ERC FUNDED PROJECTS

The architecture of DNA in the cell nucleus is an emerging epigenetic key contributor to genome function. We recently developed 4C technology, a high-throughput technique that combines state-of-the-art 3C technology with tailored micro-arrays to uniquely allow for an unbiased genome-wide search for DNA loci that interact in the nuclear space. Based on 4C technology, we were the first to provide a comprehensive overview of long-range DNA contacts of selected loci. The data showed that active and inactive chromatin domains contact many distinct regions within and between chromosomes and genes switch long-range DNA contacts in relation to their expression status. 4C technology not only allows investigating the three-dimensional structure of DNA in the nucleus, it also accurately reconstructs at least 10 megabases of the one-dimensional chromosome sequence map around the target sequence. Changes in this physical map as a result of genomic rearrangements are therefore identified by 4C technology. We recently demonstrated that 4C detects deletions, balanced inversions and translocations in patient samples at a resolution (~7kb) that allowed immediate sequencing of the breakpoints. Excitingly, 4C technology therefore offers the first high-resolution genomic approach that can identify both balanced and unbalanced genomic rearrangements. 4C is expected to become an important tool in clinical diagnosis and prognosis. Key objectives of this proposal are: 1. Explore the functional significance of DNA folding in the nucleus by systematically applying 4C technology to differentially expressed gene loci. 2. Adapt 4C technology such that it allows for massive parallel analysis of DNA interactions between regulatory elements and gene promoters. This method would greatly facilitate the identification of functionally relevant DNA elements in the genome. 3. Develop 4C technology into a clinical diagnostic tool for the accurate detection of balanced and unbalanced rearrangements.

1 225 000 €

Start date: 2008-09-01, End date: 2013-08-31

The Tera-Hertz portion of the spectrum presents unique potentials for advanced applications. Currently the THz spectrum is revealing the mechanisms at the origin of our universe and provides the means to monitor the health of our planet via satellite based sensing of critical gases. Potentially time domain sensing of the THz spectrum will be the ideal tool for a vast variety of medical and security applications. Presently, systems in the THz regime are extremely expensive and consequently the THz spectrum is still the domain of only niche (expensive) scientific applications. The main problems are the lack of power and sensitivity. The wide unused THz spectral bandwidth is, herself, the only widely available resource that in the future can compensate for these problems. But, so far, when scientists try to really use the bandwidth, they run into an insurmountable physical limit: antenna dispersion. Antenna dispersion modifies the signal’s spectrum in a wavelength dependent manner in all types of radiation, but is particularly deleterious to THz signals because the spectrum is too wide and with foreseeable technology it cannot be digitized. The goal of this proposal is to introduce break-through antenna technology that will eliminate the dispersion bottle neck and revolutionize Time Domain sensing and Spectroscopic Space Science. Achieving these goals the project will pole vault THz imaging technology into the 21-th century and develop critically important enabling technologies which will satisfy the electrical engineering needs of the next 30 years and in the long run will enable multi Tera-bit wireless communications. In order to achieve these goals, I will first build upon two major breakthrough radiation mechanisms that I pioneered: Leaky Lenses and Connected Arrays. Eventually, ultra wide band imaging arrays constituted by thousands of components will be designed on the bases of the new theoretical findings and demonstrated.

1 499 487 €

Start date: 2011-11-01, End date: 2017-10-31

Cell volume regulation is crucial for any living cell because changes in volume determine the metabolic activity through e.g. changes in ionic strength, pH, macromolecular crowding and membrane tension. These physical chemical parameters influence interaction rates and affinities of biomolecules, folding rates, and fold stabilities in vivo. Understanding of the underlying volume regulatory mechanisms has immediate application in biotechnology and health, yet these factors are generally ignored in systems analyses of cellular functions. My team has uncovered a number of mechanisms and insights of cell volume regulation. The next step forward is to elucidate how the components of a cell volume regulatory circuit work together and control the physicochemical conditions of the cell. I propose construction of a synthetic cell in which an osmoregulatory transporter and mechanosensitive channel form a minimal volume regulatory network. My group has developed the technology to reconstitute membrane proteins into lipid vesicles (synthetic cells). One of the challenges is to incorporate into the vesicles an efficient pathway for ATP production and maintain energy homeostasis while the load on the system varies. We aim to control the transmembrane flux of osmolytes, which requires elucidation of the molecular mechanism of gating of the osmoregulatory transporter. We will focus on the glycine betaine ABC importer, which is one of the most complex transporters known to date with ten distinct protein domains, transiently interacting with each other. The proposed synthetic metabolic circuit constitutes a fascinating out-of-equilibrium system, allowing us to understand cell volume regulatory mechanisms in a context and at a level of complexity minimally needed for life. Analysis of this circuit will address many outstanding questions and eventually allow us to design more sophisticated vesicular systems with applications, for example as compartmentalized reaction networks.

2 247 231 €

Start date: 2015-07-01, End date: 2020-06-30

In mammals, gene dosage of X-chromosomal genes is equalized between sexes by random inactivation of either one of the two X chromosomes in female cells. In the initial phase of X chromosome inactivation (XCI), a counting and initiation process determines the number of X chromosomes per nucleus, and elects the future inactive X chromosome (Xi). Xist is an X-encoded gene that plays a crucial role in the XCI process. At the start of XCI Xist expression is up-regulated and Xist RNA accumulates on the future Xi thereby initiating silencing in cis. Recent work performed in my laboratory indicates that the counting and initiation process is directed by a stochastic mechanism, in which each X chromosome has an independent probability to be inactivated. We also found that this probability is determined by the X:ploïdy ratio. These results indicated the presence of at least one X-linked activator of XCI. With a BAC screen we recently identified X-encoded RNF12 to be a dose-dependent activator of XCI. Expression of RNF12 correlates with Xist expression, and a heterozygous deletion of Rnf12 results in a marked loss of XCI in female cells. The presence of a small proportion of cells that still initiate XCI, in Rnf12+/- cells, also indicated that more XCI-activators are involved in XCI. Here, we propose to investigate the molecular mechanism by which RNF12 activates XCI in mouse and human, and to search for additional XCI-activators. We will also attempt to establish the role of different inhibitors of XCI, including CTCF and the pluripotency factors OCT4, SOX2 and NANOG. We anticipate that these studies will significantly advance our understanding of XCI mechanisms, which is highly relevant for a better insight in the manifestation of X-linked diseases that are affected by XCI.

1 500 000 €

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