Identification of a broad lipid repertoire associated to the endothelial cell protein C receptor (EPCR)

Abstract

Evidence is mounting that the nature of the lipid bound to the endothelial cell protein C receptor (EPCR) has an impact on its biological roles, as observed in anticoagulation and more recently, in autoimmune disease. Phosphatidylethanolamine and phosphatidylcholine species dominate the EPCR lipid cargo, yet, the extent of diversity in the EPCR-associated lipid repertoire is still unknown and remains to be uncovered. We undertook mass spectrometry analyses to decipher the EPCR lipidome, and identified species not yet described as EPCR ligands, such as phosphatidylinositols and phosphatidylserines. Remarkably, we found further, more structurally divergent lipids classes, represented by ceramides and sphingomyelins, both in less abundant quantities. In support of our mass spectrometry results and previous studies, high-resolution crystal structures of EPCR in three different space groups point to a prevalent diacyl phospholipid moiety in EPCR’s pocket but a mobile and ambiguous lipid polar head group. In sum, these studies indicate that EPCR can associate with varied lipid classes, which might impact its properties in anticoagulation and the onset of autoimmune disease.

Structural vulnerability in EPCR suggests functional modulation

“…Consequently, the nature and arrangement of the lipid in the new structure are of interest. In the novel structure, the electron density suggests that the lipid bound is phosphatidylethanolamine, which is consistent with previous reports 5 , 7 , 10 . Concerning the position of the lipid, we did not observe any relevant discrepancies with respect to previously reported EPCR structures (Supplementary Fig.…”

“…1 ). While the overall structure of the receptor adopts the canonical shape of EPCR 5 , 7 , i.e., a CD1/MHC-class I-like platform defined by two alpha helices on a β-sheet platform creating a tunnel for lipid binding, we noticed a striking conformational change in the α2 helix (Fig. 1 A–D).…”

The structural basis of the EPCR-APC complex induced biased PAR1 signaling

“…The extracellular part of the EPCR structure contains two parallel alpha-helices on top of six beta-sheet platform (figure 1B). A comprehensive analysis of the lipids, found in the EPCR groove, has been reported (45). Lipids bound to EPCR extracellular domain can modulate APC binding, and some lipids have been shown to prevent APC binding.…”

“…However, the nature of bound phospholipid has a strong effect on APC binding affinity. The lipid-binding groove of EPCR can accommodate many types of lipids (45) resulting in a lipid-dependent change of APC binding. Multiple γ- carboxyglutamic acid-bound Ca 2+ ions support binding conformation of the APC Gla-domain, but calcium does not participate directly in the EPCR-APC binding (44).…”

“…A comprehensive analysis of the lipids, found in the EPCR groove, has been reported (45). Lipids bound to EPCR extracellular domain can modulate APC binding, and some lipids have been shown to prevent APC binding.…”