scholarly journals Steroidogenic acute regulatory protein mRNA expression in adrenal tumours

2000 ◽  
pp. 294-299 ◽  
Author(s):  
S Zenkert ◽  
B Schubert ◽  
M Fassnacht ◽  
F Beuschlein ◽  
B Allolio ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Cell Line ◽  
Tumour Cell ◽  
Regulatory Protein ◽  
Steroidogenic Acute Regulatory Protein ◽  
Tumour Cell Line ◽  
Inner Mitochondrial Membrane ◽  
Endocrine Activity ◽  
Protein Factor ◽  
Steroidogenic Acute Regulatory

The rate limiting step in steroidogenesis is cholesterol transport through the outer to the inner mitochondrial membrane and the cytochrome P450 side chain cleavage (P450scc) complex. The protein factor responsible for this transport, and as such necessary for regulating the acute production of steroids, has been identified and named the steroidogenic acute regulatory protein (StAR). We investigated the expression of StAR in functional and non-functional adrenal neoplasms and compared the expression with that of P450scc. Poly A RNA was extracted from normal adrenal glands (NAG, n=5), aldosterone producing adenomas (APA, n=4), cortisol producing adenomas (CPA, n=5), adrenocortical carcinomas (ACC, n=6) and non-functional adenomas (NFA, n=3), electrophoresed through a 1% agarose gel, blotted and hybridised with a PCR-generated cDNA labelled with [(32)P]CTP. The blots were stripped and re-hybridised with a P450scc cDNA and a mouse beta-actin probe. Compared with P450scc, StAR mRNA expression showed little variability in the magnitude of expression and did not correlate with the endocrine profiles (NAG: StAR 100+/-16%, P450scc 100+/-14%; APA: StAR 80+/-3%, P450scc 94+/-13%; CPA: StAR 71+/-10%, P450scc 109+/-15%; NFA: StAR 64+/-9.5%, P450scc 18+/-5%; means+/-s.e.m.). ACC expressed low levels of both genes probably as a result of dedifferentiation (StAR 29+/-9%, P450scc 46+/-18%). Incubation of the NCI-h295 tumour cell line with 10nmol ACTH and 10micromol forskolin induced an increase in the abundance of StAR and P450scc mRNA, demonstrating gene regulation by the cAMP protein kinase A pathway. Furthermore, we incubated the NCI-h295 tumour cell line with the adrenostatic compounds, aminoglutethimide and metyrapone. We could not detect an effect on the expression of StAR mRNA, whereas the expression of P450scc mRNA was significantly reduced. We conclude that, in contrast to P450scc, StAR seems to be evenly expressed in adrenocortical adenomas. Therefore, the endocrine activity of a given tumour cannot be explained by the abundance of StAR expression. In ACC, both StAR and P450scc expression is low, explaining the relatively inefficient steroid production of these tumours.

scholarly journals Oxygen Concentration Is an Important Factor for Modulating Progesterone Synthesis in Bovine Corpus Luteum

Endocrinology ◽  
2006 ◽  
Vol 147 (9) ◽  
pp. 4273-4280 ◽  
Author(s):  
Ryo Nishimura ◽  
Ryosuke Sakumoto ◽  
Yuko Tatsukawa ◽  
Tomas J. Acosta ◽  
Kiyoshi Okuda
Keyword(s):  
Mrna Expression ◽  
Corpus Luteum ◽  
Oxygen Deficiency ◽  
Regulatory Protein ◽  
Steroidogenic Acute Regulatory Protein ◽  
Low Oxygen ◽  
Luteal Cells ◽  
O2 Concentration ◽  
Oxygen Condition ◽  
Steroidogenic Acute Regulatory

Oxygen deficiency caused by a decrease in the blood supply is known to induce various responses of cells. Because luteal blood flow has been shown to decrease during luteolysis, a low-oxygen condition seems to be an integral part of the environment during luteolysis. To determine whether a low-oxygen condition is associated with functional luteolysis, we examined the influence of reduced oxygen tension on the luteal progesterone (P4) generating system in cultured bovine midluteal cells. Luteal cells obtained from midcycle corpus luteum (d 8–12) were incubated under different O2 concentrations (20, 10, 5, 3% O2) with or without LH for 24 h. P4 production decreased with decreasing O2 concentration but was significantly stimulated by LH regardless of O2 concentration. After 8 h of culture, both basal and LH-stimulated P4 production was significantly lower under 3% O2 than under 20% O2. Low-oxygen condition also inhibited pregnenolone production. Cytochrome P450 side-chain cleavage enzyme (P450scc) mRNA expression, measured by quantitative PCR, decreased under low-oxygen condition in both non-LH-treated and LH-treated cells. Low-oxygen condition did not affect the expressions of steroidogenic acute regulatory protein mRNA or protein, whereas steroidogenic acute regulatory protein mRNA expression was stimulated by LH during 4 h of culture. Low-oxygen condition also did not affect 3β-hydroxysteroid dehydrogenase/Δ5-Δ4 isomerase mRNA expression or the activity of the enzyme in the cells, regardless of the incubation period. The overall results indicate that a low-oxygen condition decreases P4 synthesis by attenuating P450scc production and P450scc activity in bovine luteal cells and suggest that oxygen deficiency is an essential condition for the progression of luteolysis in cattle.

Changes of steroidogenic acute regulatory protein mRNA expression in postnatal rat development

2002 ◽  
Vol 139 (2) ◽  
pp. 247-254 ◽  
Author(s):  
Hyun Joon Kim ◽  
Chang Hwan Park ◽  
Gu Seob Roh ◽  
Sang Soo Kang ◽  
Gyeong Jae Cho ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Regulatory Protein ◽  
Steroidogenic Acute Regulatory Protein ◽  
Rat Development ◽  
Steroidogenic Acute Regulatory

scholarly journals Steroidogenic acute regulatory protein and luteinizing hormone are required for normal ovarian steroidogenesis and oocyte maturation in zebrafish†

2019 ◽  
Vol 101 (4) ◽  
pp. 760-770 ◽  
Author(s):  
Guohui Shang ◽  
Xuyan Peng ◽  
Cheng Ji ◽  
Gang Zhai ◽  
Yonglin Ruan ◽  
...  
Keyword(s):  
Luteinizing Hormone ◽  
Oocyte Maturation ◽  
Mitochondrial Membrane ◽  
Regulatory Protein ◽  
Steroidogenic Acute Regulatory Protein ◽  
Transcription Activator ◽  
Inner Mitochondrial Membrane ◽  
Steroidogenic Acute Regulatory ◽  
Functional Deficiency

Abstract In recent studies, luteinizing hormone (LH) was reported to play important roles in oocyte maturation. However, the mechanism by which LH signaling, especially regarding the steroidogenesis process, affects oocyte maturation has not been clarified. In this study, zebrafish models with a functional deficiency in luteinizing hormone beta (Lhb) or steroidogenic acute regulatory protein (Star), an enzyme that promotes the transport of cholesterol into the inner mitochondrial membrane for maturation-induced hormone (MIH) production, were generated using transcription activator-like effector nucleases (TALENs). Similar phenotypes of the maturation-arrested oocytes in both female mutants have been observed. The levels of MIH in the oocytes of the female mutants were clearly decreased in both the lhb and star knockout zebrafish. The expression of star was dramatically down-regulated in the lhb mutant follicles and was clearly promoted by forskolin and hCG in vitro. Furthermore, treatment with the MIH precursors, pregnenolone or progesterone, as well as with MIH itself rescued the maturation-arrested oocyte phenotypes in both lhb and star mutants. The plasma levels of other steroids, including testosterone, estradiol, and cortisol, were not affected in the lhb mutants, while the levels of gonad hormones testosterone and estradiol were significantly increased in the star mutants. The cortisol levels were decreased in the star mutants. Collectively, our results confirm that LH plays important roles in the initiation of MIH synthesis from cholesterol and maintains oocyte maturation in zebrafish, as well as provide evidence that Star might act downstream of LH signaling in steroidogenesis.

scholarly journals Interleukin-1 inhibits Leydig cell steroidogenesis without affecting steroidogenic acute regulatory protein messenger ribonucleic acid or protein levels

1998 ◽  
Vol 156 (3) ◽  
pp. 461-467 ◽  
Author(s):  
T Lin ◽  
D Wang ◽  
DM Stocco
Keyword(s):  
Mrna Expression ◽  
Leydig Cell ◽  
Leydig Cells ◽  
Interleukin 1 ◽  
Mrna Level ◽  
Primary Cultures ◽  
Regulatory Protein ◽  
Steroidogenic Acute Regulatory Protein ◽  
Protein Levels ◽  
Steroidogenic Acute Regulatory

The rate-limiting step of steroidogenesis is the transport of the substrate cholesterol from the outer to the inner mitochondrial membrane which involves a cycloheximide-sensitive newly synthesized protein. A protein believed to carry out this function was recently cloned from MA-10 mouse Leydig tumor cells and named the steroidogenic acute regulatory protein (StAR). In the present study, we evaluated the expression and regulation of StAR in primary cultures of rat Leydig cells. StAR mRNA was expressed in Leydig cells as two major transcripts of 3.8 and 1.7 kb and one minor transcript of 1.2 kb. Induction of StAR mRNA transcripts could be detected as early as 30 min after the addition of human choriogonadotropin (hCG) with peak levels attained between 2 and 4 h. hCG in concentrations of 0.1-10 ng/ml caused a dose-dependent increase in StAR mRNA expression. hCG administered at a dose of 10 ng/ml increased the 3.8 kb StAR mRNA level about 14-fold and the 1.7 kb StAR mRNA level about 13.6-fold. hCG-stimulated StAR mRNA was associated with increased StAR protein levels as determined by immunoblot analysis (a 4.5-fold increase). Murine interleukin-1 alpha (mIL-1 alpha) at a concentration of 100 ng/ml inhibited hCG-induced cytochrome P450 side-chain cleavage (P450 scc) mRNA expression and testosterone formation almost completely. Interestingly, mIL-1 alpha had no effect on hCG-induced StAR mRNA or protein levels. Furthermore, mIL-1 alpha (10 ng/ml) decreased conversion of (22R)-hydroxycholesterol to testosterone while the conversion of pregnenolone, 17-hydroxypregnenolone, dehydroepiandrosterone and androstenedione to testosterone were not affected. These results indicate that the major inhibitory effect of IL-1 on Leydig cell function occurs at the level of P450 scc.

Molten-globule structure and membrane binding of the N-terminal protease-resistant domain (63–193) of the steroidogenic acute regulatory protein (StAR)

2001 ◽  
Vol 356 (1) ◽  
pp. 151-158
Author(s):  
Maengseok SONG ◽  
Haiyan SHAO ◽  
Anwer MUJEEB ◽  
Thomas L. JAMES ◽  
Walter L. MILLER
Keyword(s):  
Secondary Structure ◽  
Tertiary Structure ◽  
Molten Globule ◽  
Regulatory Protein ◽  
Limited Proteolysis ◽  
Proteinase K ◽  
Steroidogenic Acute Regulatory Protein ◽  
Inner Mitochondrial Membrane ◽  
Terminal Domain ◽  
Steroidogenic Acute Regulatory

The first step in steroidogenesis is the movement of cholesterol from the outer to inner mitochondrial membrane; this movement is facilitated by the steroidogenic acute regulatory protein (StAR). StAR has molten-globule properties at low pH and a protease-resistant N-terminal domain at pH4 and pH8 comprising residues 63–193. To explore the mechanism of action of StAR we investigated the structural properties of the bacterially expressed N-terminal domain (63–193 StAR) using CD, limited proteolysis and NMR. Far- and near-UV CD showed that the amount of secondary structure was greater at acidic than at neutral pH, but there was little tertiary structure at any pH. Unlike 63–193 StAR liberated from N-62 StAR by proteolysis, biosynthetic 63–193 StAR was no longer resistant to trypsin or proteinase K at pH7, or to pepsin at pH4. Addition of trifluoroethanol and SDS increased secondary structure at pH7, and dodecylphosphocholine and CHAPS increased secondary structure at pH2, pH4 and pH7. However, none of these conditions induced tertiary structure, as monitored by near-UV CD or NMR. Liposomes of phosphatidylcholine, phosphatidylserine and their mixture increased secondary structure of 63–193 StAR at pH7, as monitored by far-UV CD, and stable protein–liposome complexes were identified by gel-permeation chromatography. These results provide further evidence that the N-terminal domain of StAR is a molten globule, and provide evidence that this conformation facilitates the interaction of the N-terminal domain of StAR with membranes. We suggest that this interaction is the key to understanding the mechanism of StAR's action.

scholarly journals The Contribution of Serine 194 Phosphorylation to Steroidogenic Acute Regulatory Protein Function

2014 ◽  
Vol 28 (7) ◽  
pp. 1088-1096 ◽  
Author(s):  
Goro Sasaki ◽  
Mohamad Zubair ◽  
Tomohiro Ishii ◽  
Toshikatsu Mitsui ◽  
Tomonobu Hasegawa ◽  
...  
Keyword(s):  
Adrenal Cortex ◽  
Protein Function ◽  
Regulatory Protein ◽  
Initial Step ◽  
Artificial Chromosome ◽  
Steroidogenic Acute Regulatory Protein ◽  
Inner Mitochondrial Membrane ◽  
Steroidogenic Acute Regulatory

The steroidogenic acute regulatory protein (StAR) facilitates the delivery of cholesterol to the inner mitochondrial membrane, where the cholesterol side-chain cleavage enzyme catalyzes the initial step of steroid hormone biosynthesis. StAR was initially identified in adrenocortical cells as a phosphoprotein, the expression and phosphorylation of which were stimulated by corticotropin. A number of in vitro studies have implicated cAMP-dependent phosphorylation at serine 194 (S194, S195 in human StAR) as an important residue for StAR activity. To explore the importance of S194 phosphorylation in StAR function in vivo, we developed a transgenic model using a bacterial artificial chromosome expressing either wild-type (WT) StAR or StAR mutation S194A to rescue StAR knockout (KO) mice. Despite StAR protein expression comparable to or higher than amounts seen with control animals or rescue with WT StAR, S194A StAR did not rescue the neonatal lethality and only partially rescued the sex reversal in male mice observed uniformly in StAR KO mice. Like the StAR KO mice, the adrenal cortex and testicular Leydig cells contained abundant lipid deposits when stained with oil red O. Adrenal StAR from S194A rescue animals lacks an acidic species, which appears upon corticotropin stimulation in animals rescued with WT StAR, consistent with defective StAR phosphorylation. These findings demonstrate that S194 is an essential residue for normal StAR function in the adrenal cortex and testes of mice.

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