Expression of cholesterol 7alpha-hydroxylase restores bile acid synthesis in McArdle RH7777 cells.
Bile acid synthesis involves several enzymes and occurs only in liver cells. The first and rate-determining step is catalyzed by cholesterol 7alpha-hydroxylase (cyp7a). McArdle RH7777 hepatoma cells do not synthesize bile acids and do not express the cyp7a gene. A synthetic cyp7a gene was stably expressed in this cell line to determine if restoration of cyp7a activity is sufficient to reconstitute the bile acid synthetic pathway. The transfected cells contained the recombinant cyp7a mRNA and the corresponding protein. Microsomes from recombinant cells converted cholesterol into 7alpha-hydroxycholesterol, indicating that the recombinant enzyme was active. Radiolabeled bile acids, originated from exogenously supplied radiolabeled cholesterol, were detected in the culture medium of recombinant cells. Thus, expression of cyp7a is sufficient in restoring bile acid synthesis in McArdle RH7777 cells. The results also show that the additional complement of enzymatic activities required to convert cholesterol into bile acids has remained active in this cell line.
Labonté
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Department of Biochemistry, University of Alberta, Edmonton, Canada.
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Agellon
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D000818:Animals; D001647:Bile Acids and Salts / Q000096:biosynthesis*; D002784:Cholesterol / Q000378:metabolism; D002790:Cholesterol 7-alpha-Hydroxylase / Q000235:genetics / Q000378:metabolism*; D015870:Gene Expression; D008114:Liver Neoplasms, Experimental / Q000235:genetics / Q000378:metabolism; D051381:Rats; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D014162:Transfection; D014407:Tumor Cells, Cultured
D001647:Bile Acids and Salts; D011994:Recombinant Proteins; D002784:Cholesterol; D002790:Cholesterol 7-alpha-Hydroxylase