1 kb) that determine the timing, location and levels of gene transcription. They harbor specific binding sites for transcription factors (TFs) that control the enhancer. While the necessity of particular TF binding sites (TFBSs) can be assayed with mutations, it is not yet understood which DNA features in an enhancer sequence collectively give rise to the regulatory activity. To identify the molecular determinants that impart a regulatory activity to a DNA sequence element, we have devised a quantitative experimental paradigm and propose to apply statistical analysis and machine learning approaches to the molecular dissection of a model enhancer.
Specifically, we will use a model enhancer driving patterned spatial expression in the wings of fruit flies (Drosophila). First, we will create tens of variants of this enhancer, introducing mutations along its sequence, and describe their regulatory effect. To this end, we will build an automated imaging pipeline to measure the levels and spatial distribution of reporter gene in the wings. The resulting quantitative expression data in a flat tissue will define a morphospace, i.e., a mathematical multidimensional space representing the possible variation of enhancer activity. Second, we will extract DNA feature sets to quantitatively describe molecular variation along the sequence of our mutant enhancers. This will capture structural changes (DNA shape readout), changes in nucleotide sequence per se (DNA base readout), as well as other features (e.g., TFBSs). Third, leaning on dimensionality reduction techniques and machine learning, we will develop predictive models that describe the relationship between DNA features and morphospace. Finally, we will test our predictions of enhancer functionality using synthetic enhancers in transgenic flies. Because enhancers are complex biological objects, we aim with this proposal at developing appropriate mathematical tools to capture the essence of this complexity.
Since this proposal aims at developing a comprehensive mathematical approach to modeling of enhancer functionality, it has the potential to unravel complex molecular mechanisms underlying transcriptional regulation.

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

Toxoplasma is a common parasite of birds, rodents, and humans during the asexual phase of its life cycle. But, having sex for Toxoplasma is limited to cat intestines. It enters cat when an infected rat or bird is eaten up. This creates an interesting problem because rats do not prefer to be eaten up. Toxoplasma blocks the fear of cats from infected rats. This is often taken to mean that infected rats will approach cat without fear and be eaten up at higher rates. We have no direct evidence for this indirect assumption. We will test this in the first part of this project. Kangaroo Island in Australia plans to fence off one of its parts and exterminate all cats from there. The island is doing so to protect wildlife and to reduce Toxoplasmosis in livestock. From our point of view, this is a golden opportunity to test if Toxoplasma increases predation of rats by cats. We can now compare the survival of infected rats and mice vis-à-vis uninfected rats and mice in the same landscape before and after cat removal. We expect to see more infected animals dying than uninfected when cats are around; and no difference when cats are removed.
While having sex is limited to cats, Toxoplasma can still sustain its asexual life cycle through carnivory or sexual reproduction. The sexual reproduction is exciting because such infections usually have less virulence. They do not kill a host very often as dead hosts do not reproduce to further transmit the parasite. Parasites moving across the food chain, from rats to cats for example, do not have such limitation and often increase virulence. But what happens when an infection is simultaneously transmitted by sexual and going-up-food chain mode? Both paths must trade-off with each other. When cats are plentiful it pays to increase predation (by reducing fear in infected rats) even if it means a bit less of sexual transmission and vice versa. This is a difficult assumption to test directly. The situation in Dudley Peninsula provides a perfect opportunity. We will collect Toxoplasma from bodies of infected rats and mice before and after cat extermination. We will bring these bugs in the lab and ask if cat removal means that the parasite now concentrates more on sexual transmission. That would mean a robust invasion of testes, more pheromones and more liking elicited from uninfected females. This is the second part of this project.