ONECUT2 regulates RANKL-dependent enterocyte and microfold cell differentiation in the small intestine; a multi-omics study

Abstract

Microfold (M) cells reside in the intestinal epithelium of Peyer's patches (PP). Their unique ability to take up and transport antigens from the intestinal lumen to the underlying lymphoid tissue is key in the regulation of the gut-associated immune response. Here, we applied a multi-omics approach to investigate the molecular mechanisms that drive M cell differentiation in mouse small intestinal organoids. We generated a comprehensive profile of chromatin accessibility changes and transcription factor dynamics during in vitro M cell differentiation, allowing us to uncover numerous cell type-specific regulatory elements and associated transcription factors. By using single-cell RNA sequencing, we identified an enterocyte and M cell precursor population. We used our newly developed computational tool SCEPIA to link precursor cell-specific gene expression to transcription factor motif activity in cis-regulatory elements, uncovering high expression of and motif activity for the transcription factor ONECUT2. Subsequent in vitro and in vivo perturbation experiments revealed that ONECUT2 acts downstream of the RANK/RANKL signalling axis to support enterocyte differentiation, thereby restricting M cell lineage specification. This study sheds new light on the mechanism regulating cell fate balance in the PP, and it provides a powerful blueprint for investigation of cell fate switches in the intestinal epithelium.

Control of Paneth cell function by HuR regulates gut mucosal growth by altering stem cell activity

“…Secretory cells in the epithelial compartment of the small intestine consisted of PCs, goblet cells, tuft cells, and enteroendocrine cells in both IE-HuR −/− and littermate mice, whereas absorptive cells were marked by various enterocytes (Fig 2A). Notably, PCs were marked by high expression of Defe5 and Rg3a mRNAs, whereas ISCs were marked by transcripts including Ascl2, Lgr5, Olfm4, Rgmb, and Smoc2 mRNAs, as reported previously (Elmentaite et al, 2021;Luna Velez et al, 2023). Cell cluster analysis revealed that the numbers of PCs and ISCs decreased in IE-HuR −/− mice compared with control littermate mice, which were consistent with the findings obtained from studies using immunostaining assays as shown in Fig 1.…”

Transcriptional Landscape of CUT-Class Homeobox Genes in Blastic Plasmacytoid Dendritic Cell Neoplasm

“…For example, CUX1 affects both myeloid and lymphoid differentiation, and CUX2 controls HOX gene expression during forelimb development [ 41 , 42 ]. ONECUT1 and ONECUT2 regulate early retinal cell fates, while the latter also regulates intestinal cell differentiation [ 43 , 44 ]. SATB1 supports self-renewal of HSCs inhibiting their differentiation and orchestrates erythroid development via regulation of globin genes [ 45 , 46 ].…”

“…BMP-signaling inhibits ONECUT2 in trigeminal sensory neuron differentiation [ 54 ], demonstrating that this pathway plays a physiological role in ONECUT2 regulation in some cell types. We failed to detect any regulatory impact for TNFRSF11A (also named RANK); although, this receptor has been shown to activate ONECUT2 in enterocytes of the small intestine [ 43 ]. In T-cells, ONECUT2 is regulated by TBX21/T-bet [ 55 ].…”

Organoids in gastrointestinal diseases: from bench to clinic

“…Moreover, they might be used to define how different microorganisms or patient‐specific microorganisms, even the whole microbiota of gut, regulate intestinal physiological processes and pathological progression (Figure 5B). The close‐knit integration with gene editing 233,234 and multiomics, 235,236 which will provide important insights into the mechanisms underlying microbial–host interactions, is also being expected (Figure 5C). These technologies will enable comprehensive and detailed detection of the modification or expression levels of various components, from DNA to RNA to protein, so as to clarify the functional changes after the interaction between intestinal cells and bacteria.…”