Integrative multi‐omics analysis of intestinal organoid differentiation

Abstract

Intestinal organoids accurately recapitulate epithelial homeostasis in vivo, thereby representing a powerful in vitro system to investigate lineage specification and cellular differentiation. Here, we applied a multi‐omics framework on stem cell‐enriched and stem cell‐depleted mouse intestinal organoids to obtain a holistic view of the molecular mechanisms that drive differential gene expression during adult intestinal stem cell differentiation. Our data revealed a global rewiring of the transcriptome and proteome between intestinal stem cells and enterocytes, with the majority of dynamic protein expression being transcription‐driven. Integrating absolute mRNA and protein copy numbers revealed post‐transcriptional regulation of gene expression. Probing the epigenetic landscape identified a large number of cell‐type‐specific regulatory elements, which revealed Hnf4g as a major driver of enterocyte differentiation. In summary, by applying an integrative systems biology approach, we uncovered multiple layers of gene expression regulation, which contribute to lineage specification and plasticity of the mouse small intestinal epithelium.

Germ‐free and microbiota‐associated mice yield small intestinal epithelial organoids with equivalent and robust transcriptome/proteome expression phenotypes

“…Our work validates that murine small intestinal organoids are an experimental system with a robust expression pattern that resembles the phenotype of the homeostatic intestinal epithelium (Janeckova et al, ; Lindeboom et al, ). Importantly, our work extends previous data by demonstrating that this phenotype is largely independent from exposure with SPF microbiota.…”

“…The observed culture‐to‐culture variation in expression profiles may stem from bottleneck effects during early organoid establishment and/or adaptation to the culture conditions. In addition, the differentiation state and cell type composition of organoids is highly sensitive to the concentration of growth factors provided in the culture medium (e.g., Noggin, R‐spondin and EGF), or produced by the organoids themselves (e.g., Wnt3a; Farin et al, ; Kim et al, ; Lehmann et al, ; Lindeboom et al, ; Sato et al, ; van der Flier & Clevers, ). Fluctuation of these, often unstable, proteinaceous factors provides another plausible source of culture‐to‐culture variability.…”

Intestinal Organoids: A Tool for Modelling Diet–Microbiome–Host Interactions

“…A multi-omics approach was recently used to obtain a holistic view of molecular mechanisms in mouse IOs cultured in different defined mediums [100]. The research by Lindeboom and co-authors resulted in a novel workflow to investigate metabolites and lipids of IOs and can be used to explain the potential of lipidomics and metabolomics approaches aim at studying chemical signatures on IOs for diet-microbiome-host interactions.…”

Diabetes through a 3D lens: organoid models

“…A further key driver of their utility, in addition to being a more physiologically relevant model, is their amenability to both standard and high-tech laboratory techniques and their genetic and molecular tractability. For example, organoids can easily be manipulated using viral and nonviral mediated delivery of CRISPR-Cas9 gene editing [22], they can be investigated using mass spectrometry [23], flow cytometry [24], multiple single cell 'omics' technologies [25] and naturally their 3D structure lends themselves to all manner of imaging technologies [26] (Fig. 1).…”