Quantitative analysis of the expression of ACAT genes in human tissues by real-time PCR.
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 beta-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.
Smith
Jeffery L
JL
Department of Biochemistry and Molecular Biology, University of Queensland, Brisbane 4072, Australia. jefferysmith@optusnet.com.au
Rangaraj
Kavitha
K
Simpson
Robert
R
Maclean
Donald J
DJ
Nathanson
Les K
LK
Stuart
Katherine A
KA
Scott
Shaun P
SP
Ramm
Grant A
GA
de Jersey
John
J
D004386:Duodenum / Q000378:metabolism; D020869:Gene Expression Profiling / Q000379:methods*; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000378:metabolism; D008099:Liver / Q000378:metabolism; D009928:Organ Specificity; D016133:Polymerase Chain Reaction / Q000379:methods; D012333:RNA, Messenger / Q000032:analysis; D002785:Sterol O-Acyltransferase / Q000235:genetics*; D014018:Tissue Distribution; D000097587:Sterol O-Acyltransferase 2
D012333:RNA, Messenger; D002785:Sterol O-Acyltransferase; D000097587:Sterol O-Acyltransferase 2; C493492:sterol O-acyltransferase 1