Junctional uvomorulin/E-cadherin and phosphotyrosine-modified protein content are correlated with paracellular permeability in Madin-Darby canine kidney (MDCK) epithelia

C B Collares-Buzato, M A Jepson, G T McEwan, N L Simmons, B H Hirst

Research output: Contribution to journalArticle

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Abstract

Strains I and II of Madin-Darby canine kidney (MDCK) cells, which differ markedly in transepithelial resistance (RT) and paracellular permeability, have been used to investigate whether differences in the cellular content of uvomorulin/E-cadherin and phosphotyrosine may be correlated with junctional properties. Using immunocytochemistry, the strain I "tight" epithelia showed significantly stronger uvomorulin staining at regions of cell-cell contact compared with strain II "leaky" MDCK epithelia. In contrast, strain I MDCK cells showed a relatively faint phosphotyrosine staining, distributed evenly throughout the cytoplasm, while strain II MDCK cells displayed intense staining for phosphotyrosine residues in the junctional region and the lateral cell membrane with additional labelling of the cytoplasm. Exposure to vanadate in conjunction with H2O2 (which are potent inhibitors of protein tyrosine phosphatases) resulted in a dramatic increase in phosphotyrosine staining at the intercellular area and, concomitantly, induced changes in cell morphology, a significant decrease in RT, increase in paracellular inulin permeability, and time-dependent disappearance of uvomorulin from the cell-cell contact sites. Moreover, the effects of vanadate/H2O2 treatment were more dramatic in strain II compared with strain I cells, consistent with greater generation of tyrosine-modified protein in strain II cells. An inverse relationship was demonstrated between membrane-associated uvomorulin/E-cadherin and cellular phosphotyrosine content, which varied between the two strains of MDCK cells and when phosphotyrosine was directly manipulated. These data support the hypothesis that regulation of paracellular permeability may result from specific tyrosine phosphorylation of protein components of the junctional complex.
Original languageEnglish
Pages (from-to)185-94
Number of pages10
JournalJournal of Histochemistry and Cytochemistry
Volume101
Issue number3
Publication statusPublished - Mar 1994

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Phosphotyrosine
cadherins
Cadherins
Canidae
Permeability
permeability
epithelium
Epithelium
protein content
kidneys
Madin Darby Canine Kidney Cells
Kidney
dogs
kidney cells
Proteins
Staining and Labeling
Vanadates
cells
Tyrosine
Cytoplasm

Keywords

  • Animals
  • Cadherins
  • Cell Line
  • Cell Membrane
  • Cell Membrane Permeability
  • Dogs
  • Electric Conductivity
  • Electrophysiology
  • Epithelium
  • Hydrogen Peroxide
  • Intercellular Junctions
  • Kidney
  • Membrane Potentials
  • Microscopy, Electron
  • Microscopy, Electron, Scanning
  • Phosphoproteins
  • Phosphotyrosine
  • Tyrosine
  • Vanadates

Cite this

Junctional uvomorulin/E-cadherin and phosphotyrosine-modified protein content are correlated with paracellular permeability in Madin-Darby canine kidney (MDCK) epithelia. / Collares-Buzato, C B; Jepson, M A; McEwan, G T; Simmons, N L; Hirst, B H.

In: Journal of Histochemistry and Cytochemistry, Vol. 101, No. 3, 03.1994, p. 185-94.

Research output: Contribution to journalArticle

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abstract = "Strains I and II of Madin-Darby canine kidney (MDCK) cells, which differ markedly in transepithelial resistance (RT) and paracellular permeability, have been used to investigate whether differences in the cellular content of uvomorulin/E-cadherin and phosphotyrosine may be correlated with junctional properties. Using immunocytochemistry, the strain I {"}tight{"} epithelia showed significantly stronger uvomorulin staining at regions of cell-cell contact compared with strain II {"}leaky{"} MDCK epithelia. In contrast, strain I MDCK cells showed a relatively faint phosphotyrosine staining, distributed evenly throughout the cytoplasm, while strain II MDCK cells displayed intense staining for phosphotyrosine residues in the junctional region and the lateral cell membrane with additional labelling of the cytoplasm. Exposure to vanadate in conjunction with H2O2 (which are potent inhibitors of protein tyrosine phosphatases) resulted in a dramatic increase in phosphotyrosine staining at the intercellular area and, concomitantly, induced changes in cell morphology, a significant decrease in RT, increase in paracellular inulin permeability, and time-dependent disappearance of uvomorulin from the cell-cell contact sites. Moreover, the effects of vanadate/H2O2 treatment were more dramatic in strain II compared with strain I cells, consistent with greater generation of tyrosine-modified protein in strain II cells. An inverse relationship was demonstrated between membrane-associated uvomorulin/E-cadherin and cellular phosphotyrosine content, which varied between the two strains of MDCK cells and when phosphotyrosine was directly manipulated. These data support the hypothesis that regulation of paracellular permeability may result from specific tyrosine phosphorylation of protein components of the junctional complex.",
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TY - JOUR

T1 - Junctional uvomorulin/E-cadherin and phosphotyrosine-modified protein content are correlated with paracellular permeability in Madin-Darby canine kidney (MDCK) epithelia

AU - Collares-Buzato, C B

AU - Jepson, M A

AU - McEwan, G T

AU - Simmons, N L

AU - Hirst, B H

PY - 1994/3

Y1 - 1994/3

N2 - Strains I and II of Madin-Darby canine kidney (MDCK) cells, which differ markedly in transepithelial resistance (RT) and paracellular permeability, have been used to investigate whether differences in the cellular content of uvomorulin/E-cadherin and phosphotyrosine may be correlated with junctional properties. Using immunocytochemistry, the strain I "tight" epithelia showed significantly stronger uvomorulin staining at regions of cell-cell contact compared with strain II "leaky" MDCK epithelia. In contrast, strain I MDCK cells showed a relatively faint phosphotyrosine staining, distributed evenly throughout the cytoplasm, while strain II MDCK cells displayed intense staining for phosphotyrosine residues in the junctional region and the lateral cell membrane with additional labelling of the cytoplasm. Exposure to vanadate in conjunction with H2O2 (which are potent inhibitors of protein tyrosine phosphatases) resulted in a dramatic increase in phosphotyrosine staining at the intercellular area and, concomitantly, induced changes in cell morphology, a significant decrease in RT, increase in paracellular inulin permeability, and time-dependent disappearance of uvomorulin from the cell-cell contact sites. Moreover, the effects of vanadate/H2O2 treatment were more dramatic in strain II compared with strain I cells, consistent with greater generation of tyrosine-modified protein in strain II cells. An inverse relationship was demonstrated between membrane-associated uvomorulin/E-cadherin and cellular phosphotyrosine content, which varied between the two strains of MDCK cells and when phosphotyrosine was directly manipulated. These data support the hypothesis that regulation of paracellular permeability may result from specific tyrosine phosphorylation of protein components of the junctional complex.

AB - Strains I and II of Madin-Darby canine kidney (MDCK) cells, which differ markedly in transepithelial resistance (RT) and paracellular permeability, have been used to investigate whether differences in the cellular content of uvomorulin/E-cadherin and phosphotyrosine may be correlated with junctional properties. Using immunocytochemistry, the strain I "tight" epithelia showed significantly stronger uvomorulin staining at regions of cell-cell contact compared with strain II "leaky" MDCK epithelia. In contrast, strain I MDCK cells showed a relatively faint phosphotyrosine staining, distributed evenly throughout the cytoplasm, while strain II MDCK cells displayed intense staining for phosphotyrosine residues in the junctional region and the lateral cell membrane with additional labelling of the cytoplasm. Exposure to vanadate in conjunction with H2O2 (which are potent inhibitors of protein tyrosine phosphatases) resulted in a dramatic increase in phosphotyrosine staining at the intercellular area and, concomitantly, induced changes in cell morphology, a significant decrease in RT, increase in paracellular inulin permeability, and time-dependent disappearance of uvomorulin from the cell-cell contact sites. Moreover, the effects of vanadate/H2O2 treatment were more dramatic in strain II compared with strain I cells, consistent with greater generation of tyrosine-modified protein in strain II cells. An inverse relationship was demonstrated between membrane-associated uvomorulin/E-cadherin and cellular phosphotyrosine content, which varied between the two strains of MDCK cells and when phosphotyrosine was directly manipulated. These data support the hypothesis that regulation of paracellular permeability may result from specific tyrosine phosphorylation of protein components of the junctional complex.

KW - Animals

KW - Cadherins

KW - Cell Line

KW - Cell Membrane

KW - Cell Membrane Permeability

KW - Dogs

KW - Electric Conductivity

KW - Electrophysiology

KW - Epithelium

KW - Hydrogen Peroxide

KW - Intercellular Junctions

KW - Kidney

KW - Membrane Potentials

KW - Microscopy, Electron

KW - Microscopy, Electron, Scanning

KW - Phosphoproteins

KW - Phosphotyrosine

KW - Tyrosine

KW - Vanadates

M3 - Article

VL - 101

SP - 185

EP - 194

JO - Journal of Histochemistry and Cytochemistry

JF - Journal of Histochemistry and Cytochemistry

SN - 0022-1554

IS - 3

ER -