Relevance Malaria is one of the most threatening infectious diseases, placing almost half of the world-population at risk, and is responsible for nearly 1 million deaths annually. Despite intensive research, no effective vaccine is available and resistance is spreading against the current, and only available, line of medication. Therefore, new drug-targets and novel strategies to fight this disease are urgently required. Histone deacetylase (HDAC) inhibitors, successfully used in cancer-therapy, pose anti-malarial activity against in vitro human and in vivo rodent malaria infections. However, their exact targets and the molecular underpinning of their anti-malarial effect so-far remain completely unspecified, but are essential to improve therapeutic efficacy. Project HDACs are prime enzymes in epigenetic regulation of gene expression and genome-organization. In eukaryotes, they function in large multi-protein complexes, and when targeted to their sites-of-action orchestrate transcriptional repression via their various epigenetic activities. However, the composition of the presumed mega-dalton HDAC-complexes in malaria parasites is completely unknown. I aim to: 1. Characterize the HDAC-(sub)complexes in the deadly human malaria parasite Plasmodium falciparum utilising the latest proteomic approaches, 2. Identify the genome-wide localization of these various (sub)complexes using state-of-the-art next-generation-sequencing, 3. Identify chemical compounds that specifically inhibit the catalytic HDAC-activity or complex-recruitment employing high-tech chemoproteomics. By means of this combined approach of complex-characterization and ?epidrug? identification, I aim to uncover Plasmodium(-specific) epigenetic mechanisms and identify promising targets for rational-based drug design. Innovation This is a timely, ambitious and ground-breaking project, which will most likely result in: - Elucidation of the composition of HDAC-(sub)complexes - Genome-wide ChIP-seq localization, which I have pioneered for Plasmodium, of HDAC-complexes - Testing of candidate-epidrugs on physiologically relevant native, mega-dalton protein complexes (instead of physiologically less representative singular recombinant HDAC proteins) as a prelude to novel therapy - Adaptation of state-of-the-art proteomics & top-notch chemoproteomic approaches to malaria research.

During embryonic development in mammals, female cells silence one of the two X chromosomes in a process called X-chromosome inactivation. In this project, we have applied powerful new labelling strategies to identify 41 proteins that are involved in X inactivation. Future functional studies are required to further explore the exact functions of these proteins in the X inactivation process. Thus, our all-encompassing approach revealed for the first time all proteins and protein complexes that cooperate in the silencing of the X chromosome, and thereby this project has been very successful.

Stem cell therapy offers therapeutic opportunities for diseases such as leukemia and diabetes, during which disfunctioning or diseased cells are replaced with healthy cells. For these therapies, a promising stem cell source which does not require invasive procedures and/or elicit ethical debate are trophoblast cells retrieved from the placenta after birth. However, methods to efficiently reprogram these cells to pluripotent stem cells to enable directed differentiation to any cell type of interest are currently lacking. Therefore, we aim to identify small-molecule compounds that enable efficient reprogramming as critical step towards using placental cells for stem cell therapies.

A cell in the body is rarely alone and even when it is, it continuously receives signals from the environment. These signals are crucial for early development of the embryo, but also for the repair of tissues after damage or during disease. This project tries to understand how cells choose how to react to these signals and which processes in the cell influence this choice.

With approximately 1,400 known and predicted members,1 transcription factors (TFs) represent the largest class of human proteins.2 Obtaining a complete picture of the regulatory networks defined by the orchestrated binding of TFs in human cells would significantly increase our understanding of biological processes such as development and disease and provide invaluable knowledge for regenerative medicine.3 A comprehensive catalogue of DNA recognition motifs and their corresponding TFs would allow the construction of genome wide TF binding maps of all human tissues present in the large repositories of DNaseI footprinting data.4-6 However, due to the limited throughput of current technologies this catalogue so far only covers a minority of TFs and motifs. To bridge this gap, I aim to develop an all-in-one, unbiased, high-throughput compatible quantitative mass-spectrometry-based proteomics approach that allows the identification of protein-DNA interactions, including their affinity and the stoichiometry of their complexes. I will focus on novel recognition motifs discovered by DNaseI footprinting in human embryonic stem cells (hESC) and neuroectoderm, eventually allowing the comparison of their TF regulatory networks. Key objectives: 1. Development of a high-throughput compatible approach for the identification of Protein-DNA interactions including their Affinity and Complex stoichiometry (iPAC) 2. Identification of the TFs that bind to novel motifs discovered in DNase I footprints of hESCs and neuroectoderm 3. Generation of comprehensive TF binding maps for hESCs and neuroectoderm 4. Bioinformatic analysis of (cell-type specific) TF modules including their functional impact on pluripotency and neuronal development iPAC will be a generic tool to answer a plethora of biological questions centred around protein-nucleic acid interactions, including the functional effects of disease-linked single nucleotide polymorphisms.7 iPAC combined with available genomics data will thus be an extremely useful resource to model disease and identify factors of disease, and will therefore form a basis for the development of targeted medication.