Phenotypic landscape of intestinal organoid regeneration

Abstract

The development of intestinal organoids from single adult intestinal stem cells in vitro recapitulates the regenerative capacity of the intestinal epithelium 1 , 2 . Here we unravel the mechanisms that orchestrate both organoid formation and the regeneration of intestinal tissue, using an image-based screen to assay an annotated library of compounds. We generate multivariate feature profiles for hundreds of thousands of organoids to quantitatively describe their phenotypic landscape. We then use these phenotypic fingerprints to infer regulatory genetic interactions, establishing a new approach to the mapping of genetic interactions in an emergent system. This allows us to identify genes that regulate cell-fate transitions and maintain the balance between regeneration and homeostasis, unravelling previously unknown roles for several pathways, among them retinoic acid signalling. We then characterize a crucial role for retinoic acid nuclear receptors in controlling exit from the regenerative state and driving enterocyte differentiation. By combining quantitative imaging with RNA sequencing, we show the role of endogenous retinoic acid metabolism in initiating transcriptional programs that guide the cell-fate transitions of intestinal epithelium, and we identify an inhibitor of the retinoid X receptor that improves intestinal regeneration in vivo.

Human colorectal pre-cancer atlas identifies distinct molecular programs underlying two major subclasses of pre-malignant tumors

“…For SSCs, significantly represented subnetworks form a super module pointing towards interleukin signaling and microbiota interaction through receptors for retinoic acid (RA) (RXRA/RARA), vitamin D (VDR), and aryl hydrocarbon (AHR) (Figure 3A,G; Figure S9F-H). Importantly, a recent paper reported exogenous RA signaling suppressed the YAP-dependent regenerative stem state and promoted absorptive cell differentiation, consistent with the "absorptive" identity of SSCs (Lukonin et al, 2020). We furthered gleaned signature genes from organoids under RA signaling activation and found similar genes, such as ALDOB, to be upregulated in SSCs ( Figure 3A).…”

Establishment of intestinal organoid cultures modeling injury-associated epithelial regeneration

“…Recent reports demonstrated a critical role of RXR signaling in the regulation of the regeneration state in intestinal organoids. 20 Interestingly, compared to organoids cultured in the 8C minus VPA/EPZ6438 condition, Hyper-organoids showed a slightly decreased expression of target genes of RXR signaling (Supplementary information, Fig. S10g, h).…”

“…Although activation of regeneration is observed during the early stages of organoid formation from single stem cells, this regeneration process is transient which cannot persist during the growth of organoids. 20 In the gut, for example, regeneration of damaged intestinal epithelium differs significantly in cellular composition and stem cell function from normal intestinal epithelial homeostasis. 15,[21][22][23][24] Homeostasis of the intestinal epithelium primarily relies on active cycling of Lgr5 + crypt-base columnar cells (CBCs), which give rise to all the major intestinal epithelial cell types.…”

Regenerative Intestinal Stem Cells Induced by Acute and Chronic Injury: The Saving Grace of the Epithelium?

“…Importantly, they also showed that ISLR is overexpressed in human tissues from patients with IBD and colorectal cancers, suggesting that inflammation drives the expression of ISLR. Finally, Lukonin et al (2020) used a single cell imaging platform to identify a novel role of the orphan retinoid X receptor RXR in IEC proliferation and differentiation. They found that RXR activation using all-trans retinoic acid or a chemical activator led to maturation and differentiation of enteroids, associated with Yap cytosolic localization.…”