Molecular cloning, functional expression and tissue distribution of rat acyl-coenzyme A:cholesterol acyltransferase.
Acyl-coenzyme A:cholesterol acyltransferase (ACAT) is an enzyme catalyzing the intracellular formation of cholesteryl esters from free cholesterol and fatty acyl-CoA. In the present study, we cloned rat ACAT cDNA and determined its tissue distribution. Rat ACAT cDNA, having a coding region of 1635 bp with its deduced protein sequence of 545 amino acids and two typical motifs such as signature sequences and leucine heptad motif, showed 83, 92 and 90% identity with human, mouse, and hamster ACAT, respectively. Expression of rat ACAT cDNA in A293 cells and CHO cells resulted in a 3.0 to 3.5-fold increase in the enzyme activity. Among twelve tissues examined, ACAT activity was highest in adrenal followed by liver and intestine while that of aorta was extremely low. The mRNA level was also the highest in adrenal among four tissues examined. However, in contrast to its high ACAT activity, the liver mRNA level was extremely low (adrenal >> intestine > aorta >> liver). Consistent with mRNA levels, immunohistochemical analyses with a specific ACAT antibody detected significant ACAT signals in adrenal and intestine but a negligible signal in liver. These results indicate that adrenal most abundantly expresses ACAT in rat. Furthermore, rat liver showed a high ACAT activity but an extremely low ACAT mRNA and negligible immunohistochemical reactivity, suggesting the presence of a structurally different ACAT protein(s) in rat liver.
Matsuda
H
H
Department of Biochemistry, Kumamoto University School of Medicine, Honjo 2-2-1, Kumamoto 860-0811, Japan.
Hakamata
H
H
Kawasaki
T
T
Sakashita
N
N
Miyazaki
A
A
Takahashi
K
K
Shichiri
M
M
Horiuchi
S
S
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D003001:Cloning, Molecular; D006224:Cricetinae; D017931:DNA Primers / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics; D015870:Gene Expression; D006801:Humans; D007150:Immunohistochemistry; D051379:Mice; D008969:Molecular Sequence Data; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D017386:Sequence Homology, Amino Acid; D002785:Sterol O-Acyltransferase / Q000235:genetics* / Q000378:metabolism*; D014018:Tissue Distribution
D017931:DNA Primers; D018076:DNA, Complementary; D012333:RNA, Messenger; D002785:Sterol O-Acyltransferase