Unravelling the metaphase epigenome: Differences in DNA accessibility between homologous chromosomes

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My research focuses on understanding the structure and organization of metaphase chromosomes in mitosis. High fidelity mitotic metaphase chromosome condensation is essential for accurate transmission of the genome into daughter cells. Despite advances in modeling higher order chromosome condensation, locus-specific accessibility of chromatin within highly condensed metaphase chromosomes is not well understood. Furthermore, it is believed that the pattern of condensation along the chromosome's length differs between homologous chromosome pairs but that individual chromosomes in a pair share the same pattern. Our recent studies using 3D super-resolution microscopy and unique sequence DNA FISH probes developed from our genomic technology demonstrate that this is not the case (Khan et al, Mol Cytogen 7:70, 2014). By color labeling short DNA sequences from parts of specific genes or intergenic regions and tagging them to their complementary sequence on the chromosome pair using fluorescence in situ hybridization (FISH), we have identified regions of individual chromosome pairs that differ from each other. These differences do not occur on all chromosome regions but occur on most if not all chromosomes. These differences are nonrandom, reproducible and share the same pattern on chromosomes from many unrelated individuals. The DNA in DA regions is more compacted on one member of the chromosome pair which is more open (referred to as differential accessibility or DA). We have also been able to modify metaphase chromosome structure with a chromatin condensation inhibitor to remove these structural differences by relaxing the chromosome (Khan et al, Mol Cytogen, 8:65, 2015). We have studied 14 DNA probes (9 that show differential accessibility [DA] and 5 controls that do not) on metaphase chromosomes from blood tissue. These findings have raised many questions. In this research program, we want to understand the significance of DA by broadening our studies to include: A) whether DA occurs in other tissues and is identical between tissues; B) whether it is heritable and related to parental origin; C) whether studying a larger number of loci will reveal common sequence-based or structural properties of DA regions; D) if DA is widespread DA in the genome; E) whether the super-helical differences that characterize DA arise directly or indirectly from topoisomerase binding to DNA; F) whether DA is established during development or differentiation; and G) if it is an early event that remains stable once it occurs. DA may represent a structural code of chromatin regulation, and its occurrence on homologous metaphase chromosomes may be a way for the cell to re-establish identity after mitosis. We propose a laboratory based program that addresses each of these questions.