TNFAIP8 controls murine intestinal stem cell homeostasis and regeneration by regulating microbiome-induced Akt signaling

ARTICLE https://doi.org/10.1038/s41467-020-16379-2 OPEN 1234567890():,; TNFAIP8 controls murine intestinal stem cell homeostasis and regeneration by regulating microbiome-induced Akt signaling Jason R. Goldsmith 1 ✉, Nina Spitofsky1, Ali Zamani1, Ryan Hood1, Amanda Boggs1, Xinyuan Li1, Mingyue Li1, Elizabeth Reiner1,2, Arshad Ayyaz3, Zienab Etwebi1, Ling Lu1, Javier Rivera Guzman1,4, Mayassa J. Bou-Dargham 1, Terry Cathoupolis1, Hakon Hakonarson 5,6, Honghong Sun1, Jeffrey L. Wrana3,7, Michael V. Gonzalez5,6 & Youhai H. Chen1 ✉ The intestine is a highly dynamic environment that requires tight control of the various inputs to maintain homeostasis and allow for proper responses to injury. It was recently found that the stem cell niche and epithelium is regenerated after injury by de-differentiated adult cells, through a process that gives rise to Sca1+ fetal-like cells and is driven by a transient population of Clu+ revival stem cells (revSCs). However, the molecular mechanisms that regulate this dynamic process have not been fully defined. Here we show that TNFAIP8 (also known as TIPE0) is a regulator of intestinal homeostasis that is vital for proper regeneration. TIPE0 functions through inhibiting basal Akt activation by the commensal microbiota via modulating membrane phospholipid abundance. Loss of TIPE0 in mice results in injuryresistant enterocytes, that are hyperproliferative, yet have regenerative deficits and are shifted towards a de-differentiated state. Tipe0−/− enterocytes show basal induction of the Clu+ regenerative program and a fetal gene expression signature marked by Sca1, but upon injury are unable to generate Sca-1+/Clu+ revSCs and could not regenerate the epithelium. This work demonstrates the role of TIPE0 in regulating the dynamic signaling that determines the injury response and enables intestinal epithelial cell regenerative plasticity. 1 Department of Pathology and Laboratory Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA. 2 University of Pikeville—Kentucky School of Osteopathic Medicine, Pikeville, KY 41501, USA. 3 Centre for Systems Biology, Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, Toronto, ON, Canada. 4 Department of Biology, University of Maryland Baltimore College, Baltimore, MD 21250, USA. 5 Center for Applied Genomics, The Children′s Hospital of Philadelphia, Philadelphia, PA 19104, USA. 6 Department of Pediatrics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA. 7 Department of Molecular Genetics, University of Toronto, Toronto, ON, Canada. ✉email: ; NATURE COMMUNICATIONS | (2020)11:2591 | https://doi.org/10.1038/s41467-020-16379-2 | www.nature.com/naturecommunications 1 ARTICLE NATURE COMMUNICATIONS | https://doi.org/10.1038/s41467-020-16379-2 I t was recently found that the stem cell niche and epithelium is regenerated after injury by de-differentiated adult cells1, through a process that gives rise to Sca1+ fetal-like cells and is driven by a transient population of Clu+ revival stem cells (revSCs)2–4. The intestinal epithelial cells (IECs) that comprise the intestinal barrier protect the body from harmful luminal contents and play critical roles in nutrient absorption and waste excretion5. In a healthy gut, IECs are regenerated about every 5 days. During and after intestinal injury, such as through radiation, colitis, or ischemia, intestinal regeneration becomes more pronounced as the disrupted barrier is restored. Lgr5+/CBC (crypt basal columnar) stem cells were previously identified as responsible for daily homeostatic regeneration6; however, they are particularly susceptible to cell death during injury, and yet the intestine is still able to regenerate after injury. This has led to ongoing efforts to understand the molecular biology of injuryinduced intestine regeneration6. Recently, it has been appreciated that all partially and fully differentiated epithelial cells studied thus far can revert to novel stem-cell states post injury and restore the homeostatic stem cell niche; a process named paligenosis6,7. A population of intestinal stem cells, known as “fetal-like” stem cells because of their similarity to fetal intestinal stem cells8, has been identified in adult mice that are Sca-1+ and Lgr5−, and only appear in appreciable numbers after injury3,4. This fetal-like, Sca-1+ stemcell program appears to require functional YAP/TAZ-signaling9, which is known to be repressed by β-catenin signaling, the central pathway mediating intestinal stemness, regeneration, and differentiation6. Other recent work demonstrated that this regenerative program requires the transient induction of a small subset of Clu + “revival stem cells” or rev-SCs (that are also Sca-1+), that are directly responsible for post-injury regeneration2. Thus, in the intestine, regeneration requires Sca-1+ and Clu+ cell states. However, the general signaling processes and critical proteins required to induce regenerative paligenosis after injury remain unknown. The TNFAIP8-like (Tumor necrosis factor-alpha-induced protein 8-like, or TIPE) family of proteins (comprised of TNFAIP8 and TNFAIP8L1, TNFAIP8L2, and TNFAIP8L3) are homologous lipid transport proteins that help regulate PI3Kmediated signaling, and play crucial roles in inflammation, cell migration, and cell survival10. TNFAIP8L2, (aka TIPE2), the most well-studied member, is restricted to the immune compartment. Loss of TIPE2 results in defective leukocyte chemotaxis11–13, and decreased dextran sodium sulfate (DSS)-induced colitis, with reduced immune cell migration to the gut14. In contrast, loss of TNFAIP8 (hereafter TIPE0), which is ubiquitously expressed11,12, results in more severe DSS colitis; this effect was found to be independent of immune cells15. We report here that TIPE0 is a regulator of the intestinal injury response and controls intestinal cell stemness and plasticity during injury, by regulating basal Akt activation induced by the microbiota. Results Tipe0−/− mice are resistant to intestinal ischemia. To determine if TIPE0 modulated intestinal injury in an acute injury model without a regenerative component, we subjected both Tipe0−/− and Tipe2−/− mice to 60 min of distal ileal ischemia followed by 90 min of reperfusion (I/R90′). Compared with wildtype (WT) mice, Tipe0−/− mice were resistant to intestinal injury, as were the Tipe2−/− mice, but to a lesser degree Fig. 1a, b). The protection in the knockouts could not be explained by global differences in cytokine expression, which was induced to similar levels in all genotypes exposed to I/R90′ (Fig. 1c). Furthermore, we saw increased inflammatory cytokine expression in healthy 2 Tipe2−/− mice as compared with WT controls, and increases in Tnf in the healthy Tipe0−/− mice vs WT, all of which was in accordance with previous literature on these mice10,14–17 and further suggested that decreased inflammatory signaling did not explain the p (...truncated)