Direct association of connexin36 with zonula occludens-2 and zonula occludens-3

Xinbo Li, Shijun Lu, James I. Nagy

Research output: Contribution to journalArticlepeer-review

29 Scopus citations

Abstract

The gap junction protein connexin36 (Cx36) is widely expressed in neurons and was previously shown to interact with the PDZ domain-containing protein zonula occludens-1 (ZO-1). We investigated whether Cx36 is also able to interact with other members of zonula occludens family of proteins, namely, ZO-2 and ZO-3, the former of which was reported to be co-localized with Cx36 at gap junctions in mouse retina. HeLa cells transfected with Cx36 and cultured βTC-3 cells were found to express ZO-2 and ZO-3, and both of these ZO proteins were co-localized with Cx36 at gap junctional cell-cell contacts. In lysates of Cx36-transfected HeLa cells, ZO-2 and ZO-3 were shown to co-immunoprecipitate with Cx36, whereas Cx36/ZO-2 association was absent in cells transfected with truncated Cx36 lacking its C-terminus SAYV PDZ interaction motif. In vitro pull-down assays revealed that Cx36 interacts with the PDZ1, but not with the other two PDZ domains in ZO-2 or ZO-3. Truncated Cx36 lacking its PDZ binding motif failed to bind the PDZ1 domain of either ZO-2 or ZO-3. A 14 amino acid peptide corresponding to the C-terminus of Cx36 was also shown to interact with the PDZ1 domains of ZO-2 and ZO-3, and this peptide inhibited the association of Cx36 with the PDZ1 domains of these ZO proteins. These results indicate that Cx36 associates with the first PDZ domain of ZO-2 and ZO-3 and that this association requires the C-terminus SAYV sequence in Cx36. These findings, together with the known association of ZO-2 with a variety of proteins, including transcription factors, suggest that ZO-2 may serve to anchor regulatory proteins at gap junctions composed of Cx36.

Original languageEnglish (US)
Pages (from-to)393-402
Number of pages10
JournalNeurochemistry International
Volume54
Issue number5-6
DOIs
StatePublished - May 2009
Externally publishedYes

Keywords

  • Connexin36
  • PDZ domain
  • Zonula occludens-2
  • Zonula occludens-3
  • βTC-3 cells

ASJC Scopus subject areas

  • Cellular and Molecular Neuroscience
  • Cell Biology

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