Neddylated Cullin 3 is required for vascular endothelial-cadherin-mediated endothelial barrier function

Tomohisa Sakaue, Ayako Fujisaki, Hironao Nakayama, Masashi Maekawa, Hiromi Hiyoshi, Eiji Kubota, Takashi Joh, Hironori Izutani, Shigeki Higashiyama

Research output: Contribution to journalArticlepeer-review

13 Citations (Scopus)

Abstract

Vascular endothelial (VE)-cadherin, a major endothelial adhesion molecule, regulates vascular permeability, and increased vascular permeability has been observed in several cancers. The aim of this study was to elucidate the role of the NEDD8-Cullin E3 ligase, in maintaining barrier permeability. To this end, we investigated the effects of the inhibition of Cullin E3 ligases, by using inhibitors and knockdown techniques in HUVECs. Furthermore, we analyzed the mRNA and protein levels of the ligases by quantitative RT-PCR and Western blotting, respectively. The results revealed that NEDD8-conjugated Cullin 3 is required for VE-cadherin-mediated endothelial barrier functions. Treatment of HUVECs with MLN4924, a chemical inhibitor of the NEDD8-activating enzyme, led to high vascular permeability due to impaired cell–cell contact. Similar results were obtained when HUVECs were treated with siRNA directed against Cullin 3, one of the target substrates of NEDD8. Immunocytochemical staining showed that both treatments equally depleted VE-cadherin protein localized at the cell–cell borders. However, quantitative RT-PCR showed that there was no significant difference in the VE-cadherin mRNA levels between the treatment and control groups. In addition, cycloheximide chase assay revealed that the half-life of VE-cadherin protein was dramatically reduced by Cullin 3 depletion. Together, these findings suggest that neddylated Cullin 3 plays a crucial role in endothelial cell barrier function by regulating VE-cadherin.

Original languageEnglish
Pages (from-to)208-215
Number of pages8
JournalCancer science
Volume108
Issue number2
DOIs
Publication statusPublished - 2017 Feb 1
Externally publishedYes

Keywords

  • Cullin 3
  • endothelial barrier function
  • MLN4924
  • neddylation
  • vascular endothelial-cadherin

ASJC Scopus subject areas

  • Oncology
  • Cancer Research

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