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  • Title: Critical role for intracellular calcium in tight junction biogenesis.
    Author: Stuart RO, Sun A, Panichas M, Hebert SC, Brenner BM, Nigam SK.
    Journal: J Cell Physiol; 1994 Jun; 159(3):423-33. PubMed ID: 8188760.
    Abstract:
    Using the Madin Darby canine kidney (MDCK) cell "calcium switch," we have previously demonstrated that, as MDCK cells establish contact and ultimately form tight junctions, there are marked global and localized changes in intracellular calcium at the sites of cell-cell contact (Nigam et al., 1992, Proc. Natl. Acad. Sci. USA, 89:6162-6166). We have now examined whether intracellular Ca++ is critical to the biogenesis of tight junctions by chelating this ion and monitoring the formation of junctions by electrical, immunocytochemical, and biochemical criteria. Intracellular Ca++ was chelated with the cell-permeant chelators, dimethyl-BAPTA-AM and BAPTA-AM. By digital imaging of fura-2 loaded cells, it was demonstrated that both agents efficiently chelated Ca++ during the "switch" in a dose-dependent manner which paralleled their respective in vitro affinities for Ca++. Chelation of Ca++ during the switch markedly attenuated the development of transepithelial electrical resistance (TER), a measure of tight junction assembly. Immunofluorescent staining of the tight junctional protein, zonula occludens-1 (ZO-1), revealed that chelation of intracellular Ca++ retarded the movement of ZO-1 from intracellular sites to the plasma membrane during the switch. During the development of tight junctions, a fraction of ZO-1 redistributed from the Triton X-100 soluble to the Triton X-100 insoluble pool; chelation of Ca++ during the induction of cell-cell contact prevented this stabilization into the Triton X-100 insoluble fraction. Taken together, these data indicate an important role for intracellular Ca++ in tight junction biogenesis and suggest a specific role for calcium in the early sorting and possible cytoskeletal association of tight junction components.
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