Congenital Lipoid Adrenal Hyperplasia Research Articles

Prenatal Diagnosis of Congenital Lipoid Adrenal Hyperplasia (CLAH) by Molecular Genetic Testing in Korean Siblings

Congenital lipoid adrenal hyperplasia (CLAH) is caused by mutations to the steroidogenic acute regulatory protein (StAR) gene associated with the inability to synthesize all adrenal and gonadal steroids. Inadequate treatment in an infant with this condition may result in sudden death from an adrenal crisis. We report a case in which CLAH developed in Korean siblings; the second child was prenatally diagnosed because the first child was affected and low maternal serum estriol was detected in a prenatal screening test. To our knowledge, this is the first prenatal diagnosis of the Q258X StAR mutation, which is the only consistent genetic cluster identified to date in Japanese and Korean populations.

Partial Defect in the Cholesterol Side-Chain Cleavage Enzyme P450scc (CYP11A1) Resembling Nonclassic Congenital Lipoid Adrenal Hyperplasia

The cholesterol side-chain cleavage enzyme (P450scc), encoded by the CYP11A1 gene, converts cholesterol to pregnenolone to initiate steroidogenesis. Genetic defects in P450scc cause a rare autosomal recessive disorder that is clinically indistinguishable from congenital lipoid adrenal hyperplasia (lipoid CAH). Nonclassic lipoid CAH is a recently recognized disorder caused by mutations in the steroidogenic acute regulatory protein (StAR) that retain partial function. We describe two siblings with hormonal findings suggesting nonclassic lipoid CAH, who had a P450scc mutation that retains partial function. A 46,XY male presented with underdeveloped genitalia and partial adrenal insufficiency; his 46,XX sister presented with adrenal insufficiency. Hormonal studies suggested nonclassic lipoid CAH. Sequencing of the StAR gene was normal, but compound heterozygous mutations were found in the CYP11A1 gene. Mutations were recreated in the F2 plasmid expressing a fusion protein of the cholesterol side-chain cleavage system. P450scc activity was measured as Vmax/Km for pregnenolone production in transfected COS-1 cells. The patients were compound heterozygous for the previously described frameshift mutation 835delA and the novel missense mutation A269V. When expressed in the P450scc moiety of F2, the A269V mutant retained 11% activity of the wild-type F2 protein. There is a broad clinical spectrum of P450scc deficiency. Partial loss-of-function CYP11A1 mutation can present with a hormonal phenotype indistinguishable from nonclassic lipoid CAH.

Ovarian cyst torsion in a patient with congenital lipoid adrenal hyperplasia

Congenital lipoid adrenal hyperplasia (CLAH) is the most severe form of congenital adrenal hyperplasia, characterized by lack of synthesis of all gonadal and adrenal steroid hormones. Ovarian cysts can develop as complications of CLAH. However, the precise mechanism of development and natural history of such cysts in patients with CLAH have not yet been determined. We recently treated a 14-year-old female patient with CLAH and ovarian cyst torsion. As a neonate, she presented with vomiting, skin pigmentation, and electrolyte imbalance. At that time, her serum adrenocorticotropic hormone concentration was elevated, while 17-hydroxypregnenolone level was normal, as were her normal female external genitalia. Mutation analysis showed that she was homozygous for p.Q258X in the StAR gene. She underwent spontaneous puberty and menstruation. At 14 years of age, she received salpingo-oophorectomy because of ovarian cyst torsion presenting as acute pelvic pain. Ovarian cysts are important clinical manifestations in 46,XX patients with CLAH, although onset time and severity can vary depending on individuals. Thus, patients with CLAH should be carefully monitored for ovarian complications to preserve as much ovarian function as possible.

Clinical, Genetic, and Functional Characterization of Four Patients Carrying Partial Loss-of-Function Mutations in the Steroidogenic Acute Regulatory Protein (StAR)

Nonclassic congenital lipoid adrenal hyperplasia (lipoid CAH) is a recently recognized disorder caused by mutations in the steroidogenic acute regulatory protein (StAR) that retain partial function. Affected individuals can present with a phenotype of late onset adrenal insufficiency with only mild or minimally disordered sexual development. The aim was to delineate the clinical spectrum of StAR mutations and correlate phenotype with StAR activity. Four patients had nonclassic/atypical lipoid CAH. Adrenal insufficiency was manifested at birth in two patients and at 11 months and 4 yr in the other two. Three were 46,XY with underdeveloped genitalia. The StAR gene was sequenced, mutations were recreated in expression vectors, and StAR activity was measured as pregnenolone production in COS-1 cells cotransfected with the cholesterol side-chain cleavage system. StAR mutants were expressed as N-62 StAR in bacteria, and purified proteins were tested for activity with isolated steroidogenic mitochondria and for cholesterol-binding capacity. DNA sequencing identified mutations on all alleles. Missense mutations were R188C, G221D, L260P, and F267S; we also tested R192C described by others. The respective activities of R188C, R192C, G221D, L260P, and F267S were 8.0, 39.4, 2.4, 3.1, and 6.1% of wild-type in transfected cells, and 12.8, 54.8, 6.3, 1.8, and 9.5% with isolated mitochondria. Cholesterol binding capacities of R188C, R192C, G221D, L260P, and F267S were 6.7, 55.3, 10.2, 4.6, and 20.9%. These data are correlated to the three-dimensional structure of StAR. There is a broad clinical spectrum of StAR mutations; StAR activities in vitro correlate well with clinical phenotypes.

Signal Transduction of Steroidogenic Hormones to the Adrenal and Gonadal Mitochondria and the Possibilities for Combating Lipoid Congenital Adrenal Hyperplasia

In steroidogenic tissue the first and rate limiting step of steroid hormone synthesis is the transport of cholesterol from the outer mitochondrial membrane to the inner membrane where it is enzymatically converted by cytochrome P450 side chain cleavage (P450scc) to pregnenolone, which is the metabolic precursor for all steroid hormones. This step is accelerated by steroidogenic hormones but how the signal from the periphery is transmitted to the mitochondria is only partially established. The steroidogenic acute regulatory protein (StAR) is known to accelerate the export of cholesterol from the outer mitochondrial membrane. StAR may bind cholesterol in its hydrophobic pocket and then transport or propel cholesterol towards the inner membrane. This action is thought to be dependent on interaction with proteins in the contact sites between the inner and outer mitochondrial membranes, such as the translocator protein and the voltage-dependent anion channel. Mutations in StAR often rob the protein of its steroidogenic properties and result in lipoid congenital adrenal hyperplasia (LCAH), a condition that is characterized by a build up of cholesterol and cholesterol esters in steroidogenic cells, deficiencies in mineralocorticoids and glucocorticoids and XY genetic males with female phenotypic external genitalia. StAR expression in healthy individuals is known to be regulated by protein kinase A and the transcription factor Nurr77. A complete understanding the workings of the signal transduction machinery in steroidogenic cells may allow future plans to combat LCAH by using gene therapy to restore appropriate activity to StAR gene products.

Congenital Lipoid Adrenal Hyperplasia: Functional Characterization of Three Novel Mutations in the STAR Gene

The steroidogenic acute regulatory protein (StAR) has been shown to be essential for steroidogenesis by mediating cholesterol transfer into mitochondria. Inactivating StAR mutations cause the typical clinical picture of congenital lipoid adrenal hyperplasia. The objective of the investigation was to study the functional and structural consequences of three novel StAR mutations (p.N148K in an Italian patient; p.P129fs and p.Q128R in a Turkish patient). Transient in vitro expression of the mutant proteins together with P450 side-chain cleavage enzyme, adrenodoxin, and adrenodoxin reductase yielded severely diminished cholesterol conversion of the p.N148K mutant, the combined p.P129fs and p.Q128R mutant, and the p.P129fs mutant by itself. The p.Q128R mutant led to a higher cholesterol conversion than the wild-type StAR protein. As derived from three-dimensional protein modeling, the residue N148 is lining the ligand cavity of StAR. A positively charged lysine residue at position 148 disturbs the hydrophobic cluster formed by the alpha4-helix and the sterol binding pocket. The frame shift mutation p.P129fs truncates the StAR protein. Residue p.Q128 is situated at the surface of the molecule and is not part of any functionally characterized region of the protein. The mutations p.N148K and p.P129fs cause adrenal insufficiency in both cases and lead to a disorder of sex development with complete sex reversal in the 46, XY case. The mutation p.Q128R, which is not relevant for the patient's phenotype, is the first reported variant showing a gain of function. We speculate that the substitution of hydrophilic glutamine with basic arginine at the surface of the molecule may accelerate cholesterol transfer.

Polycystic Ovaries and Adrenal Insufficiency in a Young Pubescent Female with Lipoid Congenital Adrenal Hyperplasia Due to Splice Mutation of the StAR Gene: A Case Report and Review of the Literature

We report a case of Lipoid Congenital Adrenal Hyperplasia (LCAH) secondary to Steroidogenic Acute Regulatory (StAR) gene mutation in an adolescent female with bilateral ovarian cysts. StAR gene defects follow an autosomal recessive mode of inheritance and typically present with severe adrenal insufficiency during infancy. Both sexes can be affected equally. XY males often present with sex reversal, while XX females may develop gonadal failure later in life due to premature loss of ovarian follicles. Recently there have been reported cases of successful fertility outcomes in women with LCAH. In our case report, we describe the clinical, biochemical and molecular analysis of a 16 year-old XX adolescent female who was suspected of having LCAH upon discovery of bilateral ovarian cysts in the context of adrenal insufficiency. Examination of the StAR gene revealed a homozygous splice site mutation. The patient is currently undergoing estradiol therapy to suppress ovarian cyst formation.

Nonclassic Lipoid Congenital Adrenal Hyperplasia Masquerading as Familial Glucocorticoid Deficiency

Familial glucocorticoid deficiency (FGD) is an autosomal recessive disorder resulting from resistance to the action of ACTH on the adrenal cortex. Affected individuals are deficient in cortisol and, if untreated, are likely to succumb to hypoglycemia and/or overwhelming infection. Mutations of the ACTH receptor (MC2R) and the melanocortin 2 receptor accessory protein (MRAP), FGD types 1 and 2 respectively, account for approximately 45% of cases. A locus on chromosome 8 has previously been linked to the disease in three families, but no underlying gene defect has to date been identified. The study design comprised single-nucleotide polymorphism genotyping and mutation detection. The study was conducted at secondary and tertiary referral centers. Eighty probands from families referred for investigation of the genetic cause of FGD participated in the study. There were no interventions. Analysis by single-nucleotide polymorphism array of the genotype of one individual with FGD previously linked to chromosome 8 revealed a large region of homozygosity encompassing the steroidogenic acute regulatory protein gene, STAR. We identified homozygous STAR mutations in this patient and his affected siblings. Screening of our total FGD patient cohort revealed homozygous STAR mutations in a further nine individuals from four other families. Mutations in STAR usually cause lipoid congenital adrenal hyperplasia, a disorder characterized by both gonadal and adrenal steroid deficiency. Our results demonstrate that certain mutations in STAR (R192C and the previously reported R188C) can present with a phenotype indistinguishable from that seen in FGD.

Lipoid Congenital Adrenal Hyperplasia: Pathologic Features of the Testis

Lipoid congenital adrenal hyperplasia (lipoid CAH) is a rare disorder of steroid biosynthesis that is caused by a molecular defect in the steroidogenic acute regulatory protein. Patients with lipoid CAH usually experience life-threatening adrenal insufficiency and develop female external genitalia in both genetic sexes because of deficient gonadal steroid synthesis. Little is known regarding testicular histology in patients with lipoid CAH, and less is known about the changes in testicular histology with growth. This article describes testicular histopathology in a patient with 46, XY lipoid CAH who underwent orchiectomy at the age of 8 years.

An update on the mechanism of action of the Steroidogenic Acute Regulatory (StAR) protein.

The Steroidogenic Acute Regulatory (StAR) protein has been demonstrated to be an indispensable component in the acute regulation of steroid hormone biosynthesis. The StAR protein, which is rapidly synthesized in response to tropic hormone stimulation, apparently functions by mediating the transfer of the substrate for all steroid hormones, cholesterol, from the outer to the inner mitochondrial membrane where it is cleaved to pregnenolone, the first steroid formed. This transfer is the regulated and rate limiting step in steroidogenesis. Mutations in the StAR gene have been shown to be the only cause of the potentially fatal disease, lipoid congenital adrenal hyperplasia, in which the affected individual can synthesize virtually no steroids. One of the most interesting and important areas of studies on this protein encompasses the mechanism whereby the StAR protein mediates the transfer of cholesterol to the inner mitochondrial membrane. While this mechanism remains unknown at this time, this review will attempt to bring this aspect of StAR function up to date.

Adrenocortical Responsiveness to Adrenocorticotropin: StAR Is Ascendant

The circadian rhythm in glucocorticoid secretion by the adrenal cortex was long considered to be a product and direct reflection of the circadian rhythm of ACTH secretion by the anterior pituitary. However, data have accumulated for several years indicating that changes in circulating ACTH cannot fully explain the adrenal rhythm, and under many conditions there is a dissociation between ACTH and glucocorticoids (1). The circadian rhythm in plasma glucocorticoid concentration is extremely robust, and in many species has a much greater amplitude than the ACTH rhythm (2). In rats after hypophysectomy, the corticosteroid rhythm persists and exhibits 3-fold differences in response to exogenous ACTH, depending on the time of day (3, 4); and in capuchin monkeys, markedly rhythmical cortisol secretion continues after ACTH suppression with dexamethasone (5). Additional findings of nycthemeral differences in adrenal responsiveness to ACTH (6, 7), the ability of a restricted feeding schedule to alter those rhythms (8), and the demonstration that innervation of the adrenal cortex via the splanchnic nerve plays a major role in regulation of circadian glucocorticoid secretion (2, 9) have all contributed to our understanding that stimulation of adrenal steroidogenesis is subject to a number of potent influences in addition to ACTH. Circadian clocks, both the central rhythm generator in the suprachiasmatic nuclei of the hypothalamus and the peripheral oscillators that have been discovered in multiple mammalian tissues, regulate temporal output via interlocking transcriptional/ translational feedback loops involving the core clock genes Per1-2, Cry1-2, Clock, and Bmal1, and their protein products (10). In recent years the molecular machinery necessary for generation of circadian rhythms has been found in the adrenal cortex of rodents and primates, and the adrenal clock of capuchin monkeys maintains a circadian rhythm in vitro (11). These findings suggest that this molecular adrenal clock may also be among the modulators of the steroidogenic response to ACTH. In this issue of Endocrinology, Torres-Farfan et al. (12) have demonstrated that in the capuchin monkey (Cebus apella) adrenal cortex, an antisense knockdown of the expression of the adrenal clock protein cryptochrome 2 (CRY2) blocks the adrenal’s steroidogenic response to stimulation by ACTH. Another recent study showed that expressionofadifferent canonical clockprotein, BMAL1, is necessary for steroidogenesis in the Y-1 immortalized mouse adrenocortical cell line (13). In addition, a transgenic mouse line in which a part of the BMAL1 coding region was expressed in an antisense orientation under tissue-specific control failed to express rhythms of corticosteroid synthesis and secretion when the animals were maintained in constant darkness. Because steroidogenic cells store very little steroid, acute increases in secretion in response to a stimulus require rapid synthesis of new steroid, and the rate-limiting step of transfer of cholesterol across the mitochondrial membranes is dependent on de novo protein synthesis (14). Acute steroidogenesis was found to be accompanied by the synthesis of a phosphoprotein (15) that occurs much more rapidly than ACTH-elicited synthesis of steroidogenic enzymes. The protein was cloned and named steroidogenic acute regulatory protein (StAR) (16). In the aforementioned studies in which adrenal steroidogenic responses to ACTH were found to be dependent on clock proteins and in several other investigations of factors observed to modulate steroidogenesis, the effects were achieved at least partly by alteration of StAR transcription. StAR is expressed only in steroidogenic tissues [adrenal cortex, testes, ovaries, placenta, and brain (17)] and mediates the transfer of mobilized cholesterol from the outer to the inner mitochondrial membrane, the site of the first enzymatic step in steroidogenesis, side chain cleavage of cholesterol to form pregnenolone. The clinical consequence of dysfunctional StARs resulting from gene mutations is congenital lipoid adrenal hyperplasia (18). The absence of functional StAR results in a roughly 85% loss of steroidogenesis that elicits compensatory ACTH secretion and subsequent cellular damage from massive accumulation in lipid droplets of esterified cholesterol that cannot be transported into mitochondria (14).

Gonadal Function, First Cases of Pregnancy, and Child Delivery in a Woman with Lipoid Congenital Adrenal Hyperplasia

Mutations in the steroidogenic acute regulatory protein (StAR) gene often cause lipoid congenital adrenal hyperplasia (LCAH). In this disorder an impairment of steroid synthesis leads to adrenal and gonadal insufficiencies with a particular female genital phenotype in both human karyotypes. Pregnancy in LCAH has not been yet reported. We describe the first cases of pregnancy in a LCAH female patient bearing the L275P mutation in the StAR gene. We studied the gonadal function, pubertal development, and apply the appropriate hormonal therapy to support pregnancies. A 46,xx patient of French Canadian descent was diagnosed with LCAH at the age of 4.5 months. Substitution therapy with glucocorticoids and mineralocorticoids led to normal growth and development. Progressive pubertal development started at the age of 11 7/12 yr. Menarche occurred at 14 2/12 yr with normal regular menstruations thereafter but without ovulation. Clomiphene stimulation induced the first pregnancy at 25 4/12 yr of age. Spontaneous abortion occurred after 6 wk gestation. The second pregnancy (with clomiphene stimulation) was induced at the age of 26 yr. Progesterone (Prog) therapy was added at the 17th day of the cycle to protect pregnancy. Vaginal delivery of dichorionic-diamniotic twin pregnancy occurred at 30 wk gestation (two normal weight male babies). Two years later, again under clomiphene stimulation, she underwent another successful singleton pregnancy and delivered a normal weight female baby at 36 wk. The pregnancies were almost uncomplicated. Despite the dysfunctional StAR, pregnancy is possible under the proper therapeutic strategy.

The mechanism of specific binding of free cholesterol by the steroidogenic acute regulatory protein: evidence for a role of the C-terminal α-helix in the gating of the binding site

Steroidogenesis depends on the delivery of free cholesterol to the inner mitochondrial membrane by StAR (steroidogenic acute regulatory protein). Mutations in the StAR gene leads to proteins with limited cholesterol-binding capacity. This gives rise to the accumulation of cytoplasmic cholesterol, a deficit in steroid hormone production and to the medical condition of lipoid congenital adrenal hyperplasia. A detailed understanding of the mechanism of the specific binding of free cholesterol by StAR would be a critical asset in understanding the molecular origin of this disease. Previous studies have led to the proposal that the C-terminal alpha-helix 4 of StAR was undergoing a folding/unfolding transition. This transition is thought to gate the cholesterol-binding site. Moreover, a conserved salt bridge (Glu169-Arg188) in the cholesterol-binding site is also proposed to be critical to the binding process. Interestingly, some of the documented clinical mutations occur at this salt bridge (E169G, E169K and R188C) and in the C-terminal alpha-helix 4 (L275P). In the present study, using rationalized mutagenesis, activity assays, CD, thermodynamic studies and molecular modelling, we characterized the alpha-helix 4 mutations L271N and L275P, as well as the salt bridge double mutant E169M/R188M. The results provide experimental validation for the gating mechanism of the cholesterol-binding site by the C-terminal alpha-helix and the importance of the salt bridge in the binding mechanism. Altogether, our results offer a molecular framework for understanding the impact of clinical mutations on the reduction of the binding affinity of StAR for free cholesterol.

PGD for X-linked and gender-dependent disorders using a robust, flexible single-tube PCR protocol

X-linked genetic diseases include a wide range of disorders such as the dystrophinopathies. Additionally in some rare genetic diseases, severity of expression is gender dependent. Prevention of such disorders usually involves prenatal diagnosis and termination of affected pregnancies, while preimplantation genetic diagnosis (PGD) represents a specialized alternative that avoids pregnancy termination. To preclude the rejection of unaffected male embryos that cannot be differentiated from those affected when using fluorescence in-situ hybridization, a flexible protocol based on multiplex fluorescence polymerase chain reaction (PCR) was standardized and validated for gender determination in single cells, which can potentially incorporate any disease-specific locus. The final panel of nine loci included four loci on the Y chromosome, two on the X chromosome plus up to three microsatellite markers to either support the gender diagnosis or to further monitor extraneous contamination. The protocol, standardized on single lymphocytes, established a PCR efficiency of >93% for all loci with maximum allele dropout rates of 4%. Microsatellite analysis excluded external contamination and confirmed biallelic inheritance. Proof of principle for the simplicity and flexibility of the assay was demonstrated through its application to clinical PGD cycles for lipoid congenital adrenal hyperplasia, which presents a more severe clinical course in males, and Duchenne muscular dystrophy.

A Novel Homozygous Mutation inCYP11A1Gene Is Associated with Late-Onset Adrenal Insufficiency and Hypospadias in a 46,XY Patient

The first and the rate-limiting step in the biosynthesis of hormones in all steroidogenic tissues, conversion of cholesterol to pregnenolone, is catalyzed by the cholesterol side-change cleavage cytochrome P450 (P450scc) encoded by a single gene, CYP11A1. To date, mutations in CYP11A1 gene have been reported in six patients, all of whom presented with adrenal insufficiency within the first 4 yr of life and severely underandrogenized external genitalia (Prader stages 1-2). Our aim was to characterize in vitro and in vivo effects of a novel homozygous CYP11A1 gene mutation identified in a patient with an unusual presentation of P450scc deficiency. A 46,XY patient presented with mid-shaft hypospadias and cryptorchidism at birth and signs of adrenal failure at 9 yr. Mutational analysis of CYP11A1 gene was performed by PCR, followed by direct sequencing. P450scc activity was determined by measuring concentration of pregnenolone synthesized from cholesterol in the medium after a transient transfection of HEK293 cells with P450scc, adrenodoxin, adrenodoxin reductase, and steroidogenic acute regulatory protein expression plasmids. The sequencing of CYP11A1 gene in the proband revealed a novel homozygous L222P mutation, whereas both parents were heterozygous carriers for this mutation. In vitro P450scc activity of L222P mutant was approximately 7% compared with the wild type. This case represents the mildest phenotype of P450scc deficiency to be described. The phenotypic presentation was consistent with the partial reduction of P450scc activity of L222P mutant.

Toward the NMR structure of StAR

The steroidogenic acute regulatory (StAR) protein plays a crucial role in steroidogenesis, as it accelerates the transport of cholesterol to the inner mitochondrial membrane where the cytochrome P450scc enzyme is located. Mutations in the StAR gene can lead to lipoid congenital adrenal hyperplasia (LCAH), a disease that is fatal if not treated with hormone replacement therapy. Solving the structure of StAR is an important aspect of understanding LCAH. Point mutations or truncations in the StAR gene produce a partial to non-functional protein that hinders the StAR-induced delivery of cholesterol to the mitochondria during an acute hormonal stimulation of steroidogenic cells. So far, homology modeling, structure-based thermodynamics and biophysical studies have allowed us to propose the existence of an open state of StAR where the C-terminal alpha-helix 4 undergoes partial unfolding. This may act as a gating mechanism to the cholesterol binding site. Once bound, cholesterol leads to the stabilization and the refolding of alpha-helix 4, and eventually to the interaction with an import complex at the surface of the mitochondria. Though the current homology models have proven useful in understanding StAR function, only the full determination of the 3D structure of the apo- and holo-states will further validate this two-state model. In this context, we have used solution-state nuclear magnetic resonance (NMR) and obtained high-resolution (1)H-(15)N-HSQC spectra of StAR in its apo- and holo-states at physiological pH. Both spectra displayed well-dispersed resonances. However, key differences are observed on the spectra which indicate that both states have stable but slightly different tertiary structures. In conjunction with the binding/activity assays and biophysical methods, this original NMR data constitutes another structural step into the validation of the two-state model and the three-dimensional structure of StAR.

Conception and pregnancy outcome in a patient with 11-bp deletion of the steroidogenic acute regulatory protein gene

To report the pregnancy outcome of a patient with congenital lipoid adrenal hyperplasia (CLAH) due to an 11-bp deletion of the steroidogenic acute regulatory protein (StAR) gene. Case report. University-based pediatric endocrinology unit and private IVF clinic. A 24-year-old woman homozygous for a StAR gene deletion, married to a man heterozygous for the same molecular defect. Ovarian stimulation, oocyte retrieval followed by IVF, blastomere biopsy, preimplantation genetic diagnosis, and additional estrogen support until placental function initiation. Normal pregnancy outcome and delivery of a healthy newborn. A female patient with CLAH gave birth to a normal newborn after IVF and preimplantation genetic diagnosis. Pregnancy is feasible in patients with StAR gene mutations, provided that extra estrogens are offered until placental function ensues.

The Orphan Nuclear Receptor NUR77 Regulates Hormone-Induced StAR Transcription in Leydig Cells through Cooperation with Ca2+/Calmodulin-Dependent Protein Kinase I

Cholesterol transport in the mitochondrial membrane, an essential step of steroid biosynthesis, is mediated by a protein complex containing the steroidogenic acute regulatory (StAR) protein. The importance of this transporter is underscored by mutations in the human StAR gene that cause lipoid congenital adrenal hyperplasia, male pseudohermaphroditism, and adrenal insufficiency. StAR transcription in steroidogenic cells is hormonally regulated and involves several transcription factors. The nuclear receptor NUR77 is present in steroidogenic cells, and its expression is induced by hormones known to activate StAR expression. We have now established that StAR transcription in cAMP-stimulated Leydig cells requires de novo protein synthesis and involves NUR77. We found that cAMP-induced NUR77 expression precedes that of StAR both at the mRNA and protein levels in Leydig cells. In these cells, small interfering RNA-mediated NUR77 knockdown reduces cAMP-induced StAR expression. Chromatin immunoprecipitation assays revealed a cAMP-dependent increase in NUR77 recruitment to the proximal StAR promoter, whereas transient transfections in MA-10 Leydig cells confirmed that NUR77 can activate the StAR promoter and that this requires an element located at -95 bp. cAMP-induced StAR and NUR77 expression in Leydig cells was found to require a Ca2+/calmodulin-dependent protein kinase (CaMK)-dependent signaling pathway. Consistent with this, we show that within the testis, CaMKI is specifically expressed in Leydig cells. Finally, we report that CaMKI transcriptionally cooperates with NUR77, but not steroidogenic factor 1, to further enhance StAR promoter activity in Leydig cells. All together, our results implicate NUR77 as a mediator of cAMP action on StAR transcription in steroidogenic Leydig cells and identify a role for CaMKI in this process.

Von Hermaphroditos der griechischen Mythologie bis zu DSD der Moderne

We report a case of 34 year old woman how has been hospitalized at the age of 6 month with persistent vomitus. The vomitus was found to be caused by adrenal insufficiency with lack of all hormones of steroidobiosynthesis. The phenotypical femal child was diagnosed to have congenital lipoid adrenal hyperplasia with 46,XY DSD. 24 years later a homozygote mutation in the StAR-gene (L260P), which was first described in Switzerland, has been identified.

Severe Combined Adrenal and Gonadal Deficiency Caused by Novel Mutations in the Cholesterol Side Chain Cleavage Enzyme, P450scc

Mitochondrial cytochrome P450scc converts cholesterol to pregnenolone in all steroidogenic tissues. Although progesterone production from the fetally-derived placenta is necessary to maintain pregnancy to term, four patients with mutations in the gene encoding P450scc (CYP11A1), have been described, one in a 46,XX female and three in underandrogenized 46,XY individuals, all with primary adrenal failure. Our aim was to determine whether P450scc mutations might be found in other children and to explore genotype/phenotype correlations. We performed mutational analysis of CYP11A1 in individuals with 46,XY disorders of sex development and primary adrenal failure, followed by functional studies of P450scc activity and of P450scc RNA splicing. Among nine 46,XY infants with adrenal failure and disordered sexual differentiation, two infants had compound heterozygous mutations in CYP11A1. One patient harbored the novel P450scc missense mutations L141W and V415E, which retained 38 and 0% activity, respectively. The other carried a CYP11A1 frameshift mutation c835delA (0% activity) and a splice site mutation [IVS3+(2-3)insT] that prevented correct splicing of P450scc mRNA. P450scc deficiency is a recently recognized disorder that may be more frequent than originally thought. The phenotypic spectrum ranges from severe loss-of-function mutations associated with prematurity, complete underandrogenization, and severe, early-onset adrenal failure, to partial deficiencies found in children born at term with clitoromegaly and later-onset adrenal failure. In contradistinction to congenital lipoid adrenal hyperplasia caused by steroidogenic acute regulatory protein mutations, adrenal hyperplasia has not been reported in any of the six patients with P450scc deficiency.