Interaction of the human prostacyclin receptor with the PDZ adapter protein PDZK1: role in endothelial cell migration and angiogenesis

Abstract

Prostacyclin is widely implicated in re-endothelialization and angiogenesis but through unknown mechanisms. Herein the HDL scavenger receptor class B, type 1 adapter PDZK1 was identified as a direct, functional interactant of the human prostacyclin receptor and was found to influence prostacyclin-mediated endothelial migration and in vitro angiogenesis.

Prostacyclin receptors: Transcriptional regulation and novel signalling mechanisms

“…In brief, key amongst those findings include: (i) PDZK1 directly interacts with the IP involving a Class I PDZ ligand at its C-terminus; (ii) while the interaction occurs constitutively, it is dynamically regulated in response to agonist (cicaprost)-activation of the IP ( Figure 7A); (iii) while PDZK1 did not influence overall levels of the IP, it increases its functional expression at the plasma membrane enhancing agonist binding and cAMP generation; (iv) specifically, in the context of the role of prostacyclin in re-endothelialization, both cicaprost and HDL were confirmed to promote substantial EC migration and in vitro angiogenesis, and in an augmentative manner (Figure 7B), while (v) similar to that previously reported for the HDL/SR-B1-mediated EC responses 117 , siRNA-disruption of PDZK1 abolished cicaprost-induced EC migration and in vitro angiogenesis, and without affecting VEGFmediated responses 106 . A number of studies have suggested that prostacyclin-induced endothelial migration and angiogenesis occurs through its regulation of PPARδ, rather than through the IP per se 22,23 .…”

“…As previously stated, we recently identified the multi PDZ-domain protein PDZK1 (NHERF3) as a highly specific interactant of the IP (Figure 7) 106 . The intracellular scaffold or adapter protein PDZ domaincontaining protein 1 (PDZK1) is a member of the Na + , H + exchanger regulatory family (NHERF) and is predominantly expressed in the brush border of the kidney and small intestine, in epithelial and endothelial cells, in macrophages and in the liver [111][112][113] .…”

“…A number of studies have suggested that prostacyclin-induced endothelial migration and angiogenesis occurs through its regulation of PPARδ, rather than through the IP per se 22,23 . However, in those studies, iloprost was used as the IP agonist, which, unlike the highly selective IP agonist cicaprost used in our own studies 106 , is known to activate both the IP and PPARδ 3 . Moreover, in our studies, the effects of cicaprost were blocked by the IP antagonist RO1138452, confirming that the cicaprost induced EC-migration and angiogenesis are mediated through the IP rather than PPARδ 3 .…”

“…Furthermore, due to its divergent sequence and capacity to contain functionally distinct binding motifs, the intracellular C-tail domain of the GPCR is the critical binding domain for such interactions between GPCRs with their specific GIP(s) [103][104][105] . To identify novel GIPs that interact with the IP, potentially shedding new insights into the role of prostacyclin within the vasculature including in cardioprotection, a yeast-two-hybrid (Y2H)-based screen of a human kidney cDNA library was carried out where the carboxyl (C)-terminal tail domain of the IP (IP 299-386 ) was used as the initial bait protein 96,97,106,107 . Through those studies, Rab11a was identified as a direct binding partner of the IP and this IP:Rab11a interaction was found to be dependent on a 14-residue Rab11 binding domain (RBD) located within its proximal C-tail domain of the IP, comprising Val 299 -Val 307 , adjacent to its palmitoylated residues at Cys 308 -Cys 311 96, 97 .…”

“…Since the discovery of an interaction of the IP with PDZK1, we have made significant advances in validating the interaction and in establishing that, similar to that of SR-B1, the interaction is likely to be functionally important with respect to the protective role of prostacyclin/IP within the vascular endothelium 106 . In brief, key amongst those findings include: (i) PDZK1 directly interacts with the IP involving a Class I PDZ ligand at its C-terminus; (ii) while the interaction occurs constitutively, it is dynamically regulated in response to agonist (cicaprost)-activation of the IP ( Figure 7A); (iii) while PDZK1 did not influence overall levels of the IP, it increases its functional expression at the plasma membrane enhancing agonist binding and cAMP generation; (iv) specifically, in the context of the role of prostacyclin in re-endothelialization, both cicaprost and HDL were confirmed to promote substantial EC migration and in vitro angiogenesis, and in an augmentative manner (Figure 7B), while (v) similar to that previously reported for the HDL/SR-B1-mediated EC responses 117 , siRNA-disruption of PDZK1 abolished cicaprost-induced EC migration and in vitro angiogenesis, and without affecting VEGFmediated responses 106 .…”