title: Numb modulates the paracellular permeability of intestinal epithelial cells through regulating apical junctional complex assembly and myosin light chain phosphorylation.
abstract: Numb is highly expressed throughout the crypt-villus axis of intestinal mucosa and functions as cell fate determinant and integrator of cell-to-cell adhesion. Increased paracellular permeability of intestinal epithelial cells is associated with the epithelial barrier dysfunction of inflammatory bowel diseases (IBDs). The apical junctional complex (AJC) assembly and myosin light chain (MLC) phosphorylation regulate adherens junctions (AJ) and tight junctions (TJ). We determined whether and how Numb modulate the paracellular permeability of intestinal epithelial cells. Caco-2 intestinal epithelial cells and their Numb-interfered counterparts were used in the study for physiological, morphological and biological analyses. Numb, expressed in intestinal epithelial cells and located at the plasma membrane of Caco-2 cells in a basolateral to apical distribution, increased in the intestinal epithelial cells with the formation of the intestinal epithelial barrier. Numb expression decreased and accumulated in the cytoplasm of intestinal epithelial cells in a DSS-induced colitis mouse model. Numb co-localized with E-cadherin, ZO-1 and Par3 at the plasma membrane and interacted with E-cadherin and Par3. Knockdown of Numb in Caco-2 cells altered the F-actin structure during the Ca(2+) switch assay, enhanced TNFα-/INF-γ-induced intestinal epithelial barrier dysfunction and TJ destruction, and increased the Claudin-2 protein level. Immunofluorescence experiments revealed that NMIIA and F-actin co-localized at the cell surface of Caco-2 cells. Numb knockdown in Caco-2 cells increased F-actin contraction and the abundance of phosphorylated MLC. Numb modulated the intestinal epithelial barrier in a Notch signaling-independent manner. These findings suggest that Numb modulates the paracellular permeability by affecting AJC assembly and MLC phosphorylation.
authors.affiliation: Department of Gastroenterology, Southwest Hospital, Third Military Medical University, Chongqing 400038, People's Republic of China.
authors.forename: Yongtao
authors.initials: Y
authors.lastname: Yang
authors.forename: Lei
authors.initials: L
authors.lastname: Chen
authors.forename: Yin
authors.initials: Y
authors.lastname: Tian
authors.forename: Jun
authors.initials: J
authors.lastname: Ye
authors.forename: Yun
authors.initials: Y
authors.lastname: Liu
authors.forename: Lili
authors.initials: L
authors.lastname: Song
authors.forename: Qiong
authors.initials: Q
authors.lastname: Pan
authors.forename: Yonghong
authors.initials: Y
authors.lastname: He
authors.forename: Wensheng
authors.initials: W
authors.lastname: Chen
authors.forename: Zhihong
authors.initials: Z
authors.lastname: Peng
authors.forename: Rongquan
authors.initials: R
authors.lastname: Wang
mesh_terms: D018938:Caco-2 Cells; D002463:Cell Membrane Permeability; D004847:Epithelial Cells / Q000166:cytology* / Q000378:metabolism*; D006801:Humans; D007421:Intestine, Small / Q000166:cytology*; D008565:Membrane Proteins / Q000378:metabolism*; D018994:Myosin Light Chains / Q000378:metabolism*; D009419:Nerve Tissue Proteins / Q000378:metabolism*; D010766:Phosphorylation; D019108:Tight Junctions / Q000378:metabolism*; D014407:Tumor Cells, Cultured
chemical_list: D008565:Membrane Proteins; D018994:Myosin Light Chains; D009419:Nerve Tissue Proteins; C113026:NUMB protein, human
keywords: AJ; AJC; Adhesion; Apical junctional complex; DSS; IBD; MLC; Myosin light chain; NMIIA; Numb; Paracellular permeability; TEER; TJ; adherens junction; apical junction complex; dextran sulfate sodium; inflammatory bowel diseases; myosin light chain; non-muscle myosin II A; tight junction; trans-epithelial electric resistance