Regulation of MYC transcription in 3D : implications for tumor development

This thesis uncovers how chromatin organization conspires with nuclear architecture and environmental stimuli in order to regulate gene expression in disease and particular, in cancer. In Paper I, we have unraveled a mechanism of oncogenesis, previously unknown in humans, and widely known as gene gating. Specifically, we have shown that in human colon cancer cells (HCT116) the oncogenic super-enhancer (OSE) of MYC increases its expression levels post-transcriptionally, by tethering MYC to the nuclear pore complex (NPC). This process facilitates the export of MYC transcripts to the cytoplasm and enables them to escape the rapid decay taking place in the nucleus. This phenomenon does not seem to be present in the healthy counterparts of these cells, human colon epithelial cells (HCECs), indicating that this is a unique feature of cancer. Moreover, our findings show that this mechanism is mediated by AHCTF1 (also known as ELYS): a mobile nucleoporin, part of the NPC, that binds on chromatin. Finally, it is also regulated by the canonical WNT signaling pathway and the complex formation between TCF4 and β-catenin, as shown by the use of the inhibitor BC21.

In Paper II, we have further explored the molecular factors involved in the gating of MYC, as well as provided the first genetic evidence of this mechanism in humans. More precisely, by using CRISPR-Cas9 genetic engineering we generated two different clones with a mutated CTCF binding site (CTCFBS) within the OSE and showed that their inability to efficiently bind CTCF is associated with reduced MYC mRNA export. In addition, this process confers to the wild type cells a growth advantage over the mutant cells and requires the canonical WNT signaling pathway for the recruitment of the OSE from intra-nucleoplasmic positions. Our findings furthermore indicate that once the OSE has reached a peripheral position (<0.7um), the CTCFBS-mediated CCAT1 eRNA activation takes place and promotes the recruitment of AHCTF1 to the CTCFBS. That will ultimately lead to the efficient tethering of MYC to the nuclear pores and its subsequent gating, whilst pointing out the existence of a novel WNT/β-catenin-AHCTF1-CTCF-eRNA circuit in the regulation of pathogenic MYC expression.

In summary, the findings covered by the present thesis provide new insights in the regulation of oncogenic MYC expression by the 3D nuclear architecture and widen our understanding on the processes underlying tumor development. Such knowledge can improve the diagnosis, as well as potentially contribute to the identification of new therapeutic targets in cancer therapy.

List of scientific papers

I. WNT signaling and AHCTF1 promote oncogenic MYC expression through super-enhancer-mediated gene gating. Barbara A. Scholz#, Noriyuki Sumida#, Carolina Diettrich Mallet de Lima, Ilyas Chachoua, Mirco Martino, Ilias Tzelepis, Andrej Nikoshkov, Honglei Zhao, Rashid Mehmood, Emmanouil G. Sifakis, Deeksha Bhartiya, Anita Göndör* and Rolf Ohlsson*. Nature Genetics. 51, 1723–1731, 2019. #Shared first authors.
https://doi.org/10.1038/s41588-019-0535-3

II. Canonical WNT signaling-dependent gating of MYC requires a non-canonical CTCF function at a distal binding site. Ilyas Chachoua#, Ilias Tzelepis#, Hao Dai#, Jia Pei Lim#, Anna Lewandowska-Ronnegren#, Felipe Beccaria Casagrande, Shuangyang Wu, Johanna Vestlund, Carolina Diettrich Mallet de Lima, Deeksha Bhartiya, Barbara A Scholz, Mirco Martino, Rashid Mehmood and Anita Göndör*. Nature Communications, 13, 204, 2022. #Shared first authors.
https://doi.org/10.1038/s41467-021-27868-3