Defining the chromatin structure of the human genome using size-selected nucleosome mapping

The work in this thesis examines genome-wide and local changes in the patterns of nucleosome positioning throughout the human genome. Nucleosomes are the fundamental repeating unit of chromatin. Their properties and positioning in the genome dictate whether and how proteins involved in gene regulation can access DNA. Nucleosomes are dynamic; their positions can vary considerably at some loci from one cell type to another. Chromatin remodelling complexes can change the structure and the positions of nucleosomes. Their mis-regulation leads to congenital defects affecting pre-natal and early childhood development and is associated with neuro-psychiatric disorders. As mutations in genes that encode chromatin remodelling proteins are associated with human mental health disorders, the work in this thesis focusses on changes that occur in chromatin structure during early neural development. I have used MNase-seq data to construct genome-wide, high-resolution chromatin particle positioning maps from undifferentiated human induced pluripotent stem cells (hiPSC) and following differentiation to the neuro-progenitor cell (NPC) stage. These maps reveal that a small proportion of the pluripotent genome possesses well-positioned nucleosomes, the number of which increases approximately 8-fold during neural cell development. This is accompanied by changes in the distribution and localisation of nucleosomes between iPS and NPC cells. Differences in nucleosome positioning during neural cell differentiation were investigated at regulatory regions. Loss and gain of positioned nucleosomes at TSS of pluripotent and neural-specific genes was detected and correlated with gene expression. In addition I investigated the chromatin structure at the binding motifs of two important genome regulators REST and CTCF in detail. Nucleosome positioning is maintained at REST binding motifs during neural cell development. In contrast, at CTCF sites nucleosome repositioning occurs during neural cell development. This work provides insight into the role of chromatin structure in the regulation of human neural cell differentiation.