The RNA-binding protein Imp1 promotes a Spdef transcriptional program and mucus fucosylation during necrotizing enterocolitis

ABSTRACT Background In the United States over 10% of all neonates are born premature (less than 37 weeks gestational age), and many face complications related to prematurity, including necrotizing enterocolitis (NEC). NEC is the most deadly gastrointestinal disease and the leading cause of death in preterm neonates, with up to 50% mortality. Since there is no cure for NEC, prevention is the best strategy. Enhancing our understanding of intestinal epithelial cell (IEC) responses to NEC damage will provide novel therapeutic targets to prevent NEC. Published evidence suggests that the RNA-binding protein insulin-like growth factor 2 mRNA binding protein 1 (IMP1) plays roles in intestinal development, barrier function, and intestinal repair. Notably, however, roles for IMP1 in NEC are not defined. Goblet cells produce protective mucus in the intestine, and their mature function is dependent on the transcription factor Spdef. Emerging evidence suggests that goblet cell mucus complexity impacts barrier function and inflammation susceptibility. This study aimed to define the role of IMP1 in NEC pathogenesis using neonatal human enteroids and a model of NEC-like intestinal injury in mice with IEC-specific Imp1 overexpression and loss. Hypothesis IMP1 expression is protective in NEC. Methods This study used mice with intestinal epithelial Imp1 overexpression or loss and corresponding wild-type controls. At post-natal day 3, mice of both sexes were randomly assigned to control or NEC groups. NEC was induced with the well-established experimental NEC-like intestinal injury model that includes stress, formula feeding, and hypoxia. Imp1 effects on experimental NEC were assessed using RNA sequencing, western blotting, and immunostaining. Results Inflammatory bacteria induced IMP1 expression in neonatal human enteroids. Mice with Imp1 overexpression incur worse intestinal damage during NEC. Pathway analysis of RNA sequencing data revealed a significant enrichment of the Spdef transcriptional network in Imp1 IEC-OE during NEC. This included significant upregulation of Spdef target genes such as Agr2 (in NEC WT: 617.8 ± 33.56 vs Imp1 IEC-OE : 812.9 ± 111.3, p =0.02) and Fut2 (in NEC WT: 219.4 ± 34 vs Imp1 IEC-OE : 396.6 ± 62.9, p =0.05). In silico analysis predicted Imp1 binding to Spdef and mucus glycosylation mediator mRNAs. Although genotype did not affect goblet cell number, Imp1 IEC-OE mice with NEC exhibited significant increases ( p 0.05) in Spdef protein, genes responsible for goblet cell function ( Spink4, Klk1 , Tspg1 ) and mucus glycosylation ( Gcnt3 , B3gnt7, Qsox1 ). Ultimately, Imp1 overexpression led to increased mucus fucosylation during NEC. Conclusion Our data indicate that during NEC, upregulation of Imp1 promotes goblet cell function via Spdef, including enhanced goblet cell maturation and mucus fucosylation. NEW AND NOTEWORTHY Expression of the RNA-binding protein Imp1 is enhanced in neonatal human enteroids in response to inflammatory bacteria. Imp1 regulates goblet cell function in response to early intestinal inflammation in a mouse model of necrotizing enterocolitis. Specifically, Imp1 promotes a Spdef transcriptional program by upregulating Spdef, resulting in elevated gene expression of Spdef target genes. Additionally, Imp1 increases the gene expression of α(1,2)fucosyltransferase, Fut2 , and subsequently, production of fucosylated mucus marked by UEA1. Graphical Abstract