Spatial Genome Architecture in Development & Disease

Functional Characterization and Heterogeneity of the DLBCL and AML 3D Genomes


Chromatin conformation constitutes a fundamental level of eukaryotic genome regulation. However, our ability to examine its biological function and role in disease has been hindered by the large amounts of starting material required to perform current experimental approaches, limiting the examination in primary diseased tissues. As a result, we currently lack an understanding of how chromatin conformation is affected in diseased cells. Recently, we have developed a low input Hi-C method suitable for the direct examination of chromatin architecture in diseased tissue from patients, and we have demonstrated its applicability in proof-of-principle examinations of chromatin conformation in B-cells from a diffuse large B-cell lymphoma (DLBCL) patient. These experiments have resulted in interesting and valuable observations such as: i) the unbiased characterisation of patientspecific structural variation across the genome; ii) the detection of large regions with loss of heterozygosity; and, iii) unexpectedly, a global trend for the appearance of new conformational features, such as TADs, in the disease sample. In this proposal, we now aim to build on these experimental and computational methods to discover the fundamental principles and functional consequence of chromatin organisation changes in blood malignancies. In Aim 1, we will produce genome-wide chromatin conformation maps for a small cohort of DLBCL and acute myeloid leukaemia (AML) patients and a set of healthy donors as a control. These datasets will allow us to characterise the chromatin conformation heterogeneity between these samples and to identify recurrent changes in chromatin organisation in disease. In Aim 2, we will develop a computational analysis framework for the systematic and statistical analysis of chromatin architecture differences between any set of samples. These novel methods will allow us to perform an unbiased ranking of the chromatin conformation differences between disease and control samples as well as to automatically identify structural variation events. In Aim 3, we will generate functional data examining the transcriptional state of these samples, their level of chromatin accessibility and the binding state of architectural binding proteins, such as CTCF. This will allow us to evaluate the functional role of changes in chromatin conformation across samples and to pinpoint the molecular mechanisms driving these. Overall, this integrated, multidisciplinary approach will result in a deep characterisation of the changes in chromatin organisation in these malignancies and their functional implication in gene expression. In addition, our results will provide fundamental insight into how and where aberrations in chromatin architecture occur in disease, which will be an invaluable asset in future to devise personalised therapeutic intervention strategies.