Expression Cloning of an Oxysterol 7α-Hydroxylase Selective for 24-Hydroxycholesterol

Abstract

The synthesis of 7␣-hydroxylated bile acids from oxysterols requires an oxysterol 7␣-hydroxylase encoded by the Cyp7b1 locus. As expected, mice deficient in this enzyme have elevated plasma and tissue levels of 25-and 27-hydroxycholesterol; however, levels of another major oxysterol, 24-hydroxycholesterol, are not increased in these mice, suggesting the presence of another oxysterol 7␣-hydroxylase. Here, we describe the cloning and characterization of murine and human cDNAs and genes that encode a second oxysterol 7␣-hydroxylase. The genes contain 12 exons and are located on chromosome 6 in the human (CYP39A1 locus) and in a syntenic position on chromosome 17 in the mouse (Cyp39a1 locus). CYP39A1 is a microsomal cytochrome P450 enzyme that has preference for 24-hydroxycholesterol and is expressed in the liver. The levels of hepatic CYP39A1 mRNA do not change in response to dietary cholesterol, bile acids, or a bile acid-binding resin, unlike those encoding other sterol 7␣-hydroxylases. Hepatic CYP39A1 expression is sexually dimorphic (female > male), which is opposite that of CYP7B1 (male > female). We conclude that oxysterol 7␣-hydroxylases with different substrate specificities exist in mice and humans and that sexually dimorphic expression patterns of these enzymes in the mouse may underlie differences in bile acid metabolism between the sexes.Two metabolic pathways that differ in their initial steps produce 7␣-hydroxylated bile acids (1). In one pathway, cholesterol (5-cholesten-3␤-ol) is first converted into 7␣-hydroxycholesterol (cholest-5-ene-3␤,7␣-diol) by the enzyme cholesterol 7␣-hydroxylase, which is encoded by the Cyp7a1 gene in mice. In the other pathway, cholesterol is first converted into one of several oxysterols prior to being 7␣-hydroxylated by oxysterol 7␣-hydroxylase, which is encoded by the Cyp7b1 gene. The 7␣-hydroxylated intermediates produced by these different initiating steps are subsequently converted into primary bile acids by a series of shared enzymes in the liver (2).Cholesterol 7␣-hydroxylase shows a marked preference for cholesterol as a substrate and is only weakly active against other sterols (3), whereas oxysterol 7␣-hydroxylase prefers 25-hydroxycholesterol (cholest-5-ene-3␤,25-diol) and 27-hydroxycholesterol (cholest-5-ene-3␤,27-diol) (4, 5). In agreement with the latter preference, Cyp7b1 knockout mice accumulate these two oxysterols in their plasma and tissues (6). The levels of the other major oxysterol, 24-hydroxycholesterol (cholest-5-ene-3␤,24-diol), are near normal in these animals (6). In contrast, a human with a complete absence of oxysterol 7␣-hydroxylase activity accumulated 24-hydroxycholesterol as well as 25-and 27-hydroxycholesterol in his plasma (7). These observations suggest that the human enzyme, unlike the mouse enzyme, may act on all three oxysterol substrates.The observed differences in the oxysterol accumulation patterns in mice and humans that express no oxysterol 7␣-hydroxylase are not due to differences in the biosynthesis of oxysterols, as cholester...

Characterization of the dehydroepiandrosterone (DHEA) metabolism via oxysterol 7α‐hydroxylase and 17‐ketosteroid reductase activity in the human brain

“…CYP39A1 represents another oxysterol 7a-hydroxylase with a strong preference for 24S-hydroxycholesterol (Li-Hawkins et al 2000a). Conversion into 24S-hydroxycholesterol is an important brain-specific mechanism for the elimination of cholesterol across the blood-brain barrier (Lü tjohann et al 1996;Bjö rkhem et al 1997Bjö rkhem et al , 1998Bjö rkhem and Eggertsen 2001).…”

“…Previous multiple northern blot analysis showed that CYP39A1 is exclusively expressed in murine liver tissue (Li-Hawkins et al 2000a). The present block-cycler RT-PCR analysis yielded mRNA expression of the human isozyme in human liver, prostate and, as shown here for the first time, to an apparently lesser also extent in the human CNS (Fig.…”

“…Thereafter, it was identified as the hepatic oxysterol 7a-hydroxylase . Another oxysterol 7a-hydroxylase with a preference for 24S-hydroxycholesterol has previously been identified in the liver and was designated CYP39A1 (Li-Hawkins et al 2000a). Moreover, several androgen 7a-hydroxylases were identified in rodents (Nagata et al 1987;Waxman et al 1990;Kurose et al 1998Kurose et al , 1999.…”

Epigenetic regulation of oxysterol formation

“…CYP39A1 encodes an oxysterol 7α-hydroxylase enzyme that appears to have a preference for 24S-hydroxycholesterol as a substrate [53]. It was originally cloned form the liver, although low levels have been detected in the eye [54,55] and brain [56].…”

“…It was originally cloned form the liver, although low levels have been detected in the eye [54,55] and brain [56]. A notable feature is the sexually dimorphic, with females expressing a higher level of both the mRNA and the protein [53].…”