How cholesterol homeostasis is regulated by plasma membrane cholesterol in excess of phospholipids

Abstract

How do cells sense and control their cholesterol levels? Whereas most of the cell cholesterol is located in the plasma membrane, the effectors of its abundance are regulated by a small pool of cholesterol in the endoplasmic reticulum (ER). The size of the ER compartment responds rapidly and dramatically to small changes in plasma membrane cholesterol around the normal level. Consequently, increasing plasma membrane cholesterol in vivo from just below to just above the basal level evoked an acute (<2 h) and profound (Ϸ20-fold) decrease in ER 3-hydroxy-3-methylglutarylCoA reductase activity in vitro. We tested the hypothesis that the sharply inflected ER response to cholesterol is governed by the thermodynamic activity (fugacity) of plasma membrane cholesterol. The following two independent measures of plasma membrane cholesterol activity in human red cells and fibroblasts were used: susceptibility to cholesterol oxidase and cholesterol transfer to cyclodextrin. Both indicators revealed a threshold at the physiologic set point of plasma membrane cholesterol. Incrementing the phospholipid compartment in the plasma membrane with lysophosphatidylcholine, previously shown to decrease cholesterol oxidase susceptibility, reduced the transfer of plasma membrane cholesterol to cyclodextrin and to the ER. Conversely, the membrane intercalator, n-octanol, increased cholesterol oxidation, transfer, and ER pool size, perhaps by displacing cholesterol from plasma membrane phospholipids. We conclude that the activity of the fraction of cholesterol in excess of other plasma membrane lipids sets the cholesterol level in the ER. Cholesterol-sensitive elements therein respond by nulling the active plasma membrane pool, thereby keeping the cholesterol matched to the other plasma membrane lipids.

Scrambling of Phospholipids Activates Red Cell Membrane Cholesterol

“…In particular, transient increments in ER cholesterol can rapidly reduce cell cholesterol levels through interaction with at least two cholesterol-regulated systems. These are the esterification of cholesterol via acyl-cholesterol acyltransferase and the proteolysis of hydroxymethylglutaryl-CoA reductase, leading to a prompt decrease in the rate of cholesterol biosynthesis (2). In addition, the expression of multiple genes for sterol accretion is adjusted by an ER sterol-sensing protein system (21,26).…”

“…Another indicator of membrane cholesterol activity is its rate of transfer to an aqueous acceptor, MBCD (2,13,19). We therefore tested the effects of increased cytoplasmic Ca ++ on this process.…”

“…In particular, evidence from model systems suggests that the exofacial PLs might interact more strongly with sterols than the endofacial species (11,12). According to the hypothesis outlined above (2,3), the activity of cholesterol in the inner leaflet might then be higher than that in the outer leaflet.…”

Sphingolipids and cellular cholesterol homeostasis. Effect of ceramide on cholesterol trafficking and HMG CoA reductase activity

“…The direct testing of this hypothesis was not possible using the ACAT activity, because exogenous ceramides strongly inhibit this enzyme in intact cells as well as in cell-free homogenates [6,24]. Therefore we sought another method to measure the enrichment of ER cholesterol, namely the suppression of HMGCR activity, which has also been shown to be exquisitely sensitive to ER cholesterol levels [17] [18]. The results presented in Fig.…”

“…However, exogenous ceramide is known to inhibit this enzyme both in cell-free systems and in intact cells [6,16], and therefore, we employed the inhibition of HMG CoA reductase (HMGCR) activity as a measure of ER cholesterol levels in response to altered ceramide levels in the cells. This enzyme has also been shown to respond rapidly to variations in plasma membrane concentrations [17] [18], showing that its inhibition can be used as an alternate measure of ER cholesterol. The results presented here show that both exogenous and endogenous ceramides modulate the HMGCR activity independent of the membrane SM levels.…”