title: Acid ceramidase (ASAH1) is a global regulator of steroidogenic capacity and adrenocortical gene expression.
abstract: In H295R human adrenocortical cells, ACTH rapidly activates ceramide (Cer) and sphingosine (SPH) turnover with a concomitant increase in SPH-1-phosphate secretion. These bioactive lipids modulate adrenocortical steroidogenesis, primarily by acting as second messengers in the protein kinase A/cAMP-dependent pathway. Acid ceramidase (ASAH1) directly regulates the intracellular balance of Cer, SPH, and SPH-1-phosphate by catalyzing the hydrolysis of Cer into SPH. ACTH/cAMP signaling stimulates ASAH1 transcription and activity, supporting a role for this enzyme in glucocorticoid production. Here, the role of ASAH1 in regulating steroidogenic capacity was examined using a tetracycline-inducible ASAH1 short hairpin RNA H295R human adrenocortical stable cell line. We show that ASAH1 suppression increases the transcription of multiple steroidogenic genes, including Cytochrome P450 monooxygenase (CYP)17A1, CYP11B1/2, CYP21A2, steroidogenic acute regulatory protein, hormone-sensitive lipase, 18-kDa translocator protein, and the melanocortin-2 receptor. Induced gene expression positively correlated with enhanced histone H3 acetylation at target promoters. Repression of ASAH1 expression also induced the expression of members of the nuclear receptor nuclear receptor subfamily 4 (NR4A) family while concomitantly suppressing the expression of dosage-sensitive sex reversal, adrenal hypoplasia critical region, on chromosome X, gene 1. ASAH1 knockdown altered the expression of genes involved in sphingolipid metabolism and changed the cellular amounts of distinct sphingolipid species. Finally, ASAH1 silencing increased basal and cAMP-dependent cortisol and dehydroepiandrosterone secretion, establishing ASAH1 as a pivotal regulator of steroidogenic capacity in the human adrenal cortex.
authors.affiliation: School of Biology, Georgia Institute of Technology, Atlanta, Georgia 30332-0230, USA.
authors.forename: Natasha C
authors.initials: NC
authors.lastname: Lucki
authors.forename: Sibali
authors.initials: S
authors.lastname: Bandyopadhyay
authors.forename: Elaine
authors.initials: E
authors.lastname: Wang
authors.forename: Alfred H
authors.initials: AH
authors.lastname: Merrill
authors.forename: Marion B
authors.initials: MB
authors.lastname: Sewer
mesh_terms: D000107:Acetylation; D055573:Acid Ceramidase / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D000302:Adrenal Cortex / Q000378:metabolism*; D000324:Adrenocorticotropic Hormone / Q000494:pharmacology; D053898:Biosynthetic Pathways / Q000235:genetics; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D002518:Ceramides / Q000378:metabolism; D000242:Cyclic AMP / Q000378:metabolism; D056765:Cyclin B2 / Q000378:metabolism; D003687:Dehydroepiandrosterone / Q000378:metabolism; D005786:Gene Expression Regulation; D055785:Gene Knockdown Techniques; D006657:Histones / Q000378:metabolism; D006801:Humans; D006854:Hydrocortisone / Q000378:metabolism; D018809:Proliferating Cell Nuclear Antigen / Q000378:metabolism; D011401:Promoter Regions, Genetic; D034622:RNA Interference; D018160:Receptors, Cytoplasmic and Nuclear / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction; D013107:Sphingolipids / Q000235:genetics / Q000378:metabolism; D013250:Steroid Hydroxylases / Q000235:genetics* / Q000378:metabolism; D013256:Steroids / Q000096:biosynthesis*; D014158:Transcription, Genetic; D051176:beta Catenin / Q000378:metabolism
chemical_list: D002518:Ceramides; D056765:Cyclin B2; D006657:Histones; D018809:Proliferating Cell Nuclear Antigen; D018160:Receptors, Cytoplasmic and Nuclear; D013107:Sphingolipids; D013256:Steroids; D051176:beta Catenin; D003687:Dehydroepiandrosterone; D000324:Adrenocorticotropic Hormone; D000242:Cyclic AMP; D013250:Steroid Hydroxylases; C526693:ASAH1 protein, human; D055573:Acid Ceramidase; D006854:Hydrocortisone