Regulation of 3 beta-hydroxysteroid dehydrogenase/delta 5----4-isomerase expression by adrenocorticotropin in bovine adrenocortical cells.
In the steroidogenic pathway, 3 beta-hydroxysteroid dehydrogenase-isomerase (3 beta HSD) catalyzes the formation of hormonally active delta 4-3-ketosteroids from delta 5-3 beta-hydroxysteroids. In the present study the regulation of 3 beta HSD by ACTH action on bovine adrenocortical (BAC) cells in primary culture was evaluated. Western blot analysis was accomplished using an antibody against human placental 3 beta HSD. The relative molecular mass of 3 beta HSD in these cells was 45K, which was similar to that in human placenta. A significant effect of ACTH was not detected until day 6 of culture due to the high basal levels of the enzyme in BAC cells. Treatment of cells with ACTH on day 8 of culture resulted in a marked increase in the amount of 3 beta HSD protein, and this effect was correlated directly with enzymatic activity. The effects of ACTH were time and dose dependent, with an increase detectable only after 48 h of treatment; the maximal response was obtained with 10(-9) M ACTH. As demonstrated by Northern analysis, ACTH action was manifested by increasing the steady state level of 3 beta HSD mRNA. A human 3 beta HSD cDNA probe, which was used in this study, hybridized to a 1.7-kilobase species of BAC RNA. The effects of ACTH on 3 beta HSD activity and increases in 3 beta HSD protein and mRNA in BAC cells were mimicked by treatment with (Bu)2cAMP. The findings of this study suggest that ACTH controls 3 beta HSD gene expression in BAC cells by a cAMP-dependent mechanism similar to that involved in the expression of steroid hydroxylase genes. However, because the different stabilities of 3 beta HSD and hydroxylase proteins and/or mRNAs may play a critical role in determining the zone-specific steroids secreted from the adrenal cortex, other cAMP-dependent or independent regulatory mechanisms may also be important in regulating the expression of adrenal 3 beta HSD.
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Cecil H. and Ida Green Center for Reproductive Biology Sciences, Department of Obstetrics and Gynecology, University of Texas Southwestern Medical Center, Dallas 75235-9051.
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Milewich
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Mason
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D000302:Adrenal Cortex / Q000187:drug effects / Q000201:enzymology* / Q000378:metabolism; D000324:Adrenocorticotropic Hormone / Q000494:pharmacology*; D000818:Animals; D003994:Bucladesine / Q000494:pharmacology; D002417:Cattle; D002478:Cells, Cultured; D003513:Cycloheximide / Q000494:pharmacology; D015870:Gene Expression / Q000187:drug effects; D006854:Hydrocortisone / Q000378:metabolism; D007700:Kinetics; D009097:Multienzyme Complexes / Q000096:biosynthesis / Q000235:genetics*; D011375:Progesterone Reductase / Q000096:biosynthesis / Q000235:genetics*; D012333:RNA, Messenger / Q000187:drug effects / Q000378:metabolism; D013251:Steroid Isomerases / Q000096:biosynthesis / Q000235:genetics*
C021425:3 beta-hydroxysteroid oxidoreductase-delta(5) 3-ketosteroid isomerase; D009097:Multienzyme Complexes; D012333:RNA, Messenger; D003994:Bucladesine; D000324:Adrenocorticotropic Hormone; D003513:Cycloheximide; D011375:Progesterone Reductase; D013251:Steroid Isomerases; D006854:Hydrocortisone