Eisenmesser, E. Z. | Search bioRxiv

bioRxiv2023

IL-38 regulates intestinal stem cell homeostasis by inducing WNT signaling and beneficial IL-1beta secretion

The IL-1 Family member IL-38 has been characterized primarily as an anti-inflammatory cytokine in human and mouse models of systemic diseases. Here, we examined the role of IL-38 in the murine small intestine (SI). Immunostaining of SI revealed that IL-38 expression partially confines to intestinal stem cells. Cultures of intestinal organoids reveal IL-38 functions as a growth factor by increasing organoid size via inducing WNT3a. In contrast, organoids from IL-38 deficient mice develop more slowly. This reduction in size is likely due to downregulation of intestinal stemness markers (i.e., Fzd5, Ephb2, Olfm4) expression compared with wild type organoids. IL-38 binding to IL-1R6 is postulated to recruit the co-receptor IL-1R9. Therefore, to analyze the molecular mechanisms of IL-38 signaling, we also examined organoids from IL-1R9 deficient mice. Unexpectedly, these organoids, although significantly smaller than wild type, respond to IL-38, suggesting that IL-1R9 is not involved in IL-38 signaling in the stem cell crypt. Nevertheless, silencing of IL-1R6 disabled the organoid response to the growth property of IL-38, thus suggesting IL-1R6 as the main receptor used by IL-38 in the crypt compartment. In organoids from wild type mice, IL-38 stimulation induced low concentrations of IL-1{beta} which contribute to organoid growth. However, high concentrations of IL-1{beta} have detrimental effects on the cultures that were prevented by treatment with recombinant IL-38. Overall, our data demonstrate an important regulatory function of IL-38 as a growth factor, and as an anti-inflammatory molecule in the SI, maintaining homeostasis. SignificanceThe IL-1 family member IL-38 has been characterized primarily as an anti-inflammatory cytokine for systemic diseases. Here we describe a fundamental role of IL-38 in driving intestinal stem cell differentiation through the upregulation of WNT3a and IL-1{beta}. Our findings reveal a dual role of IL-38 in regulating intestinal functions; (a) in resting conditions IL-38 maintains intestinal homeostasis, driving WNT3a production and organoid budding, whereas (b) in highly inflamed conditions, IL-38 contributes to proper recovery, by exerting anti-inflammatory activities. Thus, we demonstrate a pivotal role of IL-38 in driving tissue turnover and maintenance of homeostasis in intestinal health.

cell biology↗

bioRxiv2020

The interactome of the N-terminus of band 3 regulates red blood cell metabolism and storage quality

Band 3 (anion exchanger 1 - AE1) is the most abundant membrane protein in red blood cells (RBCs), the most abundant cell in the human body. A compelling model, based on indirect evidence, posits that - at high oxygen saturation - the N-term cytosolic domain of AE1 binds to and inhibits glycolytic enzymes, thus diverting metabolic fluxes to the pentose phosphate pathway to generate reducing equivalents. Dysfunction of this mechanism occurs during RBC aging or storage under blood bank conditions, suggesting a role for AE1 in the regulation of blood storage quality and efficacy of transfusion - a life-saving intervention for millions of recipients worldwide. Here we leverage two murine models carrying genetic ablations of AE1 to provide the first direct mechanistic evidence of its role in metabolic regulation and blood storage quality. Observations in mice phenocopied those in a human subject lacking expression of AE11-11 (band 3 Neapolis), while common polymorphisms in the region coding for AE11-56 increased susceptibility to osmotic hemolysis in healthy blood donors. Through thermal proteome profiling and cross-linking proteomics, we provide the first comprehensive analysis of the RBC interactome, with a focus on AE11-56 and validate recombinant AE1 interactions with glyceraldehyde 3-phosphate dehydrogenase (GAPDH). Finally, we show that incubation with a cell-penetrating AE11-56 peptide can rescue the metabolic defect in glutathione recycling and boost post-transfusion recoveries of stored RBCs from healthy human donors and genetically ablated mice, paving the way for the in vivo metabolic manipulation of RBCs facing oxidant stress - a landmark of many diseases. Key pointsO_LIGenetic ablation of N-term of band 3 results in significant metabolic aberrations and poor post-transfusion recoveries in mice and humans; C_LIO_LIStructural studies on the N-term of band 3 reveal a complex interactome with several enzymes, including GAPDH; C_LI

biochemistry↗