Rab5a-mediated localization of claudin-1 is regulated by proteasomes in endothelial cells

Asaka M, Hirase T, Hashimoto-Komatsu A, Node K. Rab5a-mediated localization of claudin-1 is regulated by proteasomes in endothelial cells. Am J Physiol Cell Physiol 300: C87-C96, 2011. First published October 6, 2010; doi: 10.1152/ajpcell.00565.2010.Tight junctions composed of transmembrane proteins, including claudin, occludin, and tricellulin, and peripheral membrane proteins are a major barrier to endothelial permeability, whereas the role of claudin in the regulation of tight junction permeability in nonneural endothelial cells is unclear. This study demonstrates that claudin-1 is dominantly expressed and depletion of claudin-1 using small interfering RNA (siRNA) increased tight junction permeability in EA hy. 926 cells, indicating that claudin-1 is a crucial regulator of endothelial tight junction permeability. The ubiquitin-proteasome system has been implicated in the regulation of endocytotic trafficking of plasma membrane proteins. Therefore, the involvement of proteasomes in the localization of claudin-1 was investigated by pharmacological and genetic inhibition of proteasomes using a proteasome inhibitor, N-acetyl- Leu-Leu-Nle-CHO, and siRNA against the beta(5)-subunit of the 20S proteasome, respectively. Claudin-1 was localized at cell-cell contact sites in control cells. Claudin-1 was localized in the cytoplasm in association with Rab5a and EEA-1, a marker of early endosome, following inhibition of proteasomes. Depletion of Rab5a using siRNA reversed the localization of claudin-1 induced by inhibition of proteasomes. These data suggest that proteasomes regulate claudin-1 localization at the plasma membrane, which changes upon proteasomal inhibition to a Rab5a-mediated endosomal localization.