Translation initiation factor eIF2Bε promotes Wnt-mediated clonogenicity and global translation in intestinal epithelial cells.

Modulation of global mRNA translation, which is essential for intestinal stem cell function, is controlled by Wnt signaling. Loss of tumor supressor APC in stem cells drives adenoma formation through hyperactivion of Wnt signaling and dysregulated translational control. It is unclear whether factors that coordinate global translation in the intestinal epithelium are needed for APC-driven malignant transformation. Here we identified nucleotide exchange factor eIF2Bε as a translation initiation factor involved in Wnt-mediated intestinal epithelial stemness. Using eIF2BεArg191His mice with a homozygous point mutation that leads to dysfunction in the enzymatic activity, we demonstrate that eIF2Bε is involved in small intestinal crypt formation, stemness marker expression, and secreted Paneth cell-derived granule formation. Wnt hyperactivation in ex vivo eIF2BεArg191His organoids, using a GSK3ß inhibitor to mimic Apc driven transformation, shows that eIF2Bε is essential for Wnt-mediated clonogenicity and associated increase of the global translational capacity. Finally, we observe high eIF2Bε expression in human colonic adenoma tissues, exposing eIF2Bε as a potential target of CRC stem cells with aberrant Wnt signaling.

Induction of endoplasmic reticulum stress by deletion of Grp78 depletes Apc mutant intestinal epithelial stem cells.

Intestinal epithelial stem cells are highly sensitive to differentiation induced by endoplasmic reticulum (ER) stress. Colorectal cancer develops from mutated intestinal epithelial stem cells. The most frequent initiating mutation occurs in Apc, which results in hyperactivated Wnt signalling. This causes hyperproliferation and reduced sensitivity to chemotherapy, but whether these mutated stem cells are sensitive to ER stress induced differentiation remains unknown. Here we examined this by generating mice in which both Apc and ER stress repressor chaperone Grp78 can be conditionally deleted from the intestinal epithelium. For molecular studies, we used intestinal organoids derived from these mice. Homozygous loss of Apc alone resulted in crypt elongation, activation of the Wnt signature and accumulation of intestinal epithelial stem cells, as expected. This phenotype was however completely rescued on activation of ER stress by additional deletion of Grp78. In these Apc-Grp78 double mutant animals, stem cells were rapidly lost and repopulation occurred by non-mutant cells that had escaped recombination, suggesting that Apc-Grp78 double mutant stem cells had lost self-renewal capacity. Although in Apc-Grp78 double mutant mice the Wnt signature was lost, these intestines exhibited ubiquitous epithelial presence of nuclear ß-catenin. This suggests that ER stress interferes with Wnt signalling downstream of nuclear ß-catenin. In conclusion, our findings indicate that ER stress signalling results in loss of Apc mutated intestinal epithelial stem cells by interference with the Wnt signature. In contrast to many known inhibitors of Wnt signalling, ER stress acts downstream of ß-catenin. Therefore, ER stress poses a promising target in colorectal cancers, which develop as a result of Wnt activating mutations.

Oral tolerance is inefficient in neonatal mice due to a physiological vitamin A deficiency.

Increased risk of allergy during early life indicates deficient immune regulation in this period of life. To date, the cause for inefficient neonatal immune regulation has never been elucidated. We aimed to define the ontogeny of oral tolerance and to identify necessary conditions specific for this stage of life. Ovalbumin (OVA) was administered orally to mice through breast milk and efficiency of systemic tolerance to OVA was assessed in adulthood using a model of allergic airway inflammation. Oral tolerance induction was fully efficient starting third week of life. Inefficiency in neonates was a consequence of abnormal antigen transfer across the gut barrier and retinaldehyde dehydrogenase expression by mesenteric lymph node CD103(+) neonatal dendritic cells, resulting in inefficient T-cell activation. Neonates' serum retinol levels were three times lower than in adult mice, and vitamin A supplementation was sufficient to rescue neonatal defects and allow tolerance induction from birth. The establishment of oral tolerance required the differentiation of Th1 lymphocytes in both vitamin A-supplemented neonates and 3-week-old unsupplemented mice. This knowledge should guide the design of interventions for allergy prevention that are adapted to the neonatal stage of life such as vitamin A supplementation.

Endogenous c-Myc is essential for p53-induced apoptosis in response to DNA damage in vivo.

Recent studies have suggested that C-MYC may be an excellent therapeutic cancer target and a number of new agents targeting C-MYC are in preclinical development. Given most therapeutic regimes would combine C-MYC inhibition with genotoxic damage, it is important to assess the importance of C-MYC function for DNA damage signalling in vivo. In this study, we have conditionally deleted the c-Myc gene in the adult murine intestine and investigated the apoptotic response of intestinal enterocytes to DNA damage. Remarkably, c-Myc deletion completely abrogated the immediate wave of apoptosis following both ionizing irradiation and cisplatin treatment, recapitulating the phenotype of p53 deficiency in the intestine. Consistent with this, c-Myc-deficient intestinal enterocytes did not upregulate p53. Mechanistically, this was linked to an upregulation of the E3 Ubiquitin ligase Mdm2, which targets p53 for degradation in c-Myc-deficient intestinal enterocytes. Further, low level overexpression of c-Myc, which does not impact on basal levels of apoptosis, elicited sustained apoptosis in response to DNA damage, suggesting c-Myc activity acts as a crucial cell survival rheostat following DNA damage. We also identify the importance of MYC during DNA damage-induced apoptosis in several other tissues, including the thymus and spleen, using systemic deletion of c-Myc throughout the adult mouse. Together, we have elucidated for the first time in vivo an essential role for endogenous c-Myc in signalling DNA damage-induced apoptosis through the control of the p53 tumour suppressor protein.

Rage signalling promotes intestinal tumourigenesis.

Development of colon cancer is a multistep process that is regulated by intrinsic and extrinsic cellular signals. Extrinsic factors include molecular patterns that are derived from either pathogens (PAMPs) or cellular damage (DAMPs). These molecules can promote tumourigenesis by activation of the innate immune system, but the individual contribution of ligands and their receptors remains elusive. The receptor for advanced glycation end products (Rage) is a pattern recognition receptor that binds multiple ligands derived from a damaged cell environment such as Hmgb1 and S100 protein. Here we show that Rage signalling has a critical role in sporadic development of intestinal adenomas, as Apc(Min/+) Rage(-/-) mice are protected against tumourigenesis.

Role of mast cells in colorectal cancer development, the jury is still out.

The link between inflammation and colorectal cancer development is becoming increasingly clear. It had long been recognized that patients with inflammatory bowel disease are at an increased risk of colon cancer. Evidence from experimental animals now also implicates the innate immune system in the development of sporadically occurring intestinal adenomas, the precursors to colorectal cancer. Here we discuss the interaction between the immune system and the adenoma to carcinoma sequence with a special emphasis on the role of mast cells which may play a key role in adenoma development. This article is part of a Special Issue entitled: Mast cells in inflammation.