Transport of cholesterol into mitochondria is rate-limiting for bile acid synthesis via the alternative pathway in primary rat hepatocytes.
Bile acid synthesis occurs mainly via two pathways: the "classic" pathway, initiated by microsomal cholesterol 7alpha-hydroxylase (CYP7A1), and an "alternative" (acidic) pathway, initiated by sterol 27-hydroxylase (CYP27). CYP27 is located in the inner mitochondrial membrane, where cholesterol content is very low. We hypothesized that cholesterol transport into mitochondria may be rate-limiting for bile acid synthesis via the "alternative" pathway. Overexpression of the gene encoding steroidogenic acute regulatory (StAR) protein, a known mitochondrial cholesterol transport protein, led to a 5-fold increase in bile acid synthesis. An increase in StAR protein coincided with an increase in bile acid synthesis. CYP27 overexpression increased bile acid synthesis by <2-fold. The rates of bile acid synthesis following a combination of StAR plus CYP27 overexpression were similar to those obtained with StAR alone. TLC analysis of (14)C-labeled bile acids synthesized in cells overexpressing StAR showed a 5-fold increase in muricholic acid; in chloroform-extractable products, a dramatic increase was seen in bile acid biosynthesis intermediates (27- and 7,27-hydroxycholesterol). High-performance liquid chromatography analysis showed that 27-hydroxycholesterol accumulated in the mitochondria of StAR-overexpressing cells only. These findings suggest that cholesterol delivery to the inner mitochondrial membrane is the predominant rate-determining step for bile acid synthesis via the alternative pathway.
D000818:Animals; D001647:Bile Acids and Salts / Q000096:biosynthesis*; D001692:Biological Transport; D053493:Cholestanetriol 26-Monooxygenase; D002784:Cholesterol / Q000378:metabolism*; D015870:Gene Expression; D022781:Hepatocytes / Q000378:metabolism* / Q000648:ultrastructure; D008297:Male; D008862:Microsomes, Liver / Q000201:enzymology; D008928:Mitochondria / Q000378:metabolism; D010750:Phosphoproteins / Q000235:genetics; D051381:Rats; D017207:Rats, Sprague-Dawley; D013250:Steroid Hydroxylases / Q000235:genetics / Q000378:metabolism; D000098682:Steroidogenic Acute Regulatory Protein
D001647:Bile Acids and Salts; D010750:Phosphoproteins; D000098682:Steroidogenic Acute Regulatory Protein; D002784:Cholesterol; D013250:Steroid Hydroxylases; D053493:Cholestanetriol 26-Monooxygenase