RESUMO
The intestinal epithelium is the fastest regenerative tissue in the body, fueled by fast-cycling stem cells. The number and identity of these dividing and migrating stem cells are maintained by a mosaic pattern at the base of the crypt. How the underlying regulatory scheme manages this dynamic stem cell niche is not entirely clear. We stimulated intestinal organoids with Notch ligands and inhibitors and discovered that intestinal stem cells employ a positive feedback mechanism via direct Notch binding to the second intron of the Notch1 gene. Inactivation of the positive feedback by CRISPR/Cas9 mutation of the binding sequence alters the mosaic stem cell niche pattern and hinders regeneration in organoids. Dynamical system analysis and agent-based multiscale stochastic modeling suggest that the positive feedback enhances the robustness of Notch-mediated niche patterning. This study highlights the importance of feedback mechanisms in spatiotemporal control of the stem cell niche.
Assuntos
Retroalimentação Fisiológica , Intestinos/citologia , Receptor Notch1/genética , Receptores Acoplados a Proteínas G/metabolismo , Animais , Sítios de Ligação , Autorrenovação Celular , Humanos , Mucosa Intestinal/metabolismo , Camundongos , Mutação , Organoides/metabolismo , Receptor Notch1/química , Transdução de Sinais , Nicho de Células-Tronco , Processos Estocásticos , Biologia de Sistemas/métodosRESUMO
Notch signalling maintains stem cell regeneration at the mouse intestinal crypt base and balances the absorptive and secretory lineages in the upper crypt and villus. Here we report the role of Fringe family of glycosyltransferases in modulating Notch activity in the two compartments. At the crypt base, RFNG is enriched in the Paneth cells and increases cell surface expression of DLL1 and DLL4. This promotes Notch activity in the neighbouring Lgr5+ stem cells assisting their self-renewal. Expressed by various secretory cells in the upper crypt and villus, LFNG promotes DLL surface expression and suppresses the secretory lineage . Hence, in the intestinal epithelium, Fringes are present in the ligand-presenting 'sender' secretory cells and promote Notch activity in the neighbouring 'receiver' cells. Fringes thereby provide for targeted modulation of Notch activity and thus the cell fate in the stem cell zone, or the upper crypt and villus.
Assuntos
Homeostase , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Intestinos/fisiologia , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas de Membrana/metabolismo , Receptores Notch/metabolismo , Células-Tronco/citologia , Proteínas Adaptadoras de Transdução de Sinal , Animais , Proteínas de Ligação ao Cálcio , Diferenciação Celular , Proliferação de Células , Células Cultivadas , Glucosiltransferases , Glicosiltransferases , Peptídeos e Proteínas de Sinalização Intercelular/genética , Intestinos/citologia , Peptídeos e Proteínas de Sinalização Intracelular/genética , Proteínas de Membrana/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas/fisiologia , Receptores Acoplados a Proteínas G/fisiologia , Receptores Notch/genética , Transdução de Sinais , Células-Tronco/metabolismoRESUMO
Emerging evidence suggests that microRNAs can initiate asymmetric division, but whether microRNA and protein cell fate determinants coordinate with each other remains unclear. Here, we show that miR-34a directly suppresses Numb in early-stage colon cancer stem cells (CCSCs), forming an incoherent feedforward loop (IFFL) targeting Notch to separate stem and non-stem cell fates robustly. Perturbation of the IFFL leads to a new intermediate cell population with plastic and ambiguous identity. Lgr5+ mouse intestinal/colon stem cells (ISCs) predominantly undergo symmetric division but turn on asymmetric division to curb the number of ISCs when proinflammatory response causes excessive proliferation. Deletion of miR-34a inhibits asymmetric division and exacerbates Lgr5+ ISC proliferation under such stress. Collectively, our data indicate that microRNA and protein cell fate determinants coordinate to enhance robustness of cell fate decision, and they provide a safeguard mechanism against stem cell proliferation induced by inflammation or oncogenic mutation.