The physical basis of all life is the ability to store, selectively read out, duplicate and transmit genetic information. In order to achieve these goals the DNA that carries this information within each living cell is organized spatially by proteins into a highly intricate, hierarchical and dynamical structure known as chromatin (Figure 1). This amazing structure strikes a balance between the need for compaction and the need for accessibility by genomic processes (i.e. transcription, replication, repair and recombination of DNA). Understanding the constant (re)organization of the genome within the chromatin structure is needed to grasp the physical basis of life in all eukaryotes, including man. Despite this, there is a remarkable paucity in our understanding of properties and behavior of chromatin.

Hence we have started a research program with the objective to resolve the physical mechanisms that organize, regulate and maintain the DNA within the chromatin structure.

The timing of such an ambitious project is highly opportune, because - despite the progress biologists have made in identifying the many key players that interact with the genome – to date little is known about the physical processes that underpin the dynamic control of the DNA in the chromatin structure. Fortunately, over the past few years biophysicists have developed highly sophisticated single-molecule techniques that are ideally suited for investigations of the biophysical mechanisms of the protein machines that arrange and process the chromatin structure. The research groups united in this consortium have played a prominent role in the international research on this topic and combine a unique array of know-how needed to address this important problem. A major challenge for biophysical research in the coming years is to attain a higher level of complexity and study the interplay of different proteins in complex mechanisms. This research program will bundle and focus the existing excellence in nucleoprotein biophysics in the Netherlands and combine it with the expertise provided by selected eminent theoreticians and biologists to make this step and help resolve the physical mechanisms that organize, regulate and maintain the genome within the dynamic chromatin structure. Because malfunction of chromatin plays an important, but still poorly understood role in human health, including aging and cancer, we anticipate that results of our research will find applications in the biomedical and biotechnological field.

Background

Chromatin is probably the largest and most complex molecular structure existing inside cells. It can locally exist in a variety of functional states that are constantly switched in response to internal and external signals. Only in certain of these functional states can the DNA be read out or maintained. Unraveling the physical mechanisms of DNA processing and organization in chromatin requires concerted research efforts that span all length scales, from the interaction of individual proteins with a small number of base pairs (nanometer-scale) to the dynamics of full chromosome structures (micron-scale) and it poses the major challenge of functionally integrating the knowledge at these different length scales (figure 1).

Approach

Our objective of resolving physical mechanisms underlying chromatin dynamics and organization can be subdivided into three major areas: Chromatin structure (i.e. nucleosome binding & modification, domains of euchromatin and heterochromatin, ATP-dependent remodeling), DNA maintenance (i.e. repair, replication, recombination), and Gene expression (i.e. gene regulation, transcription), which are all tightly connected. In all of these cases the interaction of DNA with binding proteins such as the nucleosome, the protein complex that underlies the highly condensed structure of chromatin, plays a central role. However, at present each of these three domains developed into a separate field, complete with its own jargon, tools and selection of prototypical biochemical pathways. Rather than approaching these issues separately, we propose an integrated effort and focus on the interplay between structure, maintenance and expression. For example, how is recombination affected by DNA condensation, how do different nucleosome modifications affect transcription, and what role do regulatory networks play in transcription activity?

More specifically, we aim to elucidate and model the physical mechanisms and time frames of the following issues:

• How is the chromatin structure organized and dynamically remodeled, at the single-protein level in nucleosome-DNA interaction dynamics up to the different states of large-scale multi-component metaphase chromosomes.
• How is DNA maintained in chromatin, an investigation that involves studying the replication processes at multiple scales of complexity as well the physical mechanism of repair and homologous recombination of DNA.
• How is gene expression regulated and used within chromatin; a question that includes elucidating the mechanical aspects of the transcription machinery, DNA looping, and gene regulation in relation to the spatial organization of chromatin.

Over the past few years a number of groups working on the physics of DNA have emerged in the Netherlands, in large part due to seeding support from FOM. At the same time the importance of the mechanistic, physics-based approach to understanding the genome has become widely recognized in the international bioscience community. The proposed program brings together a number of highly successful groups in this field with a strong biophysics infrastructure, with a selected number of key biological partners, in order to establish a coherent and focused research framework. This set-up aims to optimally exploit the opportunities to embed the physics-based effort in the area of DNA within the increasingly multidisciplinary approach to understand the living cell at the systems level; an approach widely recognized as a crucial ingredient of future advances in biomedicine and biotechnology. The quality of biomolecular DNA research in the Netherlands is already recognized internationally, therefore the proposed research program intends to provide a platform of excellence with high-visibility which can be a major player in the increasingly large-scale consortia of the systems biology arena.