Structural and Mechanistic Studies of the SWI/SNF Family of Chromatin Remodelling Complexes

DNA, the blueprint of life, is found within 46 chromosomes in every human cell; if stretched out end to end, it would measure two metres in length. In order for these 46 chromosomes to fit into the "control-room" of every cell, known as the nucleus, the DNA must be very tightly packaged. This is achieved by wrapping the DNA around the surface of special proteins, called histones, that are spaced regularly along the DNA like beads on a string. Each DNA-histone bead is known as a nucleosome and each nucleosome is able to pack very closely against its neighbours to form highly compact fibres called chromatin. Although chromatin fibers are very good at compacting DNA into small spaces, they are poor at allowing other proteins access to the DNA. Many normal processes in the cell involve proteins binding to DNA, such as when genes are decoded to make protein, when chromosomes are replicated prior to cell division, and when special repair proteins are called upon to fix sites of DNA damage. Consequently, complicated organisms like humans, with highly packaged DNA, have had to develop specialized machinery for opening chromatin at very specific regions of the chromosome. This machinery includes a large number of specialized proteins. One group of proteins, known as chromatin remodelling complexes (CRCs), help to expose DNA by using energy to slide or remove nucleosomes within chromatin. CRCs are protein machines, much larger than a single nucleosome, and made up of proteins responsible for recruitment to the correct chromosome region, binding to nucleosomes, or helping to generate the force required for nucleosome sliding or removal. Multiple different CRCs exist in human cells, each made up of similar proteins that can interact with chromatin in subtly different ways or target the complex to different regions of the chromosome. The recruitment of a CRC to the start of a gene is an important early step in the process of turning on, or activating, that gene. If the cell makes a mistake, failing to recruit CRCs to their correct sites or instead recruiting those complexes to the wrong set of genes, then a dangerous cascade can result in the loss of control of normal cell events such as cell division and death. This type of gene dysregulation takes place at an early stage in the development of every human cancer. Over the past decade, advances in DNA sequencing technology have allowed scientists to identify mistakes in the DNA code, known as DNA mutations, that are found in many types of human cancer. A surprising finding from these studies was that DNA mutations leading to the loss of protein components from one single CRC, known as the BAF complex, are present in as many as 20% of all human cancers. Further investigations showed that DNA mutations altering the BAF complex cause many normal target genes to be switched off whereas new and inappropriate genes often become active. Although the BAF complex is very often the target of mutations in cancer, surprisingly little is currently known about this important machine, with many questions still unaddressed. For example, how are the different proteins organized in the BAF complex? What roles do the different proteins play in the recruitment of the complex to the correct target genes? How does the BAF complex interact with nucleosomes? How does BAF use energy to bring about nucleosome sliding or eviction? The overarching goal of my future research will be to address these questions using a repertoire of cutting-edge structural biology techniques such as cryo-electron microscopy, protein cross-linking, X-ray crystallography and computational modelling, in order to provide a detailed description of the organization, recruitment and remodelling activity of this important human complex. Such findings will provide a framework for understanding the molecular basis of BAF complex dysregulation, with broad implications for the future treatment of many human cancers.