Abstract
The synthesis of cholesterol esters by acyl-CoA:cholesterol acyltransferase (ACAT, EC 2.3.1.26) is an important component of cellular cholesterol homeostasis. Cholesterol ester formation also is hypothesized to be important in several physiologic processes, including intestinal cholesterol absorption, hepatic lipoprotein production, and macrophage foam cell formation in atherosclerotic lesions. Mouse tissue expression studies and the disruption of the mouse ACAT gene (Acact) have indicated that more than one ACAT exists in mammals and specifically that another enzyme is important in mouse liver and intestine. We now describe a second mammalian ACAT enzyme, designated ACAT-2, that is 44% identical to the first cloned mouse ACAT (henceforth designated ACAT-1). Infection of H5 insect cells with an ACAT-2 recombinant baculovirus resulted in expression of a approximately 46-kDa protein in cell membranes that was associated with high levels of cholesterol esterification activity. Both ACAT-1 and ACAT-2 also catalyzed the esterification of the 3beta-hydroxyl group of a variety of oxysterols. Cholesterol esterification activities for ACAT-1 and ACAT-2 exhibited different IC50 values when assayed in the presence of several ACAT-specific inhibitors, demonstrating that ACAT inhibitors can selectively target specific forms of ACAT. ACAT-2 was expressed primarily in mouse liver and small intestine, supporting the hypothesis that ACAT-2 contributes to cholesterol esterification in these tissues. The mouse ACAT-2 gene (Acact2) maps to chromosome 15 in a region containing a quantitative trait locus influencing plasma cholesterol levels. The identification and cloning of ACAT-2 will facilitate molecular approaches to understanding the role of ACAT enzymes in mammalian biology.
Highlights
The synthesis of cholesterol esters by acyl-CoA:cholesterol acyltransferase (ACAT, EC 2.3.1.26) is an important component of cellular cholesterol homeostasis
Each family member has a potential tyrosine phosphorylation site and a conserved serine residue that has been implicated in the catalytic activity of hamster ACAT-1 [32]
Prior studies, including the disruption of the mouse ACAT-1 gene (Acact) [15] and tissue expression studies of Acact [16], indicated that more than one ACAT exists in mammals
Summary
The synthesis of cholesterol esters by acyl-CoA:cholesterol acyltransferase (ACAT, EC 2.3.1.26) is an important component of cellular cholesterol homeostasis. Infection of H5 insect cells with an ACAT-2 recombinant baculovirus resulted in expression of a ϳ46-kDa protein in cell membranes that was associated with high levels of cholesterol esterification activity. Cholesterol is the predominant cellular sterol, and cholesterol esterification is catalyzed by the enzyme acyl-CoA:cholesterol acyltransferase (E.C. 2.3.1.26, ACAT) Because of ACAT’s apparently prominent role in cholesterol metabolism, a number of ACAT inhibitors have been developed for use as anti-atherosclerosis agents (reviewed in Ref. 10) Some of these act by inhibition of intestinal cholesterol absorption [10] or hepatic lipoprotein synthesis and secretion [11, 12], and others, as specific inhibitors of macrophage foam cell formation [13]
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