Oxysterol pathway

This is a mitochondrial enzyme that was characterized on the basis of two rather divergent catalytic activities, the 25-hydroxylation of vitamin Dj (Figure 10.16) and the oxidation of cholesterol at the C27 position (Figure 10.18). Thus, the enzyme bridges between hormone (vitamin D) and oxysterol pathways, and the clinical relevance of P450 27A1 is considerable. 

The occurrence of side chain oxysterols has been reported in plants, animals, and microorganisms (58,59). In this report, the evolution of oxysterols resulting from primary metabolism via the oxysterol pathway" will be discussed, this includes sterols oxygenated in the side chain at carbon-24 and/or-25 (Figure 2). 

Also included in this report, are two documented alternative biochemical pathways which are known to introduce oxygen functionaly at C-25 (and C-26) directly without using the oxysterol pathway . In the first, sterols may be hydroxylated at C-25 in the liver during bile acid biosynthesis (1). In the second, vitamin D3 is known to be hydroxylated directly at C-25 in mammalian systems (1). These documented instances occur in the animal kingdom. Plant metabolism, especially the biosynthesis of secondary plant metabolites, has not been examined in sufficient detail to indicate the extent to which direct introduction of a hydroxyl group at C-25 might occur. However, a great deal of evidence is available to indicate that many of these side chain oxysterols are produced via the oxysterol pathway . 

Ample evidence has been presented to show a clear lineage of C-24 and/or 25 oxysterol biosynthesis in the biosphere via the "oxysterol pathway . Those results would seem to indicate the evolution of this pathway from lower organisms and finally its occurrence and function in mammalian systems. Certainly, we are the fortunate final recipients of this evolutionary event. 

Harada, K., Ishibashi, S., Miyashita, T, Osuga, J., Yagyu, H., Ohashi, K., Yazaki, Y., and Yamada, N., 1997, Bcl-2 inhibits oxysterol-induced apoptosis through suppressing CPP32-mediated pathway, FEBS Lett. 411 63-66. 

Formation of mevalonate. The conversion of acetyl CoA to acetoacetyl CoA and then to 3-hydroxy-3-methylglutaryl CoA (3-HMG CoA) corresponds to the biosynthetic pathway for ketone bodies (details on p. 312). In this case, however, the synthesis occurs not in the mitochondria as in ketone body synthesis, but in the smooth endoplasmic reticulum. In the next step, the 3-HMG group is cleaved from the CoA and at the same time reduced to mevalonate with the help of NADPH+H 3-HMG CoA reductase is the key enzyme in cholesterol biosynthesis. It is regulated by repression of transcription (effectors oxysterols such as cholesterol) and by interconversion... 

Cholesterol is an important structural component of cellular membranes, where it plays a role in modulating membrane fluidity and phase transitions, and, together with sphingomyelin, forms lipid rafts or caveolae, which are sites where proteins involved in diverse signaling pathways become concentrated. Furthermore, cholesterol is a precursor of oxysterols, steroid hormones, and bile acids. 

The initial steps in BA synthesis are characterised by the introduction of a hy-droxylic group in the la position, or in position 27, followed by another in the la position into the cholesterol nucleus. Both synthetic pathways (the neutral and the acidic pathways) possess a distinct microsomal 7-oxysterol hydroxylase, which is regulated by different genes. The most recently described disorder of BA synthesis is cholesterol 7a-hydroxylase deficiency, in which their decreased production through the classical pathway is partially balanced by activation of the alternative pathway. Cholesterol levels increase in the liver, with a consequent low-density lipoprotein hypercholesterolemia, and cholesterol gallstones may result, although there is no liver disease. In contrast, a defect in the conversion of 27-hydroxy-cholesterol to la,27-dihydroxy-cholesterol due to deficiency of the oxysterol 7a-hydroxylase specific for the alternate pathway, causes severe neonatal liver disease [8]. 

Janowski BA, Willy PJ, Devi TR, Falck JR, Mangelsdorf DJ. An oxysterol signaling pathway mediated by the nuclear receptor LXR alpha. Nature 1996 383 728-731. 

J. L. et al. (1997) Activation of the nuclear receptor LXR by oxysterols defines a new hormone response pathway. The Journal of Biological Chemistry, 272 (6), 3137-3140. 

For example, hepatocytes express LXR (liver X receptor), a nuclear receptor that senses the levels of oxysterols. When cellular cholesterol Increases in the liver, oxysterols are generated and activate LXR. Activated LXR stimulates the expression of cholesterol 7ct-hydroxylase, the key rate-limiting enzyme In the hepatic conversion of cholesterol Into bile acids, a major pathway for disposing of excess cholesterol from the body. LXR also stimulates the expression of the ABC proteins that export cholesterol Into the bile (ABCG5/8) or onto lipoproteins in the blood (ABCAl). In... 

In the nuclear-receptor pathway, transcription factors in the cytosol are activated by intracellular lipids (e.g., high levels of oxysterols or bUe acids). The ligand-transcription factor complex then enters the nucleus and regulates the expression of specific target genes that participate in feedback regulation of the synthesis, transport, and catabolism of lipids. 

Fig. 1. Overview of the metabolic and transport pathways that control cholesterol levels in mammalian cells. Cholesterol is synthesized from acetyl-CoA and the four key enzymes that regulate cholesterol synthesis are indicated. Cells also obtain cholesterol by uptake and hydrolysis of LDL s cholesteryi esters (CE). End products derived from cholesterol or intermediates in the pathway include bile acids, oxysterols, cholesteryi esters, and non-steroidal isoprenoids. ACAT, acyl-CoA cholesterol acyltransferase.

Essential non-steroidal isoprenoids, such as dolichol, prenylated proteins, heme A, and isopentenyl adenosine-containing tRNAs, are also synthesized by this pathway. In extrahepatic tissues, most cellular cholesterol is derived from de novo synthesis [3], whereas hepatocytes obtain most of their cholesterol via the receptor-mediated uptake of plasma lipoproteins, such as low-density lipoprotein (LDL). LDL is bound and internalized by the LDL receptor and delivered to lysosomes via the endocytic pathway, where hydrolysis of the core cholesteryl esters (CE) occurs (Chapter 20). The cholesterol that is released is transported throughout the cell. Normal mammalian cells tightly regulate cholesterol synthesis and LDL uptake to maintain cellular cholesterol levels within narrow limits and supply sufficient isoprenoids to satisfy metabolic requirements of the cell. Regulation of cholesterol biosynthetic enzymes takes place at the level of gene transcription, mRNA stability, translation, enzyme phosphorylation, and enzyme degradation. Cellular cholesterol levels are also modulated by a cycle of cholesterol esterification mediated by acyl-CoA cholesterol acyltransferase (ACAT) and hydrolysis of the CE, by cholesterol metabolism to bile acids and oxysterols, and by cholesterol efflux. 

Squalene is converted into the first sterol, lanosterol, by the action of squalene epoxidase and oxidosqualene lanosterol cyclase. The catalytic mechanism for the cyclase s four cyclization reactions was revealed when the crystal stmcture of the human enzyme was obtained (R. Thoma, 2004). Oxidosqualene lanosterol cyclase is considered an attractive target for developing inhibitors of the cholesterol biosynthetic pathway because its inhibition leads to the production of 24,25-epoxycholesterol (M.W. Huff, 2005). This oxysterol is a potent ligand activator of the liver X receptor (LXR) and leads to expression of several genes that promote cellular cholesterol efflux, such as ABCAl, ABCG5, and ABCG8 (Section 4.1). Thus, inhibitors of oxidosqualene lanosterol cyclase could be therapeutically advantageous because they would reduce cholesterol synthesis and promote cholesterol efflux (M.W. Huff, 2005). 

Isoprenoid synthesis is regulated by sterol and non-sterol components of the biosynthetic pathway, oxysterols, and also by physiological factors. The cholesterol content of the... 

Fig. 4. The bile acid biosynthetic pathways. The classical pathway operates entirely in the liver and cholesterol 7a-hydroxylase (CYP7A1) initiates the pathway. In other tissues, the entry of cholesterol into the alternate pathways is facilitated by sterol 27-hydroxylase (CYP27A1), cholesterol 24-hydroxylase (CYP46A1), and cholesterol 25-hydroxylase (CH25H). The oxysterols generated by these enzynies are 7a-hydroxylated by oxysterol 7a-hydroxylases CYP7B1 and CYP39A1, and the products enter the latter steps of the classical pathway.

The existence of an alternate pathway for the synthesis of bile acids was suspected because it was possible for oxysterols to be converted into bile acids (N. Wachtel, 1968). It is now recognized that a variety of oxysterols produced by an assortment of cell types can be converted into bile acids. The production of these oxysterols is catalyzed by several sterol hydroxylases sterol 27-hydroxylase (CYP27A1) (J.J. Cali, 1991), cholesterol 25-hydroxylase (CH25H) (E.G. Lund, 1998), and cholesterol 24-hydroxylase (CYP46A1) (E.G. Lund, 1999). Cholesterol 25-hydroxylase is not a cytochrome P-450 monooxygenase, unlike the two other enzymes. Almost all of the 24-hydroxycholesterol that ends up in the liver originates from the brain, and it has been suggested that the production of... 

Fig. 7. Induction of Cyp7al gene expression by oxysterol-activated LXRa RXRa. Both the bile acid and cholesterol biosynthetic pathways generate oxysterols. The binding site of LXRaiRXRa in the Cyp7al gene promoter is a DR-4 element (a direct repeat of the hexanucleotide hormone response element separated by 4 nt).

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