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Trans-Golgi protein TVP23B regulates host-microbe interactions via Paneth cell homeostasis and Goblet cell glycosylation

ABSTRACT: A key feature in intestinal immunity is the dynamic intestinal barrier, which separates the host from resident and pathogenic microbiota through a mucus gel impregnated with antimicrobial peptides. The mechanisms underlying the maintenance and function of this intestinal barrier are not completely understood. Using a mouse forward genetic screen for defects of intestinal homeostasis, we have found a mutation in Tvp23b, which conferred susceptibility to chemically induced and infectious colitis. Golgi apparatus membrane protein TVP23 homolog B (TVP23B) is a transmembrane protein conserved from yeast to humans. In the intestine, the protein is localized to the epithelium and its deficiency in the hematopoietic extrinsic compartment was essential to the colitis phenotype. We found that TVP23B controls the homeostasis of Paneth cells and function of goblet cells in vivo, leading to a decrease in antimicrobial peptides as well as a more penetrable mucus layer. As a result, Tvp23b-/- mice displayed decreased barrier function and a loss of host-microbe separation. TVP23B-deficient colonocytes have a loss of core-3 O-glycosylation of colonic proteins, which is the major O-glycosylation present on gel forming mucins. TVP23B binds with another Golgi protein, YIPF6, which is similarly critical for intestinal homeostasis. The Golgi proteomes of YIPF6 and TVP23B-deficient colonocytes have a common deficiency of several critical glycosylation enzymes, including those necessary for core-3 glycosylation of mucins. TVP23B is necessary for the formation of the sterile mucin layer of the intestine and its absence disturbs the balance of host and microbe in vivo.

INSTRUMENT(S): Q Exactive HF

ORGANISM(S): Mus Musculus (ncbitaxon:10090)

SUBMITTER: Emre E. Turer

PROVIDER: MSV000090883 | MassIVE | Mon Dec 12 09:01:00 GMT 2022

SECONDARY ACCESSION(S): PXD038751

REPOSITORIES: MassIVE

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Trans-Golgi protein TVP23B regulates host-microbe interactions via Paneth cell homeostasis and Goblet cell glycosylation.

Song Ran R McAlpine William W Fond Aaron M AM Nair-Gill Evan E Choi Jin Huk JH Nyström Elisabeth E L EEL Arike Liisa L Field Sydney S Li Xiaohong X SoRelle Jeffrey A JA Moresco James J JJ Moresco Eva Marie Y EMY Yates John R JR Azadi Parastoo P Ni Josephine J Birchenough George M H GMH Beutler Bruce B Turer Emre E EE

Nature communications 20230620 1

A key feature in intestinal immunity is the dynamic intestinal barrier, which separates the host from resident and pathogenic microbiota through a mucus gel impregnated with antimicrobial peptides. Using a forward genetic screen, we have found a mutation in Tvp23b, which conferred susceptibility to chemically induced and infectious colitis. Trans-Golgi apparatus membrane protein TVP23 homolog B (TVP23B) is a transmembrane protein conserved from yeast to humans. We found that TVP23B controls the ...[more]

PMID: 37339972

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Project description:Background; MUC2 mucin produced by intestinal goblet cells is the major component of the intestinal mucus barrier. MUC2 homo-oligomerizes intracellularly into large secreted polymers which give mucus its viscous properties. The inflammatory bowel disease (IBD) ulcerative colitis is characterized by depleted goblet cells and a reduced mucus layer, whereas goblet cells and the mucus layer are increased in the other major inflammatory bowel disease, Crohnâs disease. Methods and Findings; By murine N-ethyl-N-nitrosourea-mutagenesis we identified two distinct non-complementing missense mutations in Muc2 exons encoding N- and C-terminal homo-oligomerization domains causing an ulcerative colitis-like phenotype. Both strains developed mild spontaneous distal intestinal inflammation, chronic diarrhea, rectal bleeding and prolapse, increased susceptibility to acute and chronic colitis induced by a luminal toxin, aberrant Muc2 biosynthesis, smaller goblet cell thecae (less stored mucin) and a diminished mucus barrier. Enhanced local production of IL-1beta, TNF-alpha and IFN-gamma was seen in the distal colon. The number of leukocytes within mesenteric lymph nodes was increased five-fold and leukocytes cultured in vitro produced both Th1 and Th2 cytokines (IFN-gamma, TNF-alpha and IL-13). Intestinal permeability was increased and the luminal bacterial flora were more heavily coated with immunoglobulin as occurs in IBD. This pathology was accompanied by accumulation of the Muc2 precursor and ultrastructural and biochemical evidence of endoplasmic reticulum (ER) stress in goblet cells, activation of the unfolded protein response, and altered intestinal expression of genes involved in ER stress, inflammation, apoptosis and wound repair. Expression of mutated Muc2 oligomerization domains in vitro demonstrated that aberrant Muc2 oligomerization underlies the ER stress. These models show that mutations in Muc2 oligomerization domains can lead to aberrant assembly of the Muc2 complex leading to ER stress, a depleted mucus barrier and intestinal inflammation. In ulcerative colitis we demonstrate similar accumulation of non-glycosylated MUC2 precursor in goblet cells together with ultrastructural and biochemical evidence of ER stress even in non-inflamed intestinal tissue. Conclusions; The observations that mucin misfolding and ER stress lead directly to intestinal inflammation and that ER stress and goblet cell pathology occur in ulcerative colitis suggest that ER stress-related mucin depletion could be a fundamental component of the pathogenesis of colitis. Experiment Overall Design: 3 individual mice from the Eeyore, Winnie or Wild-type strains were compared as groups. An Affymetrix ID was compared between groups if the ID was Present within two of the three mice within each grouping. IDs were compared by calculating the log2 of Group One average signal divided by Group 2 average signal.

Project description:Background MUC2 mucin produced by intestinal goblet cells is the major component of the intestinal mucus barrier. MUC2 homo-oligomerizes intracellularly into large secreted polymers which give mucus its viscous properties. The inflammatory bowel disease (IBD) ulcerative colitis is characterized by depleted goblet cells and a reduced mucus layer, whereas goblet cells and the mucus layer are increased in the other major inflammatory bowel disease, Crohn’s disease. Methods and Findings By murine N-ethyl-N-nitrosourea-mutagenesis we identified two distinct non-complementing missense mutations in Muc2 exons encoding N- and C-terminal homo-oligomerization domains causing an ulcerative colitis-like phenotype. Both strains developed mild spontaneous distal intestinal inflammation, chronic diarrhea, rectal bleeding and prolapse, increased susceptibility to acute and chronic colitis induced by a luminal toxin, aberrant Muc2 biosynthesis, smaller goblet cell thecae (less stored mucin) and a diminished mucus barrier. Enhanced local production of IL-1beta, TNF-alpha and IFN-gamma was seen in the distal colon. The number of leukocytes within mesenteric lymph nodes was increased five-fold and leukocytes cultured in vitro produced both Th1 and Th2 cytokines (IFN-gamma, TNF-alpha and IL-13). Intestinal permeability was increased and the luminal bacterial flora were more heavily coated with immunoglobulin as occurs in IBD. This pathology was accompanied by accumulation of the Muc2 precursor and ultrastructural and biochemical evidence of endoplasmic reticulum (ER) stress in goblet cells, activation of the unfolded protein response, and altered intestinal expression of genes involved in ER stress, inflammation, apoptosis and wound repair. Expression of mutated Muc2 oligomerization domains in vitro demonstrated that aberrant Muc2 oligomerization underlies the ER stress. These models show that mutations in Muc2 oligomerization domains can lead to aberrant assembly of the Muc2 complex leading to ER stress, a depleted mucus barrier and intestinal inflammation. In ulcerative colitis we demonstrate similar accumulation of non-glycosylated MUC2 precursor in goblet cells together with ultrastructural and biochemical evidence of ER stress even in non-inflamed intestinal tissue. Conclusions The observations that mucin misfolding and ER stress lead directly to intestinal inflammation and that ER stress and goblet cell pathology occur in ulcerative colitis suggest that ER stress-related mucin depletion could be a fundamental component of the pathogenesis of colitis. Keywords: Single gene, multiple mutant comparison