Research ArticleVASCULAR BIOLOGY

PDZRN3 destabilizes endothelial cell-cell junctions through a PKCζ-containing polarity complex to increase vascular permeability

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Sci. Signal.  31 Jan 2017:
Vol. 10, Issue 464, eaag3209
DOI: 10.1126/scisignal.aag3209

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Destabilizing endothelial cell connections

Interconnected endothelial cells that line blood vessels form a barrier between the circulatory system and tissues. The integrity of the intercellular junctions between endothelial cells regulates vascular permeability, which is the leakiness of blood vessels. In stroke patients, too much fluid can accumulate in the brain and cause irreparable damage. Sewduth et al. elucidated a signaling pathway mediated by the E3 ubiquitin ligase PDZRN3 that regulated endothelial intercellular junctions and vascular permeability. Developing mice that overexpressed Pdzrn3 in endothelial cells died of hemorrhaging in multiple tissues, whereas genetic ablation of Pdzrn3 in adult mice decreased the brain edema that occurred after stroke, an effect mimicked by a pharmacological inhibitor of PKCζ, a kinase that destabilizes endothelial intercellular junctions. Manipulating Pdzrn3 abundance in endothelial cells affected the localization of PKCζ to intercellular junctions, activation of PKCζ, and permeability. These results suggest that inhibiting PDZRN3 or its downstream effector PKCζ may prevent the pathological edema that occurs in conditions such as stroke.

Abstract

Endothelial cells serve as a barrier between blood and tissues. Maintenance of the endothelial cell barrier depends on the integrity of intercellular junctions, which is regulated by a polarity complex that includes the ζ isoform of atypical protein kinase C (PKCζ) and partitioning defective 3 (PAR3). We revealed that the E3 ubiquitin ligase PDZ domain–containing ring finger 3 (PDZRN3) regulated endothelial intercellular junction integrity. Endothelial cell–specific overexpression of Pdzrn3 led to early embryonic lethality with severe hemorrhaging and altered organization of endothelial intercellular junctions. Conversely, endothelial-specific loss of Pdzrn3 prevented vascular leakage in a mouse model of transient ischemic stroke, an effect that was mimicked by pharmacological inhibition of PKCζ. PDZRN3 regulated Wnt signaling and associated with a complex containing PAR3, PKCζ, and the multi-PDZ domain protein MUPP1 (Discs Lost–multi-PDZ domain protein 1) and targeted MUPP1 for proteasomal degradation in transfected cells. Transient ischemic stroke increased the ubiquitination of MUPP1, and deficiency of MUPP1 in endothelial cells was associated with decreased localization of PKCζ and PAR3 at intercellular junctions. In endothelial cells, Pdzrn3 overexpression increased permeability through a PKCζ-dependent pathway. In contrast, Pdzrn3 depletion enhanced PKCζ accumulation at cell-cell contacts and reinforced the cortical actin cytoskeleton under stress conditions. These findings reveal how PDZRN3 regulates vascular permeability through a PKCζ-containing complex.

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