title: JRAB/MICAL-L2 is a junctional Rab13-binding protein mediating the endocytic recycling of occludin.
abstract: The dynamic turnover of tight junctions (TJs) is essential for epithelial-mesenchymal transitions and/or mesenchymal-epithelial transitions during epithelial morphogenesis. We previously demonstrated that Rab13 specifically mediates the endocytic recycling of occludin. Here, we identified MICAL-L2 (molecule interacting with CasL-like 2) as a novel Rab13-binding protein. Immunoprecipitation and immunofluorescence microscopy showed that MICAL-L2 specifically bound to the GTP-bound form of Rab13 via its C terminus, which contained a coiled-coil domain, and localized at TJs in epithelial MTD-1A cells. Recycling assay demonstrated that a MICAL-L2 mutant lacking the Rab13-binding domain (MICAL-L2-N) specifically inhibited the endocytic recycling of occludin but not transferrin receptor. Ca2+ switch assay further revealed that MICAL-L2-N as well as Rab13 Q67L inhibited the recruitment of occludin to the plasma membrane, the development of transepithelial electrical resistance, and the formation of a paracellular diffusion barrier. MICAL-L2 was displaced from TJs upon actin depolymerization and was distributed along radiating actin cables and stress fibers in Ca2+-depleted MTD-1A and fibroblastic NIH3T3 cells, respectively. These results suggest that MICAL-L2 mediates the endocytic recycling of occludin and the formation of functional TJs by linking Rab13 to actin cytoskeleton. We rename MICAL-L2 as JRAB (junctional Rab13-binding protein).
authors.affiliation: Department of Biochemistry, Institute of Health Biosciences, University of Tokushima Graduate School, Tokushima 770-8503, Japan.
authors.forename: Tomoya
authors.initials: T
authors.lastname: Terai
authors.forename: Noriyuki
authors.initials: N
authors.lastname: Nishimura
authors.forename: Ikuno
authors.initials: I
authors.lastname: Kanda
authors.forename: Natsuo
authors.initials: N
authors.lastname: Yasui
authors.forename: Takuya
authors.initials: T
authors.lastname: Sasaki
mesh_terms: D008841:Actin Cytoskeleton / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D002118:Calcium / Q000378:metabolism; D003598:Cytoskeletal Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D004285:Dogs; D004705:Endocytosis / Q000235:genetics; D004848:Epithelium / Q000378:metabolism; D005347:Fibroblasts / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism*; D051379:Mice; D008840:Microfilament Proteins; D008969:Molecular Sequence Data; D009154:Mutation; D041681:NIH 3T3 Cells; D062793:Occludin; D017434:Protein Structure, Tertiary; D019108:Tight Junctions / Q000737:chemistry / Q000378:metabolism; D020691:rab GTP-Binding Proteins / Q000378:metabolism*
chemical_list: D003598:Cytoskeletal Proteins; C512298:JRAB protein, mouse; D008565:Membrane Proteins; D008840:Microfilament Proteins; D062793:Occludin; C568986:Ocln protein, mouse; C484177:Rab13 protein, mouse; D020691:rab GTP-Binding Proteins; D002118:Calcium