We examined the interactions between three activated transcription pathways.
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All three pathways distinctly influenced intestinal stem cells.
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Fully functional Myb was required for Wnt and Notch activities.
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Notch inhibition selectively favors survival of Wnt-activated cyst-like organoids.
Abstract
Deletion studies confirm Wnt, Notch and Myb transcriptional pathway engagement in intestinal tumorigenesis. Nevertheless, their contrasting and combined roles when activated have not been elucidated. This is important as these pathways are not ablated but rather are aberrantly activated during carcinogenesis. Using ApcMin/+ mice as a source of organoids we documented their transition, on a clone-by-clone basis, to cyst-like spheres with constitutively activated Wnt pathway, increased self-renewal and growth and reduced differentiation. We then looked at this transition when Myb and/or Notch1 are activated. Activated Notch promoted cyst-like organoids. Conversely growth and propagation of cyst-like, but not normal organoids were Notch-independent. Activated Myb promoted normal, but not cyst-like organoids. Interestingly the Wnt, Notch and Myb pathways were all involved in regulating the expression of the intestinal stem cell (ISC) gene Lgr5 in organoids, while ISC gene and Notch target Olfm4 was dominantly repressed by Wnt. These findings parallel mouse intestinal adenoma formation where Notch promoted the initiation, but not growth, of Wnt-driven Olfm4-repressed colon tumors. Also Myb was essential for colon tumor initiation and collateral mouse pathologies. These data reveal the complex interplay and hierarchy of transcriptional networks that operate in ISCs and uncover a shift in pathway-dependencies during tumor initiation.
Graphical abstract
Proposed model to depict the interaction-networks linking activated Wnt, Notch and Myb transcription factors in intestinal crypts. Genes that reside between spheres of control are suggested to be co-regulated while genes within the colored spheres are primarily regulated by that pathway. β-Catenin is shown as it is a core mediator of the Wnt canonical signaling pathway while CSL is noted as it is an activated NotchIC transcription partner.
