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Journal of Lipid Research, Vol. 45, 686-696, April 2004
Copyright © 2004 by American Society for Biochemistry and Molecular Biology

Quantitative analysis of the expression of ACAT ¹ genes in human tissues by real-time PCR²

Jeffery L. Smith^3,*,, Kavitha Rangaraj^*, Robert Simpson^*, Donald J. Maclean^*, Les K. Nathanson, Katherine A. Stuart, Shaun P. Scott^**, Grant A. Ramm^** and John de Jersey^*

^* Department of Biochemistry and Molecular Biology, University of Queensland, Brisbane 4072, Australia
Department of Surgery, University of Queensland, Brisbane 4072, Australia
Department of Gastroenterology and Hepatology, Princess Alexandra Hospital, Brisbane 4102, Australia
^** The Queensland Institute of Medical Research, Brisbane 4006, Australia

³ To whom correspondence should be addressed. e-mail: jefferysmith{at}optusnet.com.au

ACAT (also called sterol o-acyltransferase) catalyzes the esterification of cholesterol by reaction with long-chain acyl-CoA derivatives and plays a pivotal role in the regulation of cholesterol homeostasis. Although two human ACAT genes termed ACAT-1 and ACAT-2 have been reported, prior research on differential tissue expression is qualitative and incomplete. We have developed a quantitative multiplex assay for each ACAT isoform after RT treatment of total RNA using TaqMan real-time quantitative PCR normalized to ß-actin in the same reaction tube. This enabled us to calculate the relative abundance of transcripts in several human tissues as an ACAT-2/ACAT-1 ratio. In liver (n = 17), ACAT-1 transcripts were on average 9-fold (range, 1.7- to 167-fold) more abundant than ACAT-2, whereas in duodenal samples (n = 10), ACAT-2 transcripts were on average 3-fold (range, 0.39- to 12.2-fold) more abundant than ACAT-1. ACAT-2 was detected for the first time in peripheral blood mononuclear cells. Interesting differences in ACAT-2 mRNA expression were evident in subgroup analysis of samples from different sources.

These results demonstrate quantitatively that ACAT-1 transcripts predominate in human liver and ACAT-2 transcripts predominate in human duodenum and support the notion that ACAT-2 has an important regulatory role in liver and intestine.

Abbreviations: PBMC, peripheral blood mononuclear cells; RT-qPCR, RT treatment of RNA followed by quantitative real-time PCR

Supplementary key words acyl-coenzyme A:cholesterol acyltransferase • atherosclerosis • cholesterol metabolism • duodenum • gallstones • intestine • liver • sterol o-acyltransferase

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