In the human adrenal cortex, adrenocorticotropin (ACTH) activates steroid
hormone biosynthesis by acutely increasing cholesterol delivery to the mitochondria and
chronically up-regulating the transcription of steroidogenic genes (including CYP17).
Sphingolipids are a diverse family of phospholipids and glycolipids that mediate a wide
variety of cellular processes, including apoptosis, proliferation, and survival. Sterol
regulatory element binding proteins (SREBPs) are a family of transcription factors that
regulate genes that are involved in cholesterol biosynthesis and fatty acid metabolism. In
this study, we investigated the role of sphingolipids in ACTH-dependent steroidogenesis.
H295R human adrenocortical cells were treated with ACTH or dibutyryl cAMP (Bt2cAMP)
for various time periods and the content of several sphingolipid species was quantified
by mass spectrometry. Both ACTH and Bt2cAMP decreased cellular amounts of
sphingomyelin, ceramides, sphingosine (So) and sphingosine-1-phosphate (S1P).
However, both ACTH and Bt2cAMP increased the activity of sphingosine kinase and the
amounts of S1P released into the media. Both So and S1P increased CYP17 mRNA
expression and increased cortisol biosynthesis. This increase in CYP17 transcription
occurs by promoting SREBP binding to an SRE at -450/-436 basepairs upstream of the
transcription initiation site. Furthermore, chromatin immunoprecipitation (ChIP) assays
revealed that Bt2cAMP and S1P treatment results in an increase in acetylation of histone
H3 and SREBP1 binding to CYP17 promoter. Additionally, transient transfection studies
using wild type or mutated hCYP17 promoters and RNA interference (RNAi) assays
confirmed the role of SREBP1 in mediating the stimulatory effect of S1P on CYP17
transcription. In summary, our studies demonstrate a link between sphingolipid metabolism and ACTH-dependent steroidogenesis which requires the activation of
SREBP1 in human adrenal cortex. |