Enrichment of Endoplasmic Reticulum with Cholesterol Inhibits Sarcoplasmic-Endoplasmic Reticulum Calcium ATPase-2b Activity in Parallel with Increased Order of Membrane Lipids

Abstract

Macrophages in advanced atherosclerotic lesions accumulate large amounts of unesterified, or "free," cholesterol (FC). FC accumulation induces macrophage apoptosis, which likely contributes to plaque destabilization. Apoptosis is triggered by the enrichment of the endoplasmic reticulum (ER) with FC, resulting in depletion of ER calcium stores, and induction of the unfolded protein response. To explain the mechanism of ER calcium depletion, we hypothesized that FC enrichment of the normally cholesterol-poor ER membrane inhibits the macrophage ER calcium pump, sarcoplasmic-endoplasmic reticulum calcium ATPase-2b (SERCA2b). FC enrichment of ER membranes to a level similar to that occuring in vivo inhibited both the ATPase activity and calcium sequestration function of SERCA2b. Enrichment of ER with ent-cholesterol or 14:0 -18:0 phosphatidylcholine, which possess the membrane-ordering properties of cholesterol, also inhibited SERCA2b. Moreover, at various levels of FC enrichment of ER membranes, there was a very close correlation between increasing membrane lipid order, as monitored by 16-doxyl-phosphatidycholine electron spin resonance, and SERCA2b inhibition. In view of these data, we speculate that SERCA2b, a conformationally active protein with 11 membrane-spanning regions, loses function due to decreased conformational freedom in FC-ordered membranes. This biophysical model may underlie the critical connection between excess cholesterol, unfolded protein response induction, macrophage death, and plaque destabilization in advanced atherosclerosis.

Lipid signalling in disease

“…The accumulation of excess lipid, in particular saturated fatty acids and sterols, might induce cellular damage through stress on the membranes of lipid-metabolizing organelles, especially the endoplasmic reticulum (ER) and possibly mitochondria [56][57][58] . Under these conditions, the ER activates a complex response system known as the unfolded protein response (UPR) to restore the functional integrity of the organelle 59 .…”

Not Just Fat: The Structure and Function of the Lipid Droplet

“…Elevation of the FC content in the membrane may lead to an increase in membrane order, which then decreases the conformational freedom of membrane proteins and disrupts seemingly unrelated functions such as nuclear pore complex formation (Li et al 2004;Hodge et al 2010). …”

Combinatorial pattern recognition receptor signaling alters the balance of life and death in macrophages

“…Thus, fucoidan and acetyl-LDL activate TLR4-MyD88 signaling only under conditions of thapsigargin treatment or FC enrichment, respectively. Based on our previous studies, we knew that at least one role of these treatments is their ability to activate the CHOP branch of the UPR (9,30). However, these treatments, like many other UPR activators, also block calcium reuptake from the cytoplasm into the ER lumen (9,30).…”

“…Based on our previous studies, we knew that at least one role of these treatments is their ability to activate the CHOP branch of the UPR (9,30). However, these treatments, like many other UPR activators, also block calcium reuptake from the cytoplasm into the ER lumen (9,30). Therefore, we sought to determine whether perturbations of cellular calcium also might be important.…”