The small intestinal epithelium self-renews every four or five days. Intestinal stem cells (Lgr5 crypt base columnar cells (CBCs)) sustain this renewal and reside between terminally differentiated Paneth cells at the bottom of the intestinal crypt. Whereas the signalling requirements for maintaining stem cell function and crypt homeostasis have been well studied, little is known about how metabolism contributes to epithelial homeostasis. Here we show that freshly isolated Lgr5 CBCs and Paneth cells from the mouse small intestine display different metabolic programs. Compared to Paneth cells, Lgr5 CBCs display high mitochondrial activity. Inhibition of mitochondrial activity in Lgr5 CBCs or inhibition of glycolysis in Paneth cells strongly affects stem cell function, as indicated by impaired organoid formation. In addition, Paneth cells support stem cell function by providing lactate to sustain the enhanced mitochondrial oxidative phosphorylation in the Lgr5 CBCs. Mechanistically, we show that oxidative phosphorylation stimulates p38 MAPK activation by mitochondrial reactive oxygen species signalling, thereby establishing the mature crypt phenotype. Together, our results reveal a critical role for the metabolic identity of Lgr5 CBCs and Paneth cells in supporting optimal stem cell function, and we identify mitochondria and reactive oxygen species signalling as a driving force of cellular differentiation.

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.

In Saccharomyces cerevisiae, the forkhead transcription factor Hcm1 is involved in chromosome segregation, spindle pole dynamics, and budding. We found that Hcm1 interacts with the histone deacetylase Sir2 and shifts from cytoplasm to the nucleus in the G 1 /S phase or in response to oxidative stress stimuli. The nuclear localization of Hcm1 depends on the activity of Sir2 as revealed by activators and inhibitors of the sirtuins and the ⌬sir2 mutant. Hcm1-overexpressing cells display more mitochondria that can be attributed to increased amounts of Abf2, a protein involved in mitochondrial biogenesis. These cells also show higher rates of oxygen consumption and improved resistance to oxidative stress that would be explained by increased catalase and Sod2 activities and molecular chaperones such as Hsp26, Hsp30, and members of Hsp70 family. Microarray analyses also reveal increased expression of genes involved in mitochondrial energy pathways and those allowing the transition from the exponential to the stationary phase. Taken together, these results describe a new and relevant role of Hcm1 for mitochondrial functions, suggesting that this transcription factor would participate in the adaptation of cells from fermentative to respiratory metabolism.Mammalian FoxO transcription factors (FoxO1, 3, 4, and 6), a subfamily of forkhead transcription factors (FKH-TFs), 3 control various biological functions, including stress resistance, DNA repair, metabolism, cell cycle arrest, and apoptosis (reviewed in Refs. 1-3). According to this variety of functions, FoxO transcription factors are regulated in these diverse functions by a wide range of external stimuli, such as insulin, insulin-like growth factor, other growth factors, neurotrophins, nutrients, cytokines, and oxidative stress stimuli. These stimuli control FoxO protein levels, subcellular localization, DNA binding, and transcriptional activity. FoxO are subject to several post-translational modifications including phosphorylation, acetylation, and ubiquitination (1, 4