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(Arteriosclerosis, Thrombosis, and Vascular Biology. 2000;20:1769.)
© 2000 American Heart Association, Inc.

Atherosclerosis and Lipoproteins

Enrichment of Acyl Coenzyme A:Cholesterol O-Acyltransferase Near Trans-Golgi Network and Endocytic Recycling Compartment

Nadia Khelef; T. T. Soe; Oswald Quehenberger; Nanda Beatini; Ira Tabas; Frederick R. Maxfield

From the Department of Biochemistry (N.K., T.T.S., F.R.M.), Weill Medical College of Cornell University, New York, NY; Institut Pasteur (N.K.), Paris, France; the Department of Medicine (O.Q.), University of California at San Diego, La Jolla; and the Departments of Medicine (N.B., I.T.) and Anatomy & Cell Biology (I.T.), Columbia University, New York, NY.

Correspondence to Frederick R. Maxfield, PhD, Department of Biochemistry, Weill Medical College of Cornell University, 1300 York Ave, New York, NY 10021. E-mail frmaxfie{at}med.cornell.edu

Abstract—Acyl coenzyme A:cholesterol O-acyltransferase (ACAT) is the enzyme responsible for cholesterol esterification in macrophages leading to foam cell formation. The determination of its localization is a critical step in understanding its regulation by cholesterol. Using immunofluorescence and confocal microscopy, we previously showed that the enzyme colocalized with markers of the endoplasmic reticulum, but in addition, ACAT was found in an unidentified paranuclear site. In the present study, we further define the localization of paranuclear ACAT. First, we found that ACAT does not colocalize with sorting endosomes or late endosomes labeled with fluorescent ₂-macroglobulin. The paranuclear ACAT is close to the endocytic recycling compartment labeled with fluorescent transferrin. We also show that the paranuclear structure containing ACAT is very close to TGN38, a membrane protein of the trans-Golgi network (TGN), but farther from Gos28, a marker of cis, medial, and trans Golgi. After treatment with nocodazole, the central localization of ACAT did not colocalize with markers of the TGN. These data indicate that a significant fraction of ACAT resides in membranes that may be a subcompartment of the endoplasmic reticulum in proximity to the TGN and the endocytic recycling compartment. Because the TGN and the endocytic recycling compartment are engaged in extensive membrane traffic with the plasma membrane, esterification of cholesterol in these membranes may play an important role in macrophage foam cell formation during atherogenesis.

Key Words: cholesterol esterification • organelles • immunolocalization • acyl coenzyme A:cholesterol O-acyltransferase

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