Upregulation of human chorionic gonadotrophin-induced steroidogenic acute regulatory protein by insulin-like growth factor-I in rat Leydig cells.
ABSTRACT Insulin-like growth factor-I (IGF-I) plays an essential role in reproductive function. Leydig cells express specific IGF-I receptors, and IGF-I enhances human chorionic gonadorphin (hCG)-induced testosterone formation. In the present study, we evaluate the effect of IGF-I on the gene expression and protein levels of steroidogenic acute regulatory protein (StAR), the rate-limiting step in steroidogenesis. StAR mRNA is expressed in rat Leydig cells as two major transcripts of 3.8 and 1.7 kb. StAR mRNA levels (both 3.8 and 1.7 kb) were markedly induced about 20-fold by hCG (10 ng/mL). Concomitant addition of IGF-I (50 or 100 ng/mL) and hCG (10 ng/mL) resulted in significant increases in StAR and cytochrome P450 side-chain cleavage (P450scc) mRNA levels, whereas lower doses of IGF-I (1 or 10 ng/ mL) had small effects. Synergistic effects of IGF-I and hCG on StAR mRNA levels were confirmed by ribonuclease protection assay (RPA). IGF-I (100 ng/mL) enhanced hCG- and 20 OH-cholesterol + hCG-induced testosterone formation, whereas the conversions of pregnenolone, 17-OH pregnenolone, dehydroepiandrosterone, and androstenedione to testosterone were not affected. This suggests that the major effect of IGF-I is at the steps of StAR and P450scc, whereas other steroidogenic enzymes are not affected. To evaluate whether increased StAR mRNA levels induced by IGF-I and hCG are associated with increased StAR protein levels, we carried out Western blot analyses. Basal StAR protein levels were low after 24 h in culture. hCG (10 ng/mL) increased StAR protein by 4.5-fold. In the presence of IGF-I (100 ng/mL), hCG-induced StAR protein levels were further increased. In conclusion, our present study demonstrated that IGF-I enhances Leydig cell steroidogenesis by upregulating hCG-induced StAR gene expression and protein production.
Article: Detection of hCG Responsive Expression of the Steroidogenic Acute Regulatory Protein in Mouse Leydig Cells.[show abstract] [hide abstract]
ABSTRACT: The steroidogenic acute regulatory (StAR) protein, a novel mitochondrial protein, is involved in the regulation of steroid hormone biosynthesis through its mediation of the intramitochondrial transport of the steroid substrate, cholesterol, to the cytochrome P450 cholesterol side chain cleavage (P450scc) enzyme. The expression of StAR protein is regulated by cAMP-dependent signaling in steroidogenic cells. During the course of our studies in mouse Leydig cells, we employ several methods for studying the regulation of StAR protein expression by human chorionic gonadotropin (hCG). A sensitive quantitative reverse transcription and polymerase chain reaction (RT-PCR) was utilized for determining StAR mRNA expression. Stimulation of mLTC-1 mouse Leydig tumor cells with hCG resulted in the coordinate regulation of StAR mRNA expression and progesterone accumulation in a time-response manner. The validity and accuracy of quantitative RT-PCR results in mLTC-1 cells were verified by a competitive PCR approach and were further confirmed in primary cultures of isolated mouse Leydig cells. Immunoblotting studies demonstrated an increase in the levels of the StAR protein in a concentration dependent manner following hCG stimulation in mLTC-1 cells. Northern hybridization analysis revealed three StAR transcripts, all of which were of sufficient size to encode functional StAR protein, and which were coordinately expressed in response to hCG. Collectively, the experimental approaches utilized in the present investigation allow for the demonstration and characterization of hCG mediated regulation of StAR mRNA and StAR protein expression in mouse Leydig cells.Biological Procedures Online 02/2004; 6:83-93. · 1.29 Impact Factor
[show abstract] [hide abstract]
ABSTRACT: Steroid hormone biosynthesis is acutely regulated by pituitary trophic hormones and other steroidogenic stimuli. This regulation requires the synthesis of a protein whose function is to translocate cholesterol from the outer to the inner mitochondrial membrane in steroidogenic cells, the rate-limiting step in steroid hormone formation. The steroidogenic acute regulatory (StAR) protein is an indispensable component in this process and is the best candidate to fill the role of the putative regulator. StAR is expressed in steroidogenic tissues in response to agents that stimulate steroid production, and mutations in the StAR gene result in the disease congenital lipoid adrenal hyperplasia, in which steroid hormone biosynthesis is severely compromised. The StAR null mouse has a phenotype that is essentially identical to the human disease. The positive and negative expression of StAR is sensitive to agents that increase and inhibit steroid biosynthesis respectively. The mechanism by which StAR mediates cholesterol transfer in the mitochondria has not been fully characterized. However, the tertiary structure of the START domain of a StAR homolog has been solved, and identification of a cholesterol-binding hydrophobic tunnel within this domain raises the possibility that StAR acts as a cholesterol-shuttling protein.Annual Review of Physiology 02/2001; 63:193-213. · 20.83 Impact Factor
Article: Multiple signaling pathways regulating steroidogenesis and StAR expression: more complicated than we thought