DNA methylation defines regional identity of human intestinal epithelial organoids and undergoes dynamic changes during development.
Nayak, Komal M
Howell, Kate J
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Kraiczy, J., Nayak, K. M., Howell, K. J., Ross, A., Forbester, J., Salvestrini, C., Mustata, R., et al. (2019). DNA methylation defines regional identity of human intestinal epithelial organoids and undergoes dynamic changes during development.. Gut, 68 (1), 49-61. https://doi.org/10.1136/gutjnl-2017-314817
Objective: Human intestinal epithelial organoids (IEO) are increasingly being recognised as highly promising translational research tool. However, our understanding of their epigenetic molecular characteristics and behaviour in culture remain limited. Design: We performed genome-wide DNA methylation and transcriptomic profiling of human IEO derived from paediatric/adult and foetal small and large bowel as well as matching purified human gut epithelium. Furthermore, organoids were subjected to in-vitro differentiation and genome editing using CRISPR/Cas9 technology. Results: We discovered stable epigenetic signatures which define regional differences in gut epithelial function, including induction of segment-specific genes during cellular differentiation. Established DNA methylation profiles were independent of cellular environment, since organoids retained their regional DNA methylation over prolonged culture periods. In contrast to paediatric and adult organoids, foetal gut -derived organoids showed distinct dynamic changes of DNA methylation and gene expression in culture, indicative of an in-vitro maturation. By applying CRISPR/Cas9 genome editing to foetal organoids, we demonstrate that this process is partly regulated by TET1, an enzyme involved in the DNA demethylation process. Lastly, generating IEO from a child diagnosed with gastric heterotopia revealed persistent and distinct disease-associated DNA methylation differences, highlighting the use of organoids as disease-specific research models. Conclusions: Our study demonstrates striking similarities of epigenetic signatures in mucosa-derived IEO with matching primary epithelium. Moreover, these results suggest that intestinal stem cell intrinsic DNA methylation patterns establish and maintain regional gut specification and are involved in early epithelial development and disease.
Intestinal Mucosa, Organoids, Cells, Cultured, Epithelial Cells, Humans, Cell Differentiation, DNA Methylation, Epigenesis, Genetic, Transcriptome, Clustered Regularly Interspaced Short Palindromic Repeats
This work was supported by funding from the following charitable organizations: Crohn’s in Childhood Research Association (CICRA), the Evelyn Trust, Crohn’s and Colitis in Childhood (“3Cs”), Addenbrooke’s Charitable Trust (ACT), and the Newlife Foundation for Disabled Children. J.K. was funded by a CICRA PhD studentship, K.H. was funded by an EBPOD EMBL-EBI/Cambridge Computational Biomedical Postdoctoral Fellowship. B-K.K. was supported by a Sir Henry Dale Fellowship from the Wellcome Trust and the Royal Society [101241/Z/13/Z] and received core support from the Wellcome Trust and MRC to the WT - MRC Cambridge Stem Cell Institute. GD and JF received support from the Wellcome Trust. J.F. was supported by a studentship from the MRC. P.R was supported by the Deutsche Forschungsgemeinschaft EXC306 Cluster “Inflammation at Interfaces” and BMBF IHEC DEEP TP5.2.
Wellcome Trust (101241/Z/13/Z)
Evelyn Trust (42228)
CICRA, Children with Crohn's and Colitis (unknown)
CICRA, Children with Crohn's and Colitis (RP/2014/1)
Evelyn Trust (15/38)
Newlife Foundation for Disabled Children (SG/15-16/05)
National Association for Colitis and Crohn's Disease (NACC) (M16-5)
WELLCOME TRUST (202483/Z/16/Z)
External DOI: https://doi.org/10.1136/gutjnl-2017-314817
This record's URL: https://www.repository.cam.ac.uk/handle/1810/276418
Attribution 4.0 International
Licence URL: http://creativecommons.org/licenses/by/4.0/
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