Cholesterol binding, efflux, and a PDZ-interacting domain of scavenger receptor–BI mediate HDL-initiated signaling

Abstract

The binding of HDL to scavenger receptor-BI (SR-BI) mediates cholesterol movement. HDL also induces multiple cellular signals, which in endothelium occur through SR-BI and converge to activate eNOS. To determine the molecular basis of a signaling event induced by HDL, we examined the proximal mechanisms in HDL activation of eNOS. In endothelial cells, HDL and methyl-β-cyclodextrin caused comparable eNOS activation, whereas cholesterol-loaded methyl-β-cyclodextrin had no effect. Phosphatidylcholine-loaded HDL caused greater stimulation than native HDL, and blocking antibody against SR-BI, which prevents cholesterol efflux, prevented eNOS activation. In a reconstitution model in COS-M6 cells, wild-type SR-BI mediated eNOS activation by both HDL and small unilamellar vesicles (SUVs), whereas the SR-BI mutant AVI, which is incapable of efflux to SUV, transmitted signal by only HDL. In addition, eNOS activation by methyl-β-cyclodextrin was SR-BI dependent. Studies of mutant and chimeric class B scavenger receptors revealed that the C-terminal cytoplasmic PDZ-interacting domain and the C-terminal transmembrane domains of SR-BI are both necessary for HDL signaling. Furthermore, we demonstrated direct binding of cholesterol to the C-terminal transmembrane domain using a photoactivated derivative of cholesterol. Thus, HDL signaling requires cholesterol binding and efflux and C-terminal domains of SR-BI, and SR-BI serves as a cholesterol sensor on the plasma membrane. IntroductionCirculating levels of HDL are inversely related to the incidence of atherosclerosis, and clinical trials with agents that increase HDL show that elevations in the lipoprotein level decrease cardiovascular disease risk (1, 2). Thus, HDL is not merely a marker but a potent mediator of cardiovascular health. The HDL particle consists of a shell of apolipoproteins (mainly apolipoproteins apoA-I and apoA-II), phospholipids, and cholesterol surrounding a lipid core of triglycerides and cholesteryl esters. Atheroprotection by HDL is in part due to the action of HDL in delivering extrahepatic cholesterol to the liver for excretion into bile in a process known as reverse cholesterol transport. Reverse cholesterol transport involves the binding of HDL to hepatic scavenger receptor-BI (SR-BI), an 82-kDa cell surface glycoprotein. SR-BI is also expressed at high levels in the adrenal glands and gonads, where it participates in regulating steroid hormone synthesis (3, 4).

Circulating cardiovascular disease risk factors and signaling in endothelial cell caveolae

“…Furthermore, heterologous expression experiments using COS-M6 cells have revealed that wild-type SR-BI mediates eNOS activation by both HDL and small unilamellar vesicles, whereas the SR-BI mutant AVI, which is incapable of efflux to small unilamellar vesicles [30], signals only in response to HDL. Moreover, eNOS activation by both HDL and methyl-h-cyclodextrin is SR-BI-dependent [31]. These findings indicate that signaling by HDL requires cholesterol flux, that the apolipoprotein and phospholipid components of HDL are sufficient to induce signal, and that SR-BI functions as a plasma membrane cholesterol sensor.…”

“…Since these domains are not involved in regulating cholesterol flux, cholesterol movement alone is not sufficient to initiate signaling. Further studies showed that cell cholesterol binds directly to the C-terminal transmembrane domain of SR-BI [31]. This is a characteristic of other cholesterol sensing proteins such as SCAP, which undergoes a conformational change with an alteration in cholesterol binding that modifies its function [32].…”

Recent Advances on the Antiatherogenic Effects of HDL-Derived Proteins and Mimetic Peptides

“…Importantly, only sHDL containing apoA-I and phospholipids, prevent TNF -induced upregulation of adhesion molecules; neither phospholipid vesicles nor lipid-free apoA-I were found to cause the same inhibitory effect [55]. It has been also demonstrated that sHDL can downregulate IL-6 expression [56] and stimulate endothelial nitric oxide synthase (eNOS) activation in cultured human endothelial cells [57]. The HDL-induced eNOS activation involves the binding of apoA-I to the scavenger receptor-BI (SR-BI) [58].…”

Update on HDL Receptors and Cellular Cholesterol Transport

“…Finally, we would be remiss not to mention that a significant body of evidence suggests SR-BI-mediated cholesterol efflux plays a critical role in signaling mechanisms that improve endothelial function (e.g., activation of endothelial nitric oxide synthase) [19]. This has been recently reviewed by Saddar et al [20].…”