Background A dysfunctional gut epithelial barrier allows the augmented permeation of endotoxins, luminal antigens, and bacteria into the bloodstream, causing disease. The maintenance of gut epithelial barrier integrity may be regulated by multiple factors. Herein we analyze the role of leucine-rich repeat-containing protein 19 (LRRC19) in regulating the permeability of the gut epithelial barrier. Methods We utilized Lrrc19 knockout (KO) mice and clinical samples through transmission electron, intestinal permeability assay, Western blot, and immunofluorescence staining to characterize the role of LRRC19 in the permeability of the gut epithelial barrier. Results We found that LRRC19, which is expressed in gut epithelial cells, impairs gut barrier function. Transmission electron micrographs revealed a tighter junction and narrower gaps in the colon epithelium cells in LRRC19 KO mice. There were lower levels of serum lipopolysaccharide and 4 kDa-fluorescein isothiocyanate-dextran after gavage in LRRC19 KO mice than in wild-type mice. We found that LRRC19 could reduce the expression of zonula occludens (ZO)-1, ZO-3, and occludin in the colonic epithelial cells. The decreased expression of ZO-1, ZO-3, and occludin was dependent on degrading protein kinase C (PKC) ζ and PKCι/λ through K48 ubiquitination by LRRC19. The expression of LRRC19 was also negatively correlated with ZO-1, ZO-3, occludin, PKCζ, and PKCι/λ in human colorectal cancers. Conclusions The protein LRRC19 can promote the permeability of the gut epithelial barrier through degrading PKC ζ and PKCι/λ to reduce the expression of ZO-1, ZO-3, and occludin.
Background and Aims The increased E. coli in colon are related to the occurrence and development of multiple diseases. Chemokines are shown to possess potentially antimicrobial activity, including Gram positive and negative bacterial pathogens. We here investigated function(s) of chemokine CXCL9 expressed in the gut epithelial cells and mechanism(s) of CXCL9 to kill E. coli. Methods We generated CXCL9 fl/flpvillin-cre T mice (pvillin-cre positive mice) and their control CXCL9 fl/flpvillin-cre wmice (pvillin-cre negative mice), and then employed dextran sulfate sodium (DSS)-mediated colitis model to determine sensitivity of the CXCL9 fl/flpvillin-cre T mice. We analyzed composition of the gut microbiota by using 16S ribosomal RNA (V3-V4 variable region) sequencing and shotgun metagenomic analyses. We generated E. coli ∆FtsX (FtsX depleted E. coli) and E. coli ∆aceE (aceE depleted E. coli) by using bacterium red recombining system to investigate the mechanism(s) of CXCL9 to kill E. coli. Results CXCL9 fl/flpvillin-cre Tmice were more sensitive to chemically induced colitis than their control littermates CXCL9 fl/flpvillin-cre wmice. After DSS treatment, there were markedly increased gut E. coli (Escherichia-Shigella) in the colonic contents of CXCL9 fl/flpvillin-cre T mice as compared to control CXCL9 fl/flpvillin-cre w mice. The increased E. coli could promote colitis through NLRC4 and caspase 1/11-mediated IL-18, which was derived from gut epithelial cells. We finally demonstrated that CXCL9 expressed in gut epithelial cells could kill the overgrown E. coli. E. coli expressed Ftsx and PDHc subunits aceE. E.coli∆aceE but not E. coli∆FtsX were resistant to CXCL9-mediated killing. Conclusions Gut epithelial cells-derived CXCL9 can kill the expanded E. coli through aceE to remain gut homeostasis.
The activation of NLRC4 is a major host response against the infection by intracellular bacteria. However, there still remain challenges in understanding the activation upon sensing of diverse stimuli. We here found that lncRNA LNCGM1082 plays a critical role in the activation of NLRC4. LNCGM1082 in macrophages could affect maturation of interleukin (IL)-1β and pyroptotic cell death only after exposed to NLRC4 ligand. Similar to NLRC4-/mice, LNCGM1082-/mice were high sensitive to Salmonella typhimurium infection. LNCGM1082 de ciency in mouse or human macrophages had reduced IL-1β maturation and pyroptosis. Mechanistically, LNCGM1082 could induce the combination of PKCδ with NLRC4 in both mice and human. There was absence of binding of NLRC4 with PKCδ in LNCGM1082-/macrophages. This lncRNA could be induced by Salmonella typhimurium through TLR5 in the macrophages of both mice and human. Thus, our data indicate that LNCGM1082 induced by TLR5 can mediate the binding of PKCδ with NLRC4 to cause the activation of NLRC4.
The activation of NLRC4 is a major host response against the infection by intracellular bacteria. However, there still remain challenges in understanding the activation upon sensing of diverse stimuli. We here found that lncRNA LNCGM1082 plays a critical role in the activation of NLRC4. LNCGM1082 in macrophages could affect maturation of interleukin (IL)-1β and pyroptotic cell death only after exposed to NLRC4 ligand. Similar to NLRC4-/- mice, LNCGM1082-/- mice were high sensitive to Salmonella typhimurium infection. LNCGM1082 deficiency in mouse or human macrophages had reduced IL-1β maturation and pyroptosis. Mechanistically, LNCGM1082 could induce the combination of PKCδ with NLRC4 in both mice and human. There was absence of binding of NLRC4 with PKCδ in LNCGM1082-/- macrophages. This lncRNA could be induced by Salmonella typhimurium through TLR5 in the macrophages of both mice and human. Thus, our data indicate that LNCGM1082 induced by TLR5 can mediate the binding of PKCδ with NLRC4 to cause the activation of NLRC4.
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