Immunolocalization of a novel cholesteryl ester hydrolase in the endoplasmic reticulum of murine and human hepatocytes

We have recently purified a cholesteryl ester hydrolase (CEH) from rat live r microsomes. Antibodies raised against the purified protein specifically r eacted with a 106-kd protein and neutralized 90% of the CEH activity of rat liver microsomes (J Lipid Res 1999;40:715-725). In this work we have used the anti-CEH antibody to study both the subcellular distribution of the pro tein in hepatocytes as well as its tissue-specific expression in rat. Weste rn blotting of subcellular fractions obtained from isolated rat hepatocytes revealed that the immunoreactive 106-kd CEH was exclusively localized in m icrosomes. The antibody also recognized a 106-kd protein in microsomes from mouse and human liver but not from rat nonparenchymal liver cells. Confoca l microscopy of HepG2 cells revealed that CEH immunoreactive material coloc alized with calnexin, a marker of the endoplasmic reticulum. Furthermore, h igh-resolution immunoelectron microscopy of rat liver thin sections exclusi vely localized the CEH immunoreactivity to the endoplasmic reticulum of the hepatocyte. No CEH immunoreactivity was observed in microsomes derived fro m adrenal glands, ovaries, testis, pancreas, intestine, white adipose tissu e, mammary gland, lung, spleen, brain, aorta, and macrophages. We report a CEH localized to the endoplasmic reticulum, erCEH, in the mammalian hepatoc yte. The subcellular localization and tissue-restricted pattern of expressi on of erCEH suggests that it might have unique functions in liver cholester ol metabolism.