The type 3 inositol 1,4,5-trisphosphate receptor is concentrated at the tight junction level in polarized MDCK cells

The subcellular localization of inositol 1,4,5-trisphosphate (InsP(3))-induced Ca2+ signals is important for the activation of many physiological functions. In epithelial cells the spatial distribution of InsP(3) receptor is restricted to specific areas, but little is known about the relationship between the receptor's distribution and cell polarity. To investigate this relationship, the best known polarized cell model, MDCK, was examined. This cell line is characterized by a strong expression of the type 3 InsP(3) receptor and the subcellular localization of this receptor was followed during cell polarization using immunofluorescence and confocal analysis. In non-polarized cells, including ras transformed f3 MDCK cells, the type 3 InsP(3) receptor was found to colocalize with markers of the endoplasmic reticulum in the cytoplasm. In contrast, in polarized cells, this receptor was mostly distributed at the apex of the lateral plasma membrane with the markers of tight junctions, ZO-1 and occludin. The localization of the type 3 InsP(3) receptor in the vicinity of tight junctions was confirmed by immunogold electron microscopy. The culture of MDCK cells in calcium-deprived medium, led to disruption of cell polarity and receptor redistribution in the cytoplasm. Addition of calcium to these deprived cells induced the restoration of polarity and the relocalization of the receptor to the plasma membrane. MDCK cells were stably transfected with a plasmid coding the full-length mouse type 1 InsP(3) receptor tagged with EGFP at the C-terminus. The EGFP-tagged type 1 receptor and the endogenous type 3 co-localized in the cytoplasm of non-polarized cells and at the tight junction level of polarized cells. Thus, the localization of InsP(3) receptor in MDCK depends on polarity.