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 (InsP3)-induced Ca2+ signals is important for the activation of many physiological functions. In epithelial cells the spatial distribution of InsP3 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 InsP3 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 InsP3 receptor was found to co-localize 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 InsP3 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 InsP3 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 InsP3 receptor in MDCK depends on polarity.
Colosetti
Pascal
P
INSERM U-442, Signalisation cellulaire et calcium, Bât 443, Université Paris-Sud, 91405 Orsay Cedex, France.
Tunwell
Richard E A
RE
Cruttwell
Caroline
C
Arsanto
Jean-Pierre
JP
Mauger
Jean-Pierre
JP
Cassio
Doris
D
D000818:Animals; D002118:Calcium / Q000172:deficiency / Q000494:pharmacology; D015220:Calcium Channels / Q000235:genetics / Q000378:metabolism*; D020013:Calcium Signaling / Q000187:drug effects / Q000502:physiology; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism* / Q000648:ultrastructure; D016764:Cell Polarity / Q000187:drug effects / Q000502:physiology*; D003593:Cytoplasm / Q000378:metabolism; D004285:Dogs; D004721:Endoplasmic Reticulum / Q000378:metabolism / Q000648:ultrastructure; D005455:Fluorescent Antibody Technique; D007150:Immunohistochemistry; D015544:Inositol 1,4,5-Trisphosphate / Q000378:metabolism; D053496:Inositol 1,4,5-Trisphosphate Receptors; D007668:Kidney / Q000166:cytology / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D018613:Microscopy, Confocal; D008854:Microscopy, Electron; D062793:Occludin; D010750:Phosphoproteins / Q000378:metabolism; D021381:Protein Transport / Q000187:drug effects / Q000235:genetics; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics / Q000378:metabolism*; D019108:Tight Junctions / Q000378:metabolism* / Q000648:ultrastructure; D014162:Transfection; D062826:Zonula Occludens-1 Protein
D015220:Calcium Channels; D053496:Inositol 1,4,5-Trisphosphate Receptors; D008565:Membrane Proteins; D062793:Occludin; C568986:Ocln protein, mouse; D010750:Phosphoproteins; D018160:Receptors, Cytoplasmic and Nuclear; C569043:Tjp1 protein, mouse; D062826:Zonula Occludens-1 Protein; D015544:Inositol 1,4,5-Trisphosphate; D002118:Calcium