Using MUC2 Mucin Producing Tumorigenic Human Goblet-Like Cells to Uncover Functional Properties of the Mucus Barrier

Goblet cells are the guardians that produce MUC2 mucin that forms the protective mucus barrier as the first line of innate host defense against pathogen invasion while sustaining a healthy gut microbiota. Using an unbiased screen, we cloned a high LS174T colonic adenocarcinoma MUC2 mucin-producing goblet-like cell that revealed by whole genome sequencing, mutations that upregulated the expression of tumor suppressor TP53, anterior gradient 2 (AGR2) and mitogen-activated protein kinase kinases (MEK and MAPK) genes that enhanced MUC2 biosynthesis and secretion. Hypersecretory mutant (Mut) cells exhibited a phenotype characterized by high constitutive MUC2 mRNA, upregulation of glycosyltransferases, and hypersecretion of sialylated and fucosylated MUC2 mucus enumerated by glycomics analysis. Mut secreted mucus was altered quantified by increased permeability to fluorescence microsphere (0.2 μm and 1 μm) beads and to Salmonella enterica adherence, invasion, and cell death as compared to WT controls. Shotgun proteomic analysis revealed that Mut cells were upregulated in metabolic pathways for oxidative stress, HIF pathways and growth factors that enhanced wound healing, and tumorigenesis. These results highlight the importance of regulatory genes for proper mucus production in providing barrier function and homeostasis in the gastrointestinal tract that is markedly affected by metabolic stress and glycosylation prevalent in colonic carcinomas.