title: Relationship of serine/threonine phosphorylation/dephosphorylation signaling to glucocorticoid regulation of tight junction permeability and ZO-1 distribution in nontransformed mammary epithelial cells.
abstract: The synthetic glucocorticoid dexamethasone regulates tight junction permeability resulting in an increased transepithelial electrical resistance (TER) of cultured 31EG4 mammary epithelial cells. Inhibition of cellular type 1 and type 2A protein phosphatase activity by okadaic acid reduced the TER of dexamethasone-treated monolayers of 31EG4 cells to basal levels within 24 h. Coincident with the increase in tight junction permeability, immunofluorescence revealed that okadaic acid caused a partial cellular redistribution of the ZO-1 tight junction-associated protein. The potent glucocorticoid antagonist RU486 had no effect on TER or ZO-1 distribution, indicating that the effects of okadaic acid are not a result of disrupting glucocorticoid receptor function. Immunoprecipitation of 32P-labeled cells and V8 protease peptide mapping demonstrated that dexamethasone did not alter ZO-1 phosphorylation. However, consistent with the changes in TER, dexamethasone induced a 2.3-fold stimulation in ZO-1 protein levels which was reduced to 73% of basal levels by okadaic acid. No effects on ZO-1 transcript levels were observed. Monolayers grown in the presence of glucocorticoids had only 28% less junction density and 16.5% more linear junction/cell, which cannot account for the observed increases of TER and ZO-1 protein levels. Taken together, our results have shown that a disruption of phosphorylation/dephosphorylation activity overrides the glucocorticoid regulation of tight junction permeability in 31EG4 mammary cells.
authors.affiliation: Department of Molecular and Cell Biology, University of California at Berkeley 94720.
authors.forename: K L
authors.initials: KL
authors.lastname: Singer
authors.forename: B R
authors.initials: BR
authors.lastname: Stevenson
authors.forename: P L
authors.initials: PL
authors.lastname: Woo
authors.forename: G L
authors.initials: GL
authors.lastname: Firestone
mesh_terms: D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D000818:Animals; D015820:Cadherins / Q000235:genetics / Q000378:metabolism; D002451:Cell Compartmentation; D002463:Cell Membrane Permeability / Q000187:drug effects / Q000502:physiology*; D003907:Dexamethasone / Q000494:pharmacology*; D017097:Electric Impedance; D004847:Epithelial Cells; D004848:Epithelium / Q000187:drug effects / Q000502:physiology; D004988:Ethers, Cyclic / Q000494:pharmacology; D007365:Intercellular Junctions / Q000502:physiology*; D007546:Isoquinolines / Q000494:pharmacology; D008321:Mammary Glands, Animal / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism; D051379:Mice; D015735:Mifepristone / Q000494:pharmacology; D019319:Okadaic Acid; D010749:Phosphoprotein Phosphatases / Q000378:metabolism*; D010750:Phosphoproteins / Q000235:genetics / Q000378:metabolism; D010766:Phosphorylation; D010879:Piperazines / Q000494:pharmacology; D047428:Protein Kinase Inhibitors; D012333:RNA, Messenger / Q000032:analysis; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D062826:Zonula Occludens-1 Protein
chemical_list: D015820:Cadherins; D004988:Ethers, Cyclic; D007546:Isoquinolines; D008565:Membrane Proteins; D010750:Phosphoproteins; D010879:Piperazines; D047428:Protein Kinase Inhibitors; D012333:RNA, Messenger; C569043:Tjp1 protein, mouse; D062826:Zonula Occludens-1 Protein; D019319:Okadaic Acid; D015735:Mifepristone; D003907:Dexamethasone; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D010749:Phosphoprotein Phosphatases