ERC FUNDED PROJECTS

My proposal aims to take advantage of my ground-breaking finding that it is possible to mature, or activate, the [FeFe] hydrogenase enzyme (HydA) using synthetic mimics of its catalytic [2Fe] cofactor. (Berggren et al, Nature, 2013) We will now explore the chemistry and (bio-)technological potential of the enzyme using an interdisciplinary approach ranging from in vivo biochemical studies all the way to synthetic model chemistry. Hydrogenases catalyse the interconversion between protons and H2 with remarkable efficiency. Consequently, they are intensively studied as alternatives to Pt-catalysts for these reactions, and are arguably of high (bio-) technological importance in the light of a future “hydrogen society”. The project involves the preparation of novel “artificial” hydrogenases with the primary aim of designing spectroscopic model systems via modification(s) of the organometallic [2Fe] subsite. In parallel we will prepare in vitro loaded forms of the maturase HydF and study its interaction with apo-HydA in order to further elucidate the maturation process of HydA. Moreover we will develop the techniques necessary for in vivo application of the artificial activation concept, thereby paving the way for a multitude of studies including the reactivity of artificial hydrogenases inside a living cell, but also e.g. gain-of-function studies in combination with metabolomics and proteomics. Inspired by our work on the artificial maturation system we will also draw from our knowledge of Nature’s [FeS] cluster proteins in order to prepare a novel class of “miniaturized hydrogenases” combining synthetic [4Fe4S] binding oligopeptides with [2Fe] cofactor model compounds. Our interdisciplinary approach is particularly appealing as it not only provides further insight into hydrogenase chemistry and the maturation of metalloproteins, but also involves the development of novel tools and concepts applicable to the wider field of bioinorganic chemistry.

1 494 880 €

Start date: 2017-02-01, End date: 2022-01-31

The packaging of genetic information into chromatin regulates a wide range of vital processes that depend on direct access to the DNA template. Many chromatin-interacting complexes impact chromatin structure and their aberrant regulation or dysfunction has been implicated in various cancers and severe developmental disorders. A better understanding of the roles of chromatin-interacting complexes in such disease states requires a detailed mechanistic study. Many chromatin-interacting complexes modify chromatin structure, yet understanding the underlying mechanisms remains a major challenge in the field. Furthermore, how chromatin-interacting complexes are regulated to enable their various functions is incompletely understood. We will address these longstanding questions in two specific aims. Aim I: Building on our expertise in single-molecule biology, we will develop powerful single-molecule imaging approaches to monitor the action of chromatin-interacting complexes in real time. We will further probe how the diverse activities of the chromatin-associated complexes are coordinated and coupled to conformational transitions. Aim II: Drawing on our expertise in structural biology, we will use a range of structural techniques in combination with biochemical approaches to study the vital regulation of chromatin-interacting complexes by their regulatory subunits as well as by chromatin features. We expect to obtain ground-breaking insights into the mechanisms and regulation of disease-related chromatin-associated complexes, which may open up new horizons for developing therapeutic intervention strategies. Furthermore, the approaches developed here will enable the investigation of a large number of chromatin-related processes.

1 498 954 €

Start date: 2017-03-01, End date: 2022-02-28

At the dawn of the 21st century, our knowledge of the molecular mechanism of mammalian fertilization remains very limited. Different lines of evidence indicate that initial gamete recognition depends on interaction between a few distinct proteins on sperm and ZP3, a major component of the extracellular coat of oocytes, the zona pellucida (ZP). On the other hand, recent findings suggest an alternative mechanism in which cleavage of another ZP subunit, ZP2, regulates binding of gametes by altering the global structure of the ZP. Progress in the field has been hindered by the paucity and heterogeneity of native egg-sperm recognition proteins, so that novel approaches are needed to reconcile all available data into a single consistent model of fertilization. Following our recent determination of the structure of the most conserved domain of sperm receptor ZP3 by X-ray crystallography, we will conclusively establish the basis of mammalian gamete recognition by performing structural studies of homogeneous, biologically active recombinant proteins. First, we will combine crystallographic studies of isolated ZP subunits with electron microscopy analysis of their filaments to build a structural model of the ZP. Second, structures of key egg-sperm recognition protein complexes will be determined. Third, we will investigate how proteolysis of ZP2 triggers overall conformational changes of the ZP upon gamete fusion. Together with functional analysis of mutant proteins, these studies will provide atomic resolution snapshots of the most crucial step in the beginning of a new life, directly visualizing molecular determinants responsible for species-restricted gamete interaction at fertilization. The progressive decrease of births in the Western world and inadequacy of current contraceptive methods in developing countries underscore an urgent need for a modern approach to reproductive welfare. This research will not only shed light on a truly fundamental biological problem, but also constitute a solid foundation for the reproductive medicine of the future.

1 499 282 €

Start date: 2011-01-01, End date: 2015-12-31

In human cells, two meters of DNA sequence are compressed into a nucleus whose linear size is five orders of magnitude smaller. Deciphering how this amazing structural organization is achieved and how DNA functions can ensue in the environment of a cell’s nucleus represent central questions for contemporary biology. Here, I embrace this challenge by establishing a comprehensive framework of microscopy and sequencing technologies coupled with advanced analytical approaches, aimed at addressing three fundamental highly-interconnected questions: 1) What are the design principles that govern DNA compaction? 2) How does genome structure vary between different cell types as well as among cells of the same type? 3) What is the link between genome structure and function? In preliminary experiments, we have devised a powerful method for Genomic loci Positioning by Sequencing (GPSeq) in fixed cells with optimally preserved nuclear morphology. In parallel, we are developing high-end microscopy tools for simultaneous localization of dozens of genomic locations at high resolution in thousands of single cells. We will obtain first-ever genome-wide maps of radial positioning of DNA loci in the nucleus, and combine them with available DNA contact probability maps in order to build 3D models of the human genome structure in different cell types. Using microscopy, we will visualize chromosomal shapes at unprecedented resolution, and use these rich datasets to discover general DNA folding principles. Finally, by combining high-resolution chromosome visualization with gene expression profiling in single cells, we will explore the link between DNA structure and function. Our study shall illuminate the design principles that dictate how genetic information is packed and read in the human nucleus, while providing a comprehensive repertoire of tools for studying genome organization.

1 499 808 €

Start date: 2018-01-01, End date: 2022-12-31

Computer vision concerns itself with understanding the real world through the analysis of images. Typical problems are object recognition, medical image segmentation, geometric reconstruction problems and navigation of autonomous vehicles. Such problems often lead to complicated optimization problems with a mixture of discrete and continuous variables, or even infinite dimensional variables in terms of curves and surfaces. Today, state-of-the-art in solving these problems generally relies on heuristic methods that generate only local optima of various qualities. During the last few years, work by the applicant, co-workers, and others has opened new possibilities. This research project builds on this. We will in this project focus on developing new global optimization methods for computing high-quality solutions for a broad class of problems. A guiding principle will be to relax the original, complicated problem to an approximate, simpler one to which globally optimal solutions can more easily be computed. Technically, this relaxed problem often is convex. A crucial point in this approach is to estimate the quality of the exact solution of the approximate problem compared to the (unknown) global optimum of the original problem. Preliminary results have been well received by the research community and we now wish to extend this work to more difficult and more general problem settings, resulting in thorough re-examination of algorithms used widely in different and trans-disciplinary fields. This project is to be considered as a basic research project with relevance to industry. The expected outcome is new knowledge spread to a wide community through scientific papers published at international journals and conferences as well as publicly available software.

1 440 000 €

Start date: 2008-07-01, End date: 2013-06-30

The length of telomeric DNA is a critical determinant factor for aging and cancer development. In germ cells, the activation of a telomerase-dependent telomere-lengthening pathway is thought to be important in order to maintain telomeric DNA across generations, but the molecular mechanisms involved in this pathway, i.e: how and when telomerase is activated in germ cells, are largely unknown. A DNA-binding protein complex called shelterin constitutively binds telomeric DNA. However, my recent studies have suggested that a multi-subunit DNA-binding complex, TERB1-TERB2-MAJIN, takes over telomeric DNA from shelterin in mammalian germ cells in order to facilitate homologous recombination. These findings represent a hitherto unknown molecular mechanism at work on the telomeres in germ cells. In this project, I hypothesize that the drastic reformation of telomere-binding complexes in germ cells contributes also to the telomere-lengthening pathway. The aim of this project is to test this hypothesis in order to reveal the mechanism underlying the transgenerational inheritance of telomeric DNA throughout meiosis. This work is divided into three work packages. WP1: to determine the molecular rearrangements that take place at telomeres during meiosis. WP2: to determine how and when telomeres are lengthened during germ cell production. WP3: to determine how meiotic recombination is achieved. The proposed project will reveal molecular mechanisms underlying the transgenerational inheritance of genetic information after meiosis, and this will increase our understanding of the etiology of numerous human diseases caused by meiotic errors, such as congenital birth defects and aneuploidy. Further, because the misregulation of telomerase is a leading cause of cancer development, the identification of telomerase-activating mechanisms in germ cells will have multidiscipline impacts in both cancer and reproductive biology fields and will be useful for developing novel cancer therapies.

1 500 000 €

Start date: 2019-01-01, End date: 2023-12-31

Mitochondria and mitochondrial function have gained increased attention within a wide range of clinical and scientific specialities, but exactly how mitochondria impact the rest of the cell is less well understood. Not only are mitochondria implicated in a range of rare genetic disorders, but dysfunction of mitochondria or reduced bioenergetic capacity has been associated with common diseases including cancer, heart failure, neurodegeneration and diabetes mellitus, as well as natural ageing. It is becoming increasingly clear that mitochondrial dysfunction is not only a downstream event in these conditions, but plays an important role in disease progression and pathology. S-adenosylmethionine (SAM) is the dominant methyl group donor within our cells, required for a diverse set of post-translational modifications, nucleotide methylations or the synthesis of co-factors and metabolites. Mitochondria play an important part in SAM synthesis, and mitochondrial function has recently been shown to influence cellular methylation. Approximately 30% of the cellular SAM pool is located within mitochondria, advocating a central role for mitochondria in cellular methylation. The advancements in genome sequencing techniques, unprecedented depth of modern mass spectrometry analyses and our possibility to efficiently generate model systems, provides a rare opportunity to comprehensively study the role of both SAM and mitochondria in health and disease. This project plan describes the genetic, molecular, metabolic and proteomic analysis of fruit fly and mouse models with mitochondrial dysfunction and disrupted intra-mitochondrial SAM levels to identify the mitochondrial methylome, its relevance towards other cellular functions and its impact on the epigenetic control of gene regulation. My extensive research on mitochondrial function, as well as working as a physician with patients suffering from inborn errors of metabolism gives me a unique perspective in this project.

1 499 999 €

Start date: 2017-04-01, End date: 2022-03-31

Semiconductor nanowires composed of III-V materials have enormous potential to add new functionality to electronics and optical applications. However, integration of these promising structures into applications is severely limited by the current near-universal reliance on gold nanoparticles as seeds for nanowire fabrication. Although highly controlled fabrication is achieved, this metal is entirely incompatible with the Si-based electronics industry. It also presents limitations for the extension of nanowire research towards novel materials not existing in bulk. To date, exploration of alternatives has been limited to selective-area and self-seeded processes, both of which have major limitations in terms of size and morphology control, potential to combine materials, and crystal structure tuning. There is also very little understanding of precisely why gold has proven so successful for nanowire growth, and which alternatives may yield comparable or better results. The aim of this project will be to explore alternative nanoparticle seed materials to go beyond the use of gold in III-V nanowire fabrication. This will be achieved using a unique and recently developed capability for aerosol-phase fabrication of highly controlled nanoparticles directly integrated with conventional nanowire fabrication equipment. The primary goal will be to deepen the understanding of the nanowire fabrication process, and the specific advantages (and limitations) of gold as a seed material, in order to develop and optimize alternatives. The use of a wide variety of seed particle materials in nanowire fabrication will greatly broaden the variety of novel structures that can be fabricated. The results will also transform the nanowire fabrication research field, in order to develop important connections between nanowire research and the semiconductor industry, and to greatly improve the viability of nanowire integration into future devices.

1 496 246 €

Start date: 2013-09-01, End date: 2018-08-31

Protein synthesis in mitochondria is essential for the bioenergetics, whereas its counterpart in chloroplasts is responsible for the synthesis of the core proteins that ultimately converts sunlight into the chemical energy that produces oxygen and organic matter. Recent insights into the mito- and chlororibosomes have provided the first glimpses into the distinct and specialized machineries that involved in synthesizing almost exclusively hydrophobic membrane proteins. Our findings showed: 1) mitoribosomes have different exit tunnels, intrinsic GTPase in the head of the small subunit, tRNA-Val incorporated into the central protuberance; 2) chlororibosomes have divaricate tunnels; 3) ribosomes from both organelles exhibit parallel evolution. This allows contemplation of questions regarding the next level of complexity: How these ribosomes work and evolve? How the ribosomal components imported from cytosol are assembled with the organellar rRNA into a functional unit being maturated in different compartments in organelles? Which trans-factors are involved in this process? How the chlororibosomal activity is spatiotemporally coupled to the synthesis and incorporation of functionally essential pigments? What are the specific regulatory mechanisms? To address these questions, there is a need to first to characterize the process of translation in organelles on the structural level. To reveal molecular mechanisms of action, we will use antibiotics and mutants for pausing in different stages. To reconstitute the assembly, we will systematically pull-down pre-ribosomes and combine single particle with tomography to put the dynamic process in the context of the whole organelle. To understand co-translational operations, we will stall ribosomes and characterize their partner factors. To elucidate the evolution, we will analyze samples from different species. Taken together, this will provide fundamental insights into the structural and functional dynamics of organelles.

1 331 300 €

Start date: 2019-05-01, End date: 2024-04-30

Bacterial infections represent a major and global health problem, which is further aggravated by the rapid and ongoing increase in antibiotic resistance. The limited success in the development of targeted therapies for particular invasive strains can be attributed to our limited knowledge how pathogens modulate their proteome homeostasis in vivo, knowledge that has so far remained elusive due to technical limitations. Here I propose the use of proteome-wide selected reaction monitoring mass spectrometry (SRM-MS) for pathogen proteome profiling from samples obtained directly from in vivo using group A streptococci (GAS) as a model system. The proposal describes the use of SRM-MS to facilitate the construction of comprehensive and quantitative molecular anatomy knowledge models outlining spatial organization, pathway organization, absolute protein concentration estimations and interaction partners with host for complete microbial proteomes. Using the molecular anatomy as benchmark I intend compare how the proteome homeostasis is controlled in pathogens isolated directly from patients with different degree of disease severity to understand how disease severity, anatomical location and host dependencies effects the proteome homeostasis. The outlined proposal describes a generic strategy to provide comprehensive understanding of the pathogen adaption directly in vivo and represents a paradigm shift in the field of bacterial infectious disease. This proposal addresses central aspects within the medical microbiology field that has been long sought for but never studied due to technology limitations and will influence the development of the next generation targeted vaccine and therapeutic development programs.

1 498 699 €

Start date: 2013-05-01, End date: 2018-04-30

SUMMARY Mitochondria are required to convert food into usable energy forms and every cell contains thousands of them. Unlike most other cellular compartments, mitochondria have their own genomes (mtDNA) that encode for 13 of the about 90 proteins present in the respiratory chain. All proteins necessary for mtDNA replication, as well as transcription and translation of mtDNA-encoded genes, are encoded in the nucleus. Mutations in nuclear-encoded proteins required for mtDNA maintenance is an important cause of neurodegeneration and muscle diseases. The common result of these defects is either mtDNA depletion or accumulation of multiple deletions of mtDNA in postmitotic tissues. Research in my laboratory will elucidate the molecular mechanisms and regulation of mitochondrial DNA replication in human cells. We will establish how mtDNA is packaged into nucleoprotein complexes, a.k.a. nucleoids and establish how these nucleoids are selected for mtDNA replication. We will elucidate the molecular mechanisms by which specific mutations in the mtDNA replication machinery affect mtDNA maintenance and cause human disease. Mitochondrial dysfunction is not limited to rare, genetic disorders, but also associated with age-associated common diseases as well as with the normal aging process. I will use my biochemical insights in combination with mouse genetics to address the hypothesis that increased mtDNA mutation load may be an important cause of normal aging. My specific aims will be: Aim 1. To define how initiation of mtDNA replication at OriH is regulated. Aim 2. To identify and characterize regulators of mtDNA replication. Aim 3. To characterize the structure and function of the mtDNA nucleoid in DNA replication. Aim 4. To address the mitochondrial theory of ageing

1 492 684 €

Start date: 2011-04-01, End date: 2016-03-31

"The development of micro fabrication and field effect transistors are key enabling technologies for todays information society. It is hard to imagine superfast and omnipresent electronic devices, information technology, the Internet and mobile communication technologies without access to continuously cheaper and miniaturized microprocessors. The giant leaps in performance of microprocessors from the first personal computing machines to todays mobile devices are to a large extent realized via miniaturization of the active components. The ultimate limit of miniaturization of electronic components is the realization of single molecule electronics. Due to fundamental physical limitations, single molecule resolution cannot be achieved using classical top-down lithographic techniques. At the same time, existing surface functionalization schemes do not provide any means of placing a single molecule with high precision at a specific location on a nanostructure. This project has the ambitious goal of establishing the first method ever allowing for self-assembly of multiple single molecule devices in a parallel way and thereby provide the first method ever allowing for multiple individual single molecule components to operate together in the same device. The impact of the technology platforms described herein goes vastly beyond the field of single molecule electronics and utilization in ultra-sensitive plasmonic biosensors with a digital single molecule response will be explored in parallel with the main roadmaps of the project."

1 500 000 €

Start date: 2014-02-01, End date: 2019-01-31

We aim to study transcriptomes with single-cell resolution, a long-standing goal in biology, to answer fundamental questions about gene regulation. The main objective concerns gene regulation during in vivo differentiation and in pluripotent cells by studying single-cells from murine preimplantation embryos, a model system with natural single-cell resolution, important biology and medical potential. This would also allow us to explore general regulatory principles of gene expression programs of individual cells. This research program will be accomplished by novel deep sequencing technology of mRNAs (mRNA-Seq) to obtain quantitative, unbiased and genome-wide gene and isoform expression measurements. We are therefore developing new experimental and computational methods for genome-wide analyses of transcriptomes at single-cell resolution. The biological significances of the proposed research are unique insights into early embryonic development. Deep sequencing of transcriptomes will also reveal post-transcriptional gene regulation important for pluripotent cells and identified pluripotency-specific gene and isoform expressions will be important for future stem cell based therapies. The inherit single-cell nature of the model system together with its important biology makes it a model systems exceptionally well suited for a systems biology approach aiming to characterize gene regulation at single-cell resolution. The novel methodology has tremendous potential to enable complete mRNA characterization of individual cells. The deep sequencing approach with state-of-the-art computational analyses is both more quantitative than previous methods and it will give readouts on alternative isoforms generated by alternative promoters, splicing and polyadenylation.

1 654 384 €

Start date: 2010-02-01, End date: 2015-01-31

Imagine a world, in which countless embedded microelectronic components continuously monitor our health and allow us to seamlessly interact with our digital environment. One particularly promising platform for the realisation of this concept is based on wearable electronic textiles. In order for this technology to become truly pervasive, a myriad of devices will have to operate autonomously over an extended period of time without the need for additional maintenance, repair or battery replacement. The goal of this research programme is to realise textile-based thermoelectric generators that without additional cost can power built-in electronics by harvesting one of the most ubiquitous energy sources available to us: our body heat. Current thermoelectric technologies rely on toxic inorganic materials that are both expensive to produce and fragile by design, which renders them unsuitable especially for wearable applications. Instead, in this programme we will use polymer semiconductors and nanocomposites. Initially, we will focus on the preparation of materials with a thermoelectric performance significantly beyond the state-of-the-art. Then, we will exploit the ease of shaping polymers into light-weight and flexible articles such as fibres, yarns and fabrics. We will explore both, traditional weaving methods as well as emerging 3D-printing techniques, in order to realise low-cost thermoelectric textiles. Finally, within the scope of this programme we will demonstrate the ability of prototype thermoelectric textiles to harvest a small fraction of the wearer’s body heat under realistic conditions. We will achieve this through integration into clothing to power off-the-shelf sensors for health care and security applications. Eventually, it can be anticipated that the here interrogated thermoelectric design paradigms will be of significant benefit to the European textile and health care sector as well as society in general.

1 500 000 €

Start date: 2015-06-01, End date: 2020-05-31