Cloning of a cDNA for a Second Retinol Dehydrogenase Type II

doi:10.1074/jbc.270.47.28408

Cloning of a cDNA for a Second Retinol Dehydrogenase Type II

EXPRESSION OF ITS mRNA RELATIVE TO TYPE I (*)

From the Department of Biochemistry, School of Medicine and Biomedical Sciences, State University of New York at Buffalo, Buffalo, New York 14214

^§ To whom correspondence should be addressed:
140 Farber Hall, School of Medicine and Biomedical Sciences, SUNY-Buffalo, Buffalo, NY 14214.
Tel.: 716-829-2726; Fax: 716-829-2725.

Abstract

A retinol dehydrogenase, RoDH(I), which recognizes holo-cellular retinol-binding protein (CRBP) as substrate, has been cloned, expressed, and identified as a short-chain dehydrogenase/reductase (Chai, X., Boerman, M. H. E. M., Zhai, Y., and Napoli, J. L.(1995) J. Biol. Chem. 270, 3900-3904). This work reports the cloning and expression of a cDNA encoding a RoDH isozyme, RoDH(II). The predicted amino acid sequence verifies RoDH(II) as a short-chain dehydrogenase/reductase, 82% identical with RoDH(I). RoDH(II) recognized the physiological form of retinol as substrate, CRBP, with a K of 2 mM. Similar to microsomal RoDH and RoDH(I), RoDH(II) had higher activity with NADP rather than NAD, was stimulated by ethanol and phosphatidyl choline, was not inhibited by the medium-chain alcohol dehydrogenase inhibitor 4-methylpyrazole, but was inhibited by phenylarsine oxide and the short-chain dehydrogenase/reductase inhibitor carbenoxolone. Northern blot analysis detected RoDH(I) and RoDH(II) mRNA only in rat liver, but RNase protection assays revealed RoDH(I) and RoDH(II) mRNA in kidney, lung, testis, and brain. These data indicate that short-chain dehydrogenases/reductase isozymes expressed tissue-distinctively catalyze the first step of retinoic acid biogenesis from the physiologically most abundant substrate, CRBP.

Footnotes

↵* This work was supported by National Institutes of Health Grant DK36870. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by 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(TM)/EMBL Data Bank with accession number(s) RODHII and U33500[GenBank].
↵⁽⁾ The abbreviations used are:

RA

all-trans-retinoic acid

CRBP

cellular retinol-binding protein type I

PAO

phenylarsine oxide

PCR

polymerase chain reaction

10Ksup

10,000 × g supernatant of a cell homogenate.
- Received June 26, 1995.
- Revision received August 16, 1995.