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Cx50 requires an intact PDZ-binding motif and ZO-1 for the formation of functional intercellular channels

The three connexins expressed in the ocular lens each contain PDZ domain–binding motifs directing a physical association with the scaffolding protein ZO-1, but the significance of the interaction is unknown. We found that Cx50 with PDZ-binding motif mutations did not form gap junction plaques or ind...

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Detalles Bibliográficos
Autores principales: Chai, Zhifang, Goodenough, Daniel A., Paul, David L.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: The American Society for Cell Biology 2011
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3226470/
https://www.ncbi.nlm.nih.gov/pubmed/21965293
http://dx.doi.org/10.1091/mbc.E11-05-0438
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author Chai, Zhifang
Goodenough, Daniel A.
Paul, David L.
author_facet Chai, Zhifang
Goodenough, Daniel A.
Paul, David L.
author_sort Chai, Zhifang
collection PubMed
description The three connexins expressed in the ocular lens each contain PDZ domain–binding motifs directing a physical association with the scaffolding protein ZO-1, but the significance of the interaction is unknown. We found that Cx50 with PDZ-binding motif mutations did not form gap junction plaques or induce cell–cell communication in HeLa cells, whereas the addition of a seven–amino acid PDZ-binding motif restored normal function to Cx50 lacking its entire C-terminal cytoplasmic domain. C-Terminal deletion had a similar although weaker effect on Cx46 but little if any effect on targeting and function of Cx43. Furthermore, small interfering RNA knockdown of ZO-1 completely inhibited the formation of gap junctions by wild-type Cx50 in HeLa cells. Thus both a PDZ-binding motif and ZO-1 are necessary for Cx50 intercellular channel formation in HeLa cells. Knock-in mice expressing Cx50 with a PDZ-binding motif mutation phenocopied Cx50 knockouts. Furthermore, differentiating lens fibers in the knock-in displayed extensive intracellular Cx50, whereas plaques in mature fibers contained only Cx46. Thus normal Cx50 function in vivo also requires an intact PDZ domain–binding motif. This is the first demonstration of a connexin-specific requirement for a connexin-interacting protein in gap junction assembly.
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spelling pubmed-32264702012-02-16 Cx50 requires an intact PDZ-binding motif and ZO-1 for the formation of functional intercellular channels Chai, Zhifang Goodenough, Daniel A. Paul, David L. Mol Biol Cell Articles The three connexins expressed in the ocular lens each contain PDZ domain–binding motifs directing a physical association with the scaffolding protein ZO-1, but the significance of the interaction is unknown. We found that Cx50 with PDZ-binding motif mutations did not form gap junction plaques or induce cell–cell communication in HeLa cells, whereas the addition of a seven–amino acid PDZ-binding motif restored normal function to Cx50 lacking its entire C-terminal cytoplasmic domain. C-Terminal deletion had a similar although weaker effect on Cx46 but little if any effect on targeting and function of Cx43. Furthermore, small interfering RNA knockdown of ZO-1 completely inhibited the formation of gap junctions by wild-type Cx50 in HeLa cells. Thus both a PDZ-binding motif and ZO-1 are necessary for Cx50 intercellular channel formation in HeLa cells. Knock-in mice expressing Cx50 with a PDZ-binding motif mutation phenocopied Cx50 knockouts. Furthermore, differentiating lens fibers in the knock-in displayed extensive intracellular Cx50, whereas plaques in mature fibers contained only Cx46. Thus normal Cx50 function in vivo also requires an intact PDZ domain–binding motif. This is the first demonstration of a connexin-specific requirement for a connexin-interacting protein in gap junction assembly. The American Society for Cell Biology 2011-12-01 /pmc/articles/PMC3226470/ /pubmed/21965293 http://dx.doi.org/10.1091/mbc.E11-05-0438 Text en © 2011 Chai et al. This article is distributed by The American Society for Cell Biology under license from the author(s). Two months after publication it is available to the public under an Attribution–Noncommercial–Share Alike 3.0 Unported Creative Commons License (http://creativecommons.org/licenses/by-nc-sa/3.0). “ASCB®,” “The American Society for Cell Biology®,” and “Molecular Biology of the Cell®” are registered trademarks of The American Society of Cell Biology.
spellingShingle Articles
Chai, Zhifang
Goodenough, Daniel A.
Paul, David L.
Cx50 requires an intact PDZ-binding motif and ZO-1 for the formation of functional intercellular channels
title Cx50 requires an intact PDZ-binding motif and ZO-1 for the formation of functional intercellular channels
title_full Cx50 requires an intact PDZ-binding motif and ZO-1 for the formation of functional intercellular channels
title_fullStr Cx50 requires an intact PDZ-binding motif and ZO-1 for the formation of functional intercellular channels
title_full_unstemmed Cx50 requires an intact PDZ-binding motif and ZO-1 for the formation of functional intercellular channels
title_short Cx50 requires an intact PDZ-binding motif and ZO-1 for the formation of functional intercellular channels
title_sort cx50 requires an intact pdz-binding motif and zo-1 for the formation of functional intercellular channels
topic Articles
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3226470/
https://www.ncbi.nlm.nih.gov/pubmed/21965293
http://dx.doi.org/10.1091/mbc.E11-05-0438
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