Identification and Characterization of a Stereospecific Human Enzyme That Catalyzes 9-cis-Retinol Oxidation
A POSSIBLE ROLE IN 9-cis-RETINOIC ACID FORMATION*
- James R. Mertz‡,
- Enyuan Shang§¶,
- Roseann Piantedosi‖,
- Shuanghong Wei**,
- Debra J. Wolgemuth§¶‡ and
- William S. Blaner‖**§§
- From the ‡Department of Cell Biology and Anatomical Sciences, City University of New York Medical School, New York, New York 10031 and the Departments of §Genetics and Development, ¶Obstetrics and Gynecology, and ‖Medicine, the **Institute of Human Nutrition, and the‡Columbia University Cancer Center, College of Physicians and Surgeons, Columbia University, New York, New York 10032
Abstract
All-trans- and 9-cis-retinoic acid are active retinoids for regulating expression of retinoid responsive genes, serving as ligands for two classes of ligand-dependent transcription factors, the retinoic acid receptors and retinoid X receptors. Little is known, however, regarding 9-cis-retinoic acid formation. We have obtained a 1.4-kilobase cDNA clone from a normalized human breast tissue library, which when expressed in CHO cells encodes a protein that avidly catalyzes oxidation of 9-cis-retinol to 9-cis-retinaldehyde. This protein also catalyzes oxidation of 13-cis-retinol at a rate approximately 10% of that of the 9-cis isomer but does not catalyze all-trans-retinol oxidation. NAD+ was the preferred electron acceptor for oxidation of 9-cis-retinol, although NADP+ supported low rates of 9-cis-retinol oxidation. The rate of 9-cis-retinol oxidation was optimal at pHs between 7.5 and 8. Sequence analysis indicates that the cDNA encodes a protein of 319 amino acids that resembles members of the short chain alcohol dehydrogenase protein family. mRNA for the protein is most abundant in human mammary tissue followed by kidney and testis, with lower levels of expression in liver, adrenals, lung, pancreas, and skeletal muscle. We propose that this cDNA encodes a previously unknown stereospecific enzyme, 9-cis-retinol dehydrogenase, which probably plays a role in 9-cis-retinoic acid formation.
Footnotes
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↵* This work was supported by grants from the National Institutes of Health (to D. J. W. and W. S. B.), a grant from the U. S. Department of Agriculture (to W. S. B.), a grant from the Chinese Academy of Sciences (to E. S.), and a seed grant (Professional Staff Congress, City University of New York) from the City University of New York (to J. R. M.).The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
The nucleotide sequence(s) reported in this paper has been submitted to the GenBank™/EMBL Data Bank with accession number(s) U89717.
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↵§§ To whom correspondence should be addressed: Dept. of Medicine, College of Physicians and Surgeons, Columbia University, 630 W. 168th St., New York, NY 10032. Tel.: 212-305-5429; Fax: 212-305-5384; E-mail:wsb2{at}columbia.edu.
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↵1 The abbreviations used are: RAR, retinoic acid receptor; RXR, retinoid X receptors; 9cRDH, 9-cis-retinol dehydrogenase; PBS, phosphate-buffered saline (10 mm sodium phosphate, 150 mm NaCl, pH 7.4); HPLC, high performance liquid chromatography; bp, base pair(s); kb, kilobase(s); CHO, Chinese hamster ovary.
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- Received January 21, 1997.
- Revision received March 4, 1997.
