Formation and degradation of dihydrotestosterone by recombinant members of the rat 3 beta-hydroxysteroid dehydrogenase/delta 5-delta 4 isomerase family.
The structures of cDNA clones encoding four members of the rat 3 beta-hydroxysteroid dehydrogenase/delta 5-delta 4 isomerase (3 beta-HSD) family were characterized. The rat type I, type II and the novel type IV are genuine NAD+/H-dependent 3 beta-HSD isoenzymes. On the other hand, the liver-specific type III protein is a specific 3-keto-reductase (3-KSR) that catalyzes the conversion of 5 alpha-androstane-3-one-17 beta-ol (DHT) and 5 alpha-androstane 3,17-dione (A-dione) into their 3 beta-hydroxy metabolites. The aim of the present study was to further characterize the enzymatic properties of rat types I, III and IV, especially their role in the formation and degradation of DHT after transient expression in intact human HeLa cervical carcinoma, JEG-3 choriocarcinoma or SW-13 adrenal cortex adenocarcinoma cells in culture. The expressed type III 3-KSR in intact HeLa cells catalyzed the reduction of DHT into 3 beta-diol, whereas expression of type I 3 beta-HSD in these cell lines had no significant effect on the basal conversion of DHT into 3 beta-diol, but it did increase the formation of DHT from 3 beta-diol. A-dione is the predominant product obtained when DHT and 5 alpha-androstane-3 beta, 17 beta-diol (3 beta-diol) are used as substrates in intact JEG-3 and SW-13 cells transfected with rat type I 3 beta-HSD. Furthermore, this predominant 17 beta-HSD activity was also observed in SW-13 cells transfected with the novel rat type IV 3 beta-HSD. The predominance of this 'secondary' 17 beta-HSD activity is also reflected in HeLa cells transfected with type I 3 beta-HSD by the deduced predominant pathway 3 beta-diol-->DHT-->5 alpha-androstane-3 alpha,17 beta-diol (3 alpha-diol)-->androsterone (ADT), in which formation of 3 alpha-HSD activity of HeLa cells, whereas the other reactions are catalyzed by the type I 3 beta-HSD isoenzyme. This observation thus demonstrates that rat type I 3 beta-HSD may also catalyze the conversion of 3 alpha-diol into ADT through its intrinsic 17 beta-HSD activity. The predominant metabolic pathways observed in the present study could be attributed to preponderant bioavailability of NAD+ and NADPH in the intact transfected cells used.
Medical Research Council Group in Molecular Endocrinology, CHUL Research Center, Quebec, Canada.
R
R
Sanchez
Y
Y
de Launoit
F
F
Durocher
A
A
Bélanger
F
F
Labrie
J
J
Simard
D015067:17-Hydroxysteroid Dehydrogenases / Q000378:metabolism; D015096:3-Hydroxysteroid Dehydrogenases / Q000378:metabolism; D000230:Adenocarcinoma; D000306:Adrenal Cortex Neoplasms; D000818:Animals; D002822:Choriocarcinoma; D013196:Dihydrotestosterone / Q000378:metabolism*; D018014:Gene Transfer Techniques; D006367:HeLa Cells; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism; D007700:Kinetics; D009097:Multienzyme Complexes / Q000235:genetics / Q000378:metabolism*; D009243:NAD / Q000494:pharmacology; D009249:NADP / Q000494:pharmacology; D011375:Progesterone Reductase / Q000235:genetics / Q000378:metabolism*; D051381:Rats; D011994:Recombinant Proteins / Q000378:metabolism; D013251:Steroid Isomerases / Q000235:genetics / Q000378:metabolism*; D014407:Tumor Cells, Cultured
C021425:3 beta-hydroxysteroid oxidoreductase-delta(5) 3-ketosteroid isomerase; D007527:Isoenzymes; D009097:Multienzyme Complexes; D011994:Recombinant Proteins; D013196:Dihydrotestosterone; D009243:NAD; D009249:NADP; D015067:17-Hydroxysteroid Dehydrogenases; D015096:3-Hydroxysteroid Dehydrogenases; C031298:3-ketosteroid reductase; D011375:Progesterone Reductase; D013251:Steroid Isomerases