Cholesterol is an important component of animal cell membranes, where it helps to establish proper fluidity and maintain the barrier between components within cells and the extracellular environment. In addition, cholesterol is the metabolic precursor of steroid hormones such as progesterone, estrogen, and testosterone that function in sexual development and the precursor of bile acids that solubilize dietary fats and fat-soluble vitamins such as vitamin A, vitamin D, and vitamin K, aiding their intestinal absorption. Cholesterol has been implicated in intracellular transport and cell signaling; in the brain, cholesterol is present at high levels in myelin membranes of white matter that provides insulation for efficient conduction of impulses.
Despite its essential role in various physiologic processes, cholesterol has acquired a maligned reputation. This stems from abundant evidence that high levels of low-density lipoprotein (LDL), the major carrier of cholesterol in blood, are associated with increased risk for development of atherosclerosis and coronary heart disease.
Animal cells acquire cholesterol from two sources: de novo synthesis through the action of more than 20 enzymes and uptake of cholesterol-rich LDL particles. To avoid the detrimental consequences associated with overproduction of cholesterol, cells must tightly regulate both the synthesis and uptake of this important molecule. The primary target of this regulation is HMG-CoA reductase, a key enzyme that sets the rate of cholesterol synthesis in cells. HMG-CoA reductase is also the target of statins, a group of drugs that are taken by 20 million Americans daily to lower levels of LDL cholesterol in the blood and reduce the incidence of coronary heart disease and heart attacks.
On average, statins both lower blood LDL-cholesterol levels and decrease the occurrence of heart attacks by 40 percent. Although statin therapy is a major medical advance, these drugs do not completely block cholesterol synthesis, and as a result, the reduction in heart attacks is not 100 percent. The limited effect of statins in reducing heart attacks is due to a dramatic increase in the amount of HMG-CoA reductase that cells produce in an attempt to restore cholesterol synthesis. This creates a situation in which higher and higher doses of statins are required to maintain their cholesterol-lowering effects.
Studies from my laboratory indicate that statins drive up levels of HMG-CoA reductase by blocking its normal degradation. We have discovered a set of proteins that are required for the degradation of HMG-CoA reductase, but these proteins are not active enough to destroy the enormous amounts of the enzyme that accumulate during statin treatment. Our hope is that by focusing on reactions that lead to the degradation of HMG-CoA reductase, we will develop novel therapies to counteract the statin-mediated increase in HMG-CoA reductase by stimulating the activity of proteins that destroy the enzyme. If successful, these new therapies could be used in combination with statins to completely block cholesterol synthesis and reduce the occurrence of heart attacks by much more than 40 percent.
The Regulated Degradative Pathway for HMG-CoA Reductase
HMG-CoA reductase is anchored to membranes of the endoplasmic reticulum (ER) through a hydrophobic N-terminal domain with eight membrane-spanning helices (Figure 1). The hydrophilic C-terminal domain of HMG-CoA reductase projects into the cytosol, where it exerts enzymatic activity. Our studies reveal that accumulation of certain types of sterols triggers the binding of HMG-CoA reductase to ER membrane proteins called Insig-1 and Insig-2. Some of these Insig molecules, in turn, are associated with gp78, a ubiquitin-ligating enzyme that mediates transfer of the small protein ubiquitin from the ubiquitin-conjugating enzyme Ubc7 to a pair of lysine residues in the membrane domain of HMG-CoA reductase.
Once ubiquitinated, HMG-CoA reductase is somehow extracted from the ER membrane and delivered to a large, cytosolic protease, called the 26S proteasome, for degradation (Figure 2). Extraction and/or proteasomal delivery occurs through a largely undefined mechanism that likely involves the gp78-bound ATPase VCP/p97 and its associated cofactors. In addition, extraction appears to be augmented by geranylgeraniol, a 20-carbon isoprenoid that is produced in the cholesterol biosynthetic pathway.
HMG-CoA reductase is the ideal model to understand the ER-associated degradation of proteins containing multiple membrane-spanning segments. The precise mechanism through which these types of proteins are recognized for degradation, extracted from ER membranes, and delivered to cytosolic proteasomes is not clear. One of our goals is to reconstitute sterol-induced degradation of HMG-CoA reductase in vitro and delineate mechanisms for its extraction from ER membranes and delivery to proteasomes. The sterol dependence of HMG-CoA reductase degradation is a valuable control that guards against artifactual degradation that may occur when various steps in the reaction are reconstituted in vitro.
An understanding of mechanisms for HMG-CoA reductase degradation will not only have important clinical implications for statin therapy but also provide insight into the degradation of other clinically important proteins with multiple membrane-spanning domains, such as mutant forms of CFTR (cystic fibrosis), connexin-32 (X-linked Charcot-Marie-Tooth disease), and polycystin-2 (autosomal-dominant polycystic kidney disease).
Grants from the National Institutes of Health, the W.M. Keck Foundation, and the American Heart Association provided partial support for this research.
As of May 30, 2012