Crosstalk between environmental signals and 3d genome organization in the regulation of gene expression

Abstract

The thesis explores the connection between environmental stimuli and gene expression regulated by the spatial changes in genome organization. In Paper I, by applying state of the art Circular Chromosome Conformation Capture assay (4C) and Chromatin in situ Proximity (ChrISP) techniques, we show that transcriptionally active circadian genes meet in space with repressed lamina-associated domains (LADs), and that these interactions are under the control of the circadian clock. External time cues thus synchronised circadian transcriptional oscillations by repositioning clock-controlled genes from the transcriptionally permissive sub-compartment of nuclear interior to the transcriptionally repressive nuclear periphery. These processes relied on the rhythmic formation of complexes between CTCF and PARP1, two master regulators of the genome, to increase the amplitude of circadian gene expression. In Paper II we took an advantage of the novel, ultrasensitive Nodewalk technique to explore the stochastic nature of MYC interactions with its flanking enhancers. By pushing the Nodewalk limits of identification of chromatin interactions in the input material corresponding to less than 8 cells, we could show that MYC is likely screening for neighbouring interaction partners rather than vice versa. Moreover, we could show that MYC does not interact with enhancers, once its transcription had been initiated. These findings suggest that enhancer hubs simultaneously interacting with MYC are likely virtual consequences of high cell population analyses and that MYC interacts with its enhancers in a mutually exclusive manner. Paper III concentrates on the role of a CTCF binding site within the oncogenic super- enhancer (OSE) in the regulation of MYC gene gating in colon cancer cells. CRISPR induced mutations in the CTCF binding site within the OSE abrogated WNT-dependent nuclear export of MYC mRNA, providing genetic evidence to the claim of the OSE- mediated gating of active MYC alleles to the nuclear pore.This manuscript documents, moreover, that the communication between OSE and MYC, as well as their repositioning to the nuclear pore, involves PARP1 to indicate a more general role for the CTCF :PARP1 complex in gene regulation.

In summary, this thesis has uncovered novel principles underlying the roles of stochastic chromatin interactions and mobility within the 3D nuclear space to regulate gene expression with a focus on circadian transcriptional regulation and the recently discovered gene gating phenomenon in humans. These findings contribute to our understanding of principles in which the nuclear architecture and genome organisation synergize to induce or maintain the properties of the cell. By extrapolation, such findings might form a platform for identifying new therapeutic strategies to battle cancer, for example.