Gene expression is further determined by the chromatin landscape. Specific transcription factors, such as ATOH1, are critically required for acquisition of IEC effector lineage identities ( Yang et al., 2001). The balance between ISC proliferation and IEC differentiation is controlled by pathways including WNT, BMP, and NOTCH ( Gehart and Clevers, 2019). On the other hand, it is vulnerable to tumorigenesis with colorectal cancer being the second leading cause of cancer-related deaths worldwide. The intestinal epithelium exhibits high plasticity in respond to challenges ( Haber et al., 2017 Jadhav et al., 2017). ISCs are continuously dividing and give rise to progenitor cells, which differentiate into specialized intestinal epithelial cell (IEC) lineages such as absorptive enterocytes and secretory lineages such as mucus-producing goblet cells, antimicrobial-producing Paneth cells, hormone-secreting enteroendocrine cells, and chemosensory tuft cells ( Gehart and Clevers, 2019). One of the hallmarks of intestinal epithelium is the rapid turnover of 3–5 days, which is driven by LGR5+ intestinal stem cells (ISCs) that reside at the bottom of crypts. The intestinal epithelium, a single layer of cells, faces the challenge of both providing a barrier against pathogens while also being responsible for the uptake of nutrients and water. Thus, epigenetic probes are powerful tools to study intestinal epithelial biology and may have therapeutic potential. For example, we show that inhibiting type I PRMTs, which leads enhanced epithelial differentiation, blocks the growth of adenoma but not normal organoid cultures. Based on this resource dataset, we identified several targets that affected epithelial cell differentiation, including HDACs, EP300/CREBBP, LSD1, and type I PRMTs, which were verified by complementary methods. The role of epigenetic modifiers in adult stem cell systems, such as the intestinal epithelium, is still undefined. Here, we developed a semi-automated organoid screening method, which we applied to a library of highly specific chemical probes to identify epigenetic regulators of intestinal epithelial biology. Yet, the selection of analytical tools to accurately quantify heterogeneous organoid cultures remains limited. Intestinal organoids are an excellent model to study epithelial biology. 9Department of Medical Biophysics, University of Toronto, Toronto, ON, Canada.8Princess Margaret Cancer Centre, University Health Network, Toronto, ON, Canada.7Structural Genomics Consortium, University of Toronto, Toronto, ON, Canada.6Screening Unit, Leibniz-Forschungsinstitut für Molekulare Pharmakologie, Berlin, Germany.5Department of Bioinformatics, Computational Biological Unit, University of Bergen, Bergen, Norway.

4Cancer Biology Department, Institute of Functional Genomics, University of Montpellier, Montpellier, France.3Novo Nordisk Foundation Center for Stem Cell Biology, Faculty of Health and Medical Sciences, University of Copenhagen, Copenhagen, Denmark.2BRIC - Biotech Research and Innovation Centre, University of Copenhagen, Copenhagen, Denmark.1Centre of Molecular Inflammation Research (CEMIR), Department of Clinical and Molecular Medicine (IKOM), NTNU - Norwegian University of Science and Technology, Trondheim, Norway.Jensen 2,3, Cheryl Arrowsmith 7,8,9 and Menno J. Lindholm 1, Alberto Díez-Sánchez 1, Naveen Parmar 1, Silke Radetzki 6, Jens Peter von Kries 6, Philippe Jay 4, Kim B. Zwiggelaar 1, Marianne Terndrup Pedersen 2,3, François Gerbe 4, Korbinian Bösl 5, Håvard T. Run('Set Measurements.Jenny Ostrop 1 *, Rosalie T. Click and hold to erase noise in the black and white image. WaitForUser('ERASE NOISE', 'Bring up the original image to compare where is true signal. Run('Analyze Particles.', 'size=0.1-Infinity show=Masks display clear') WaitForUser('REMEMBER MEAN VALUE', 'Remember/Copy the 'mean' value. Run('Set Measurements.', 'area mean limit redirect=None decimal=3') WaitForUser('MOVE THE SQUARE', 'Please move the square to where you think is noise/background. Run('Colour Deconvolution', 'vectors=H&E hide') WaitForUser('Image selection', 'Please draw to select area you want to keep for analysis If you need to adjust, finish drawing first.

WaitForUser('Image selection', 'Please zoom in (+) to where the scale bar is and then draw a line over the scale bar.

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