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Zonula occludens-1 is a scaffolding protein for signaling molecules. Galpha(12) directly binds to the Src homology 3 domain and regulates paracellular permeability in epithelial cells

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Zonula occludens-1 is a scaffolding protein for signaling molecules. Galpha(12) directly binds to the Src homology 3 domain and regulates paracellular permeability in epithelial cells. / Meyer, Tobias N; Meyer-Schwesinger, Catherine; Denker, Bradley M.

In: J BIOL CHEM, Vol. 277, No. 28, 12.07.2002, p. 24855-8.

Research output: SCORING: Contribution to journal › SCORING: Journal article › Research › peer-review

Harvard

Meyer, TN, Meyer-Schwesinger, C & Denker, BM 2002, 'Zonula occludens-1 is a scaffolding protein for signaling molecules. Galpha(12) directly binds to the Src homology 3 domain and regulates paracellular permeability in epithelial cells', J BIOL CHEM, vol. 277, no. 28, pp. 24855-8. https://doi.org/10.1074/jbc.C200240200

APA

Meyer, T. N., Meyer-Schwesinger, C., & Denker, B. M. (2002). Zonula occludens-1 is a scaffolding protein for signaling molecules. Galpha(12) directly binds to the Src homology 3 domain and regulates paracellular permeability in epithelial cells. J BIOL CHEM, 277(28), 24855-8. https://doi.org/10.1074/jbc.C200240200

Vancouver

Meyer TN, Meyer-Schwesinger C, Denker BM. Zonula occludens-1 is a scaffolding protein for signaling molecules. Galpha(12) directly binds to the Src homology 3 domain and regulates paracellular permeability in epithelial cells. J BIOL CHEM. 2002 Jul 12;277(28):24855-8. https://doi.org/10.1074/jbc.C200240200

Bibtex

@article{ac30560abb7f4272ba1ee24bddbe57dc,
title = "Zonula occludens-1 is a scaffolding protein for signaling molecules. Galpha(12) directly binds to the Src homology 3 domain and regulates paracellular permeability in epithelial cells",
abstract = "Zonula occludens proteins are multidomain proteins usually localized at sites of intercellular junctions, yet little is known about their role in regulating junctional properties. Multiple signaling proteins regulate the junctional complex, and several (including G proteins) have been co-localized with zonula occludens-1 (ZO-1) in the tight junction of epithelial cells. However, evidence for direct interactions between signaling proteins and tight junction proteins has been lacking. In these studies, we constructed Galpha-glutathione S-transferase (GST) fusion proteins and tested for interactions with [(35)S]methionine-labeled in vitro translated ZO-1 and ZO-2. Only Galpha(12) directly interacted with in vitro translated ZO-1 and ZO-2. Using a series of ZO-1 domains expressed as GST fusion proteins and in vitro translated [(35)S]methionine-labeled Galpha(12), we found that Galpha(12) and constitutively active (Q229L) alpha(12) (QLalpha(12)) bind to the Src homology 3 (SH3) domain of ZO-1. This binding was not detected with SH3 domains from other proteins. Inducible expression of wild-type alpha(12) and QLalpha(12) in Madin-Darby canine kidney (MDCK) cells was established using the Tet-Off system. In Galpha(12)-expressing cells, we found that ZO-1 and Galpha(12) co-localize by confocal microscopy and co-immunoprecipitate. Galpha(12) from MDCK cell lysates bound to the GST-ZO-1-SH3 domain, and expression of QLalpha(12) in MDCK cells reversibly increased paracellular permeability. These studies indicated that ZO-1 directly interacts with Galpha(12) and that Galpha(12) regulates barrier function of MDCK cells.",
keywords = "Animals, Cell Line, Cell Membrane Permeability, Dogs, Epithelial Cells, GTP-Binding Protein alpha Subunits, G12-G13, Heterotrimeric GTP-Binding Proteins, Membrane Proteins, Mice, Phosphoproteins, Signal Transduction, Zonula Occludens-1 Protein, src Homology Domains",
author = "Meyer, {Tobias N} and Catherine Meyer-Schwesinger and Denker, {Bradley M}",
year = "2002",
month = jul,
day = "12",
doi = "10.1074/jbc.C200240200",
language = "English",
volume = "277",
pages = "24855--8",
journal = "J BIOL CHEM",
issn = "0021-9258",
publisher = "American Society for Biochemistry and Molecular Biology Inc.",
number = "28",

}

RIS

TY - JOUR

T1 - Zonula occludens-1 is a scaffolding protein for signaling molecules. Galpha(12) directly binds to the Src homology 3 domain and regulates paracellular permeability in epithelial cells

AU - Meyer, Tobias N

AU - Meyer-Schwesinger, Catherine

AU - Denker, Bradley M

PY - 2002/7/12

Y1 - 2002/7/12

N2 - Zonula occludens proteins are multidomain proteins usually localized at sites of intercellular junctions, yet little is known about their role in regulating junctional properties. Multiple signaling proteins regulate the junctional complex, and several (including G proteins) have been co-localized with zonula occludens-1 (ZO-1) in the tight junction of epithelial cells. However, evidence for direct interactions between signaling proteins and tight junction proteins has been lacking. In these studies, we constructed Galpha-glutathione S-transferase (GST) fusion proteins and tested for interactions with [(35)S]methionine-labeled in vitro translated ZO-1 and ZO-2. Only Galpha(12) directly interacted with in vitro translated ZO-1 and ZO-2. Using a series of ZO-1 domains expressed as GST fusion proteins and in vitro translated [(35)S]methionine-labeled Galpha(12), we found that Galpha(12) and constitutively active (Q229L) alpha(12) (QLalpha(12)) bind to the Src homology 3 (SH3) domain of ZO-1. This binding was not detected with SH3 domains from other proteins. Inducible expression of wild-type alpha(12) and QLalpha(12) in Madin-Darby canine kidney (MDCK) cells was established using the Tet-Off system. In Galpha(12)-expressing cells, we found that ZO-1 and Galpha(12) co-localize by confocal microscopy and co-immunoprecipitate. Galpha(12) from MDCK cell lysates bound to the GST-ZO-1-SH3 domain, and expression of QLalpha(12) in MDCK cells reversibly increased paracellular permeability. These studies indicated that ZO-1 directly interacts with Galpha(12) and that Galpha(12) regulates barrier function of MDCK cells.

AB - Zonula occludens proteins are multidomain proteins usually localized at sites of intercellular junctions, yet little is known about their role in regulating junctional properties. Multiple signaling proteins regulate the junctional complex, and several (including G proteins) have been co-localized with zonula occludens-1 (ZO-1) in the tight junction of epithelial cells. However, evidence for direct interactions between signaling proteins and tight junction proteins has been lacking. In these studies, we constructed Galpha-glutathione S-transferase (GST) fusion proteins and tested for interactions with [(35)S]methionine-labeled in vitro translated ZO-1 and ZO-2. Only Galpha(12) directly interacted with in vitro translated ZO-1 and ZO-2. Using a series of ZO-1 domains expressed as GST fusion proteins and in vitro translated [(35)S]methionine-labeled Galpha(12), we found that Galpha(12) and constitutively active (Q229L) alpha(12) (QLalpha(12)) bind to the Src homology 3 (SH3) domain of ZO-1. This binding was not detected with SH3 domains from other proteins. Inducible expression of wild-type alpha(12) and QLalpha(12) in Madin-Darby canine kidney (MDCK) cells was established using the Tet-Off system. In Galpha(12)-expressing cells, we found that ZO-1 and Galpha(12) co-localize by confocal microscopy and co-immunoprecipitate. Galpha(12) from MDCK cell lysates bound to the GST-ZO-1-SH3 domain, and expression of QLalpha(12) in MDCK cells reversibly increased paracellular permeability. These studies indicated that ZO-1 directly interacts with Galpha(12) and that Galpha(12) regulates barrier function of MDCK cells.

KW - Animals

KW - Cell Line

KW - Cell Membrane Permeability

KW - Dogs

KW - Epithelial Cells

KW - GTP-Binding Protein alpha Subunits, G12-G13

KW - Heterotrimeric GTP-Binding Proteins

KW - Membrane Proteins

KW - Mice

KW - Phosphoproteins

KW - Signal Transduction

KW - Zonula Occludens-1 Protein

KW - src Homology Domains

U2 - 10.1074/jbc.C200240200

DO - 10.1074/jbc.C200240200

M3 - SCORING: Journal article

C2 - 12023272

VL - 277

SP - 24855

EP - 24858

JO - J BIOL CHEM

JF - J BIOL CHEM

SN - 0021-9258

IS - 28

ER -