RESUMEN
Pathophysiology of amyotrophic lateral sclerosis (ALS) remains partially understood even though it is accepted worldwide that motor neuron death results from a pluri-factorial process with a variable role of genetic factors. Although not distinguishable from a clinical point of view, familial forms of ALS (fALS, 10% of cases) and sporadic forms (sALS, 90% of cases) can be described. Since the identification of superoxide dismutase 1 gene (SOD1) mutations, more than 30 genes have been linked to fALS. Among these genes, five (C9ORF72, SOD1, TARDBP, FUS, TBK1) seem predominant with mutation frequencies of 40%, 20%, 5%, <5%, <5% in fALS and 6%, 3%, and <1% for the last three in sALS, respectively. The situation that classically leads to request genetic screening is the presence of a familial history of motor neuron disorders (MND) or fronto-temporal lobar dementia (FTLD). However, this dichotomy between fALS and sALS based on familial history can lead to mistakes since illegitimacy, ignorance of MND, FTD or psychiatric disorders within the family due to a familial censorship or lack of familial relationship, or a recessive autosomal inheritance could wrongly lead to failing to recognize a familial form. The significant development of genetic research and easier access to genetic tests in fALS increase the number of situations for which gene mutations are identified. The consequence is an increase in genetic requests from relatives of ALS patients who are eager to know their own genetic status and their own individual risk to develop ALS. Pre-symptomatic testing is thus becoming a daily issue in ALS Centers. This led us to propose a framework for such pre-symptomatic genetic testing for people at risk for developing ALS.
Asunto(s)
Esclerosis Amiotrófica Lateral/diagnóstico , Esclerosis Amiotrófica Lateral/genética , Pruebas Genéticas/normas , Esclerosis Amiotrófica Lateral/epidemiología , Enfermedades Asintomáticas , Confidencialidad/normas , Análisis Mutacional de ADN/métodos , Análisis Mutacional de ADN/normas , Revelación/normas , Diagnóstico Precoz , Frecuencia de los Genes , Estudios de Asociación Genética , Asesoramiento Genético/métodos , Asesoramiento Genético/normas , Predisposición Genética a la Enfermedad , Pruebas Genéticas/métodos , Humanos , Técnicas de Diagnóstico Molecular/métodos , Técnicas de Diagnóstico Molecular/normas , Síntomas ProdrómicosRESUMEN
We report on a case of a man with familial, X-linked, partial androgen insensitivity, in whom a new point mutation in the androgen receptor (AR) ligand-binding domain (causing a valine-to-alanine substitution at codon 686) was identified. High-dose prolonged testosterone therapy resulted in marked progression in patient's appearance and great improvement in sperm count and characteristics. In combination with intracytoplasmic microinjection, treatment resulted in fertility. This is believed to be the first report of such a case. This case supports high-dose testosterone therapeutic trial in this condition. Furthermore, it underscores the possibility of achieving fertility with current endocrine and assisted reproduction modalities, making some of these X-linked AR mutations paternally transmissible.
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Síndrome de Resistencia Androgénica/tratamiento farmacológico , Síndrome de Resistencia Androgénica/genética , Mutación Puntual , Receptores Androgénicos/genética , Testosterona/uso terapéutico , Adulto , Sustitución de Aminoácidos , Síndrome de Resistencia Androgénica/patología , Femenino , Humanos , Recién Nacido , Infertilidad Masculina/genética , Infertilidad Masculina/patología , Infertilidad Masculina/terapia , Masculino , Embarazo , Análisis de Semen , Recuento de Espermatozoides , Inyecciones de Esperma Intracitoplasmáticas , Testosterona/administración & dosificaciónRESUMEN
The role of cholesterol in female reproductive physiology has been suspected for a long time, while the molecular bases were unknown. Cholesterol is the precursor of ovarian steroid biosynthesis and is also essential for fertility. In the uterus, cholesterol is essential to achieve correct contractions at term, but an excessive uterine cholesterol concentration has been associated with contractility defects. Liver X Receptor (LXR) α and LXR ß are nuclear receptors activated by oxysterols, oxidized derivatives of cholesterol. Since their discovery, the role of LXR in the control of cholesterol homeostasis has been widely described. Beyond their cholesterol-lowering role, more recent data have linked these nuclear receptors to various physiological processes. In particular, they control ovarian endocrine and exocrine functions, as well as uterine contractility. Their contribution to female reproductive cancers will also be discussed. This review will try to enlighten on the LXR as a molecular link between dietary cholesterol and reproductive diseases in women. In the future, a better comprehension of the various physiological processes regulated by the LXR will help to develop new ligands to prevent or to cure these pathologies in women.
Asunto(s)
Colesterol/metabolismo , Fertilidad/fisiología , Receptores Nucleares Huérfanos/metabolismo , Reproducción/fisiología , Femenino , Humanos , Receptores X del Hígado , MasculinoAsunto(s)
Esclerosis Amiotrófica Lateral/diagnóstico , Esclerosis Amiotrófica Lateral/genética , Proteína Huntingtina/genética , Enfermedad de Huntington/genética , Anciano , Femenino , Heterocigoto , Humanos , Masculino , Persona de Mediana Edad , Mutación , Núcleo Familiar , Expansión de Repetición de TrinucleótidoRESUMEN
PURPOSE: Patients with locally advanced cervical cancer (LACC) are usually treated with concurrent chemoradiotherapy. Extended-field chemoradiotherapy is indicated in case of para-aortic node involvement at initial assessment. 18-Fluoro-2-deoxy-D-glucose positron emission tomography/computed tomography (18-FDG PET/CT) is currently considered to be the most accurate method of detection of node or distant metastases. The goal of this study was to evaluate the accuracy of PET at detecting para-aortic lymph node metastases in LACC patients with a negative morphological imaging. METHODS: Patients from five French institutions with LACC and both negative morphologic (magnetic resonance imaging, CT scan) and functional (PET or PET/CT) findings at the para-aortic level and distantly were submitted to a systematic infrarenal para-aortic node dissection either by laparoscopy or laparotomy. On the basis of pathological results, sensitivity, specificity, and positive and negative predictive values of PET/CT were assessed for para-aortic lymph node involvement. RESULTS: A total of 125 LACC patients (stage IB2-IVA disease with two local recurrences) fulfilled the inclusion criteria. All had an ilio-infrarenal para-aortic lymphadenectomy, either by laparoscopy (n = 117) or laparotomy (n = 8). Twenty-one patients (16.8%) had pathologically proven para-aortic metastases. Among them, 14 (66.7%) had negative PET/CT. Overall morbidity of surgery was 7.2%. All but one of the complications were mild and did not delay chemoradiotherapy. Sensitivity, specificity, and positive and negative predictive value of the PET/CT were 33.3, 94.2, 53.8, and 87.5%, respectively, for the detection of microscopic lymph node metastases. CONCLUSIONS: Laparoscopic staging surgery seems warranted in LACC patients with negative PET scan who are candidates for definitive concurrent chemoradiotherapy or exenteration.
Asunto(s)
Adenocarcinoma de Células Claras/diagnóstico , Adenocarcinoma/diagnóstico , Carcinoma de Células Escamosas/diagnóstico , Fluorodesoxiglucosa F18 , Ganglios Linfáticos/patología , Recurrencia Local de Neoplasia/diagnóstico , Tomografía de Emisión de Positrones , Neoplasias del Cuello Uterino/diagnóstico , Adenocarcinoma/terapia , Adenocarcinoma de Células Claras/terapia , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Carcinoma de Células Escamosas/terapia , Niño , Femenino , Estudios de Seguimiento , Humanos , Metástasis Linfática , Imagen por Resonancia Magnética , Persona de Mediana Edad , Recurrencia Local de Neoplasia/terapia , Radiofármacos , Estudios Retrospectivos , Sensibilidad y Especificidad , Tasa de Supervivencia , Tomografía Computarizada por Rayos X , Resultado del Tratamiento , Neoplasias del Cuello Uterino/terapia , Adulto JovenRESUMEN
BACKGROUND: 5α steroid reductase deficiency (5αSRD) is an autosomal recessive enzymatic deficiency and mutations in the 5α steroid reductase type 2 gene (SRD5A2) result in male pseudohermaphrodism caused by decreased dihydrotestosterone (DHT) synthesis. AIM: To identify the specific mutations of the SRD5A2 gene in Cypriot patients with 5αSRD. SUBJECTS AND METHODS: Five unrelated patients with 46,XY karyotype were examined. Four of them were born with ambiguous genitalia and 1 patient, who was raised as girl, presented with primary amenorrhea. The hCG test was informative (elevated testosterone/DHT) of 5αSRD in 3 out of 4 subjects. Sequencing of the SRD5A2 gene was completed for all patients. Genomic DNA was also isolated from a total of 204 healthy unrelated Cypriot subjects. Screening for the IVS1-2A>G mutation was performed by using direct sequencing and restriction enzyme analysis. RESULTS: The IVS1-2A>G was identified in homozygosity in 3 patients and in a compound heterozygote state in the other 2 patients, in combination with p.P181L and p.R171S in exon 3, respectively. The carrier frequency in the Cypriot population for the IVS1-2A>G mutation was estimated to be 0.98% or 2 in 204. CONCLUSIONS: The same IVS1-2A>G mutation in the SRD5A2 gene seems to characterize all Cypriot patients with 5αSRD diagnosed so far. Furthermore this relatively rare genetic defect, which has only been reported previously in a single case in the Eastern Mediterranean region, is very likely to be the result of a founder effect.
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3-Oxo-5-alfa-Esteroide 4-Deshidrogenasa/deficiencia , Proteínas de la Membrana/genética , 3-Oxo-5-alfa-Esteroide 4-Deshidrogenasa/genética , Adolescente , Adulto , Secuencia de Bases , Preescolar , Gonadotropina Coriónica , Chipre , Trastornos del Desarrollo Sexual/genética , Femenino , Efecto Fundador , Humanos , Lactante , Recién Nacido , Masculino , MutaciónRESUMEN
BACKGROUND: Inactivating LH receptor (LHR) mutations have been described so far in men as well as in women. Phenotypes in men have been variable with in nearly all cases impairment of sex differentiation or azoospermia. We report a milder reproductive phenotype both in a male patient and his sister. METHODS AND RESULTS: We describe a family that carries a homozygous mutation G-->A at position -1 at the intron 10-exon 11 boundary of the LHR gene. The male patient presented with delayed puberty, micropenis and oligospermia. Two of his sisters were homozygous for the same mutation and were infertile. Surprisingly, one of them was found to have had regular ovarian cycles for years and showed normal LH values (6.5 and 10.6 mIU/ml for LH and FSH, respectively). In vitro analysis showed that this altered splicing resulted in an LHR from which eight amino acids are deleted from the extracellular domain (Delta Tyr(317)-Ser(324)). In vitro expression has shown that the receptor was expressed and capable of LH-induced signaling, albeit with reduced potency (P < 0.001). CONCLUSIONS: LHR mutations may represent an underestimated cause of infertility in women, in addition to being responsible for male hypogonadism with reduced spermatogenesis.
Asunto(s)
Empalme Alternativo , Hipogonadismo/genética , Infertilidad Femenina/genética , Oligospermia/genética , Receptores de HL/genética , Adulto , Secuencia de Bases , Células Cultivadas , Femenino , Humanos , Masculino , Ciclo Menstrual/genética , Persona de Mediana Edad , Datos de Secuencia Molecular , Linaje , Pene/anomalías , Pubertad Tardía/genética , TransfecciónRESUMEN
Adult-onset leukoencephalopathy with axonal spheroids and pigmented glia is an autosomal dominant leukoencephalopathy related to CSF1R gene mutations. A growing number of clinicoradiologic phenotypes have been described. In this study, we analyzed brain imaging findings in 16 patients with adult-onset leukoencephalopathy with axonal spheroids and pigmented glia to refine radiologic diagnostic clues. T2/FLAIR white matter hyperintensities were present in all patients with frontal or frontoparietal predilection, with asymmetric distribution in more than one-third. Brain atrophy and callosal involvement were almost constant, and corticospinal tract involvement was frequent. Moreover, deep white matter hyperintense dots on DWI and deep punctate calcifications on CT were often found. Conversely, deep gray matter nuclei, external capsules, and brain stem were rarely involved. Our series emphasized the great variability of MR imaging findings seen in adult-onset leukoencephalopathy with axonal spheroids and pigmented glia. A complete imaging screening including DWI, T2*, and CT is mandatory to accurately assess patients with suspected inherited adult-onset leukoencephalopathy.
Asunto(s)
Leucoencefalopatías/diagnóstico por imagen , Leucoencefalopatías/patología , Adulto , Femenino , Francia , Humanos , Leucoencefalopatías/genética , Imagen por Resonancia Magnética/métodos , Masculino , Persona de Mediana Edad , Neuroimagen/métodosRESUMEN
BACKGROUND: Activating mutations of the Gsalpha gene (GNAS), which encodes for the alpha-subunit of the stimulatory G protein, have been identified in patients with McCune-Albright syndrome (MAS). Accuracy and sensitivity in the molecular diagnosis of MAS is mandatory for optimal therapeutic strategy and adapted follow-up, especially for incomplete clinical forms of MAS. To date, the highly sensitive nested PCR method with intermediary digestion by a restriction enzyme at the mutation site is one of the most widely used techniques. This study evaluated a new diagnostic method using a peptidic nucleic acid (PNA) and compared it with the nested PCR method. MATERIAL AND METHODS: One hundred and forty-eight DNA samples from eighty-eight patients presenting clinical symptoms compatible with MAS were included. The DNA samples were mainly obtained from peripheral blood, ovarian tissue or cyst liquid, and bone lesions. The nested PCR method required 4 days. PNA clamping required 1.5 days and utilized the higher thermal stability and specificity of PNA-DNA coupling to inhibit PCR product formation. Direct sequencing was subsequently performed in all cases. RESULTS: The sensitivity of mutation detection was 54% (n = 80) for nested PCR and 46.6% (n = 69) for PNA (P > 0.05). The 11 cases where PNA failed to detect the mutation were mainly incomplete and atypical clinical forms of MAS (n = 10/11). The cost per sample was 50 Euros for PNA clamping versus 136 Euros for nested PCR. CONCLUSION: PNA clamping is a rapid, reliable, and economical method to diagnose MAS. It should be the first-line diagnostic method, although negative results, especially for incomplete clinical forms of MAS, should be confirmed by nested PCR.
Asunto(s)
Displasia Fibrosa Poliostótica/diagnóstico , Displasia Fibrosa Poliostótica/genética , Subunidades alfa de la Proteína de Unión al GTP Gs/genética , Pruebas Genéticas/métodos , Ácidos Nucleicos de Péptidos , Reacción en Cadena de la Polimerasa/métodos , Niño , Cromograninas , Análisis Mutacional de ADN/métodos , Análisis Mutacional de ADN/normas , Femenino , Pruebas Genéticas/normas , Humanos , Masculino , Reacción en Cadena de la Polimerasa/normas , Reproducibilidad de los Resultados , Mapeo Restrictivo/métodos , Mapeo Restrictivo/normas , Sensibilidad y EspecificidadRESUMEN
The paediatric endocrinologist is frequently asked whether pubertal development in a girl is normal, early or too early (precocious). This review will cover all clinical expression of premature development of puberty: central precocious puberty (neurogenic, secondary, and idiopathic) where treatment with GnRHa is considered, early puberty, partial puberty or pubertal variants and peripheral or pseudo precocious puberty related to an antonomous hypersecretion of estrogens by the ovaries. A special attention should be paid also to the role of environmental disruptors in the development of peripheral precocious puberty. GnRHa treatment should be considered only when evidence of central activation of the gonadotropic axis is proved by the LHRH-test.
Asunto(s)
Pubertad Precoz/diagnóstico , Niño , Enfermedades del Sistema Endocrino/complicaciones , Ambiente , Estrógenos/metabolismo , Femenino , Displasia Fibrosa Poliostótica/complicaciones , Hormona Liberadora de Gonadotropina/agonistas , Hormona Liberadora de Gonadotropina/uso terapéutico , Humanos , Ovario/metabolismo , Pubertad , Pubertad Precoz/etiologíaRESUMEN
Etiologic diagnosis of adulthood leukodystrophy is challenging in neurologic practice. We describe here the clinico-radiological features of a novel autosomal dominant leukodystrophy in a single family. Clinical and MRI features were recorded in a three generation family. Exome sequencing was performed in two affected relatives and one healthy member. Four total relatives (3 women and 1 man, mean age at onset: 45, range 32-59) were followed: 2 for migraine and 2 for cognitive loss. MRI features were homogeneous in the four affected relatives: extensive and symmetrical white matter hyperintensities on T2-weighted images, with a posterior predominance, involvement of the middle cerebellar peduncles, corpus callosum and the posterior limb of the internal capsules. An extensive metabolic screening was negative. In addition, sequencing of pathogenic genes involved in dominant leukodystrophies (NOTCH3, LMNB1, GFAP, CSF1R) was negative. No mutation has been identified yet with exome sequencing. This report is peculiar because of dominant inheritance, adult onset, highly homogeneous white matter hyperintensities on T2-weighted MR images, predominant in the middle cerebellar peduncles and posterior part of internal capsule and absence of mutation of the genes involved in dominant leukodystrophies.
Asunto(s)
Encéfalo/patología , Leucoencefalopatías/diagnóstico , Imagen por Resonancia Magnética , Adulto , Exoma , Salud de la Familia , Femenino , Pruebas Genéticas , Proteína Ácida Fibrilar de la Glía/genética , Humanos , Lamina Tipo B/genética , Leucoencefalopatías/líquido cefalorraquídeo , Leucoencefalopatías/genética , Masculino , Persona de Mediana Edad , Mutación , Receptor Notch3 , Receptores Notch/genéticaRESUMEN
We investigated the interferences of the normal or mutated androgen receptor with the activator protein-1 (AP-1) by assessing their effects on transcriptional activity in CV-1 cells. A luciferase reporter gene was constructed downstream from either a promoter for the mouse vas deferens protein, or a trimerized 12-O-tetradecanoyl phorbol-13-acetate-response element site whose transcriptions are activated by androgen and 12-O-tetradecanoyl phorbol-13-acetate, a potent AP-1 activator. The blockade of dephosphorylation by protein phosphatases identifies the protein phosphatases that modulate the AP-1/androgen receptor cross-talk. Using engineered or naturally occurring androgen receptor mutants that are responsible for complete or partial androgen insensitivity syndromes, we defined the subregions involved in the cross-talk of the androgen receptor with the AP-1 factors. First, it appears that the 188 first amino acids of the N-terminal domain of the androgen receptor are necessary to obtain a full transrepression. Second, a functional and intact ligand binding domain is critical for the modulation of androgen/AP-1 pathway interactions. Third, normal DNA binding capacity of the androgen receptor is not required. Two mutants at positions 568 and 581 of the DNA binding domain demonstrate that the transactivation and transrepression functions of the androgen receptor can be dissociated. Collectively, these data indicate that several segments of the androgen receptor are involved in cross-talk with the AP-1 pathway. Mutations within the DNA binding domain of the androgen receptor highly impair these interferences.
Asunto(s)
Aldehído Reductasa , Receptores Androgénicos/química , Receptores Androgénicos/genética , Factor de Transcripción AP-1/metabolismo , Transcripción Genética , Andrógenos/metabolismo , Animales , Neoplasias de la Mama Masculina/metabolismo , Células COS , ADN/metabolismo , Inhibidores Enzimáticos/farmacología , Genes Reporteros , Ingeniería Genética , Humanos , Luciferasas/genética , Masculino , Toxinas Marinas , Ratones , Mutación , Ácido Ocadaico/farmacología , Oxazoles/farmacología , Proteínas/genética , Proteínas/metabolismo , Proteínas Proto-Oncogénicas c-fos/metabolismo , Proteínas Proto-Oncogénicas c-jun/metabolismo , Estaurosporina/farmacología , Relación Estructura-Actividad , Acetato de Tetradecanoilforbol/farmacología , Transcripción Genética/efectos de los fármacos , TransfecciónRESUMEN
The naturally occurring mutations of the androgen receptor (AR), detected in patients with androgen insensitivity syndrome (AIS), are currently analyzed by in vitro assays. Unfortunately, these assays do not always permit the demonstration of a direct relationship between the in vitro activity of the receptor and the severity of the phenotype (in particular, for mutations detected in patients with partial AIS). We recently studied the trafficking of wild-type AR, fused to the green fluorescent protein (GFP) in living cells. In the present study, we applied this method for the analysis of AR mutants to find out whether it could be a complementary method of investigation of AIS. After construction of the GFP-AR mutant fusion proteins, the androgen-binding characteristics, nuclear transfer capacities, and transcriptional activities were evaluated. The nuclear transfer was quantified in the presence of various concentrations of dihydrotestosterone (DHT). We studied two mutants associated with partial AIS: G743V and R840C. The androgen-binding characteristics of both mutants were affected, in comparison with normal AR. Although the affinities were similar, the dissociation rate of GFP-AR-G743V was twice that of GFP-AR-R840C. In transcriptional assay, both mutants were active only at high concentrations of androgen. The nuclear trafficking of the mutants was evaluated by two parameters: 1) the rate of nuclear transfer; and 2) the maximal amount of receptors imported into the nucleus. At 10(-6) mol/L DHT, the GFP-AR mutants entered into the nucleus in a fashion similar to that of GFP-AR-wt. At 10(-7) mol/L DHT, the rate and maximal degree of nuclear import were both reduced, even more, for GFP-AR-G743V. The difference between mutants was more pronounced at 10(-9) mol/L DHT, because GFP-AR-G743V entered into the nucleus with even slower kinetics. Though the androgen-binding affinity and transcriptional activity assays did not reveal major differences between mutants, the dissociation rate and the trafficking capacity measurements permitted the activity of the mutants to be differentiated. We observed that the nuclear transfer capacities of these mutants are in correlation with the severity of the phenotype. The GFP-AR model provides an opportunity both to observe the dynamics of the hormone/receptor complex in living cells and to study the impact of the ligand-binding domain mutation, as opposed to certain in vitro techniques. Because the nuclear import capacity correlates well with the degree of androgen insensitivity, the GFP-AR is a useful complementary tool to understanding the phenotype/genotype relationship of AR function in patients with AIS.
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Andrógenos/fisiología , Proteínas Luminiscentes/genética , Mutación/genética , Receptores Androgénicos/genética , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/metabolismo , Andrógenos/metabolismo , Línea Celular Transformada , Resistencia a Medicamentos/genética , Proteínas Fluorescentes Verdes , Humanos , Membranas Intracelulares/metabolismo , Síndrome , Transcripción Genética/genéticaRESUMEN
A wide spectrum of androgen receptor (AR) gene mutations has been reported in complete androgen insensitivity syndromes. The molecular basis of androgen resistance was investigated in a female newborn with complete testicular feminization. Sequencing identified a point mutation in exon 4 responsible for a leucine (CTG) to arginine (CGG) replacement at codon 707. This novel mutation is located in the amino-terminal part of the ligand-binding domain of the AR. To determine the functional properties of the mutated AR and to establish the correlation with the clinical phenotype of androgen resistance, the mutation was reproduced in AR wild-type complementary DNA, and the plasmid was transfected into AR-free mammalian cells. In vitro studies showed that the mutant AR was functionally defective as an androgen-binding molecule. Electrophoretic mobility shift assay revealed that the binding of mutated AR to DNA was reduced. Finally, the mutant was unable to induce the transcriptional activation of androgen-responsive reporter gene. This amino acid defect in the primary sequence probably involves the rupture of hydropathicity in a region that is conserved among members of the steroid receptor subfamily. Our data substantiate the major contribution of leucine 707 to normal AR function and demonstrate that its substitution by an arginine caused the complete androgen insensitivity in this patient. Our findings also contribute to the elaboration of the structure-function map of the AR based on naturally occurring mutations.
Asunto(s)
Síndrome de Resistencia Androgénica/genética , Andrógenos , Resistencia a Medicamentos/genética , Receptores Androgénicos/genética , Secuencia de Aminoácidos , Secuencia de Bases , ADN/metabolismo , Exones , Humanos , Recién Nacido , Luciferasas/biosíntesis , Masculino , Metribolona/metabolismo , Datos de Secuencia Molecular , Receptores Androgénicos/metabolismoRESUMEN
We report an unusual observation of a 3.8-yr-old boy with McCune-Albright syndrome (MAS) associated with abnormal prepubertal testis enlargement and no sexual precocity. Physical examination showed café-au-lait skin lesions, enlarged testes, prepubertal sized penis, and no pubic or axillary hair. Skeletal radiography disclosed fibrous dysplasia. The serum testosterone level was 0.58 nmol/L and remained below 1.4 nmol/L during the 4-yr follow-up. By contrast, serum inhibin B and anti-Mullerian hormone concentrations were abnormally increased up to 255 pg/mL (childhood range, 35--180) and 792 pmol/L (childhood range, 309--566), respectively. The LH response to a GnRH test was in the prepubertal range, whereas the FSH response was blunted. This abnormal hormone concentration profile indicates autonomous hyperfunction of Sertoli cells, with no evidence of Leydig cell activation. Testicular histology showed tubules with marked Sertoli cell hyperplasia and very rare germinal cells, and interstitial tissue containing mesenchymal cells but no mature Leydig cells. DNA sequence analysis from bone and testis tissues detected the known activating mutation in MAS that results in replacement of Arg by His at codon 201 of the G(s)alpha protein. Other endocrine tests showed excessive GH secretion and moderate adrenal androgen hypersecretion. These findings are consistent with the occurrence of an activating mutation of the G(s)alpha gene mainly expressed in Sertoli cells and weakly expressed or absent in Leydig cells. Abnormal prepubertal testicular enlargement extends the clinical spectrum of MAS, suggesting that determination of serum inhibin B and anti-Mullerian hormone should be considered in boys with this syndrome. This observation demonstrates the usefulness of detailed molecular and biological investigations in atypical cases of MAS.
Asunto(s)
Displasia Fibrosa Poliostótica/complicaciones , Subunidades alfa de la Proteína de Unión al GTP Gs/genética , Mutación/fisiología , Células de Sertoli/fisiología , Testículo/anomalías , Secuencia de Bases/genética , Preescolar , Anomalías Congénitas/etiología , Anomalías Congénitas/patología , Humanos , Masculino , Mutación/genética , Isoformas de Proteínas/genética , Pubertad , Testículo/patologíaRESUMEN
The molecular basis of 5 alpha-reductase (5 alpha R) deficiency was investigated in four patients from three European families. In the French family, the first patient was raised as a female, and gonadectomy was performed before puberty. The second sibling, also raised as female, differed in that gonadal removal was performed after the onset of pubertal masculinization. The other two patients, both from Polish families, developed masculinization of external genitalia during puberty. All patients developed a female sexual identity. In all cases, no known consanguinity or family history of 5 alpha R deficiency was reported. The genomic DNAs of the patients were sequenced after polymerase chain reaction amplification of the five exons of the 5 alpha R type 2 gene. We found two homozygous mutations responsible for glutamine to arginine and histidine to arginine substitution in families 1 and 3, respectively. In family 2, we found a heterozygous mutation responsible for an asparagine to serine substitution at position 193. The glutamine/arginine 126 mutation in the French family was previously reported in a Creole ethnic group, and the Polish histidine/arginine 231 mutation was previously reported in a patient from Chicago. Moreover, all of the mutations created new restriction sites, which were used to determine the kindred carrier status in the three families. Because 5 alpha R deficiency is known to be a heterogenous disease in terms of clinical and biochemical expression, our data suggest that molecular biology analysis of the type 2 gene could be an essential step in diagnosing 5 alpha R deficiency.
Asunto(s)
3-Oxo-5-alfa-Esteroide 4-Deshidrogenasa/deficiencia , 3-Oxo-5-alfa-Esteroide 4-Deshidrogenasa/genética , Trastornos del Desarrollo Sexual/genética , Mutación Puntual , Adolescente , Secuencia de Aminoácidos , Arginina , Asparagina , Secuencia de Bases , Trastornos del Desarrollo Sexual/enzimología , Exones , Femenino , Francia , Glutamina , Histidina , Humanos , Intrones , Masculino , Datos de Secuencia Molecular , Sistemas de Lectura Abierta , Polonia/etnología , SerinaRESUMEN
Over the past 5 yr several inactivating mutations in the LH receptor gene have been demonstrated to cause Leydig cell hypoplasia, a rare autosomal recessive form of male pseudohermaphroditism. Here, we report the identification of two new LH receptor mutations in a compound heterozygous case of complete Leydig hypoplasia and determine the cause of the signaling deficiency at a molecular level. On the paternal allele of the patient we identified in codon 343 a T to A transversion that changes a conserved cysteine in the hinge region of the receptor to serine (C343S); on the maternal allele a T to C transition causes another conserved cysteine at codon 543 in trans-membrane segment 5 to be altered to arginine (C543R). Both of these mutant receptors are completely devoid of hormone-induced cAMP reporter gene activation. Using Western blotting of expressed LH receptor protein with a hemagglutinin tag, we further show that despite complete absence of total and cell surface hormone binding, protein levels of both mutant LH receptors are only moderately affected. The expression and study of enhanced green fluorescent protein-tagged receptors confirmed this view and further indicated that initial translocation to the endoplasmic reticulum of these mutant receptors is normal. After that, however, translocation is halted or misrouted, and as a result, neither mutant ever reaches the cell surface, and they cannot bind hormone. This lack of processing is also indicated by reduced presence of an 80-kDa protein, the only N-linked glycosylated protein in the LH receptor protein profile. Thus, complete lack of signaling by the identified mutant LH receptors is caused by insufficient processing from the endoplasmic reticulum to the cell surface and results in complete Leydig cell hypoplasia in this patient.
Asunto(s)
Trastornos del Desarrollo Sexual/genética , Trastornos del Desarrollo Sexual/patología , Heterocigoto , Células Intersticiales del Testículo/patología , Mutación/fisiología , Receptores de HL/genética , Secuencia de Aminoácidos/genética , Secuencia de Bases/genética , Niño , Trastornos del Desarrollo Sexual/fisiopatología , Exones , Femenino , Humanos , Membranas Intracelulares/metabolismo , Masculino , Datos de Secuencia Molecular , Linaje , Procesamiento Proteico-Postraduccional , Receptores de HL/fisiología , Transducción de SeñalRESUMEN
Androgen insensitivity syndrome (AIS) is an X-linked disease caused by mutations in the androgen receptor (AR) resulting in various degrees of defective masculinization in 46,XY individuals. In the present study, we describe a novel mutation in exon 7 of the AR gene in an Egyptian patient with partial AIS (PAIS). Sequencing analysis of the AR gene revealed a novel missense mutation, P817A, within the ligand-binding domain (LBD). This is the first report of a mutation within the short amino acid motif (codons 815-817) of the beta-strand lying between helices H8 and H9 of the AR LBD. The functional defects of the mutated protein were characterized by in vitro study and included significantly decreased ligand-binding affinity and impaired transactivation potential. Limited proteolysis assays performed with the wild-type and mutant AR receptors incubated with the synthetic agonist R1881 revealed that the P817A mutation resulted in a reduced stabilization of the AR active conformation. Structural analyses showed that this mutation is likely to perturb the beta-sheet interaction between residues 815-817 and 911-913. This structural alteration destabilizes the position of the C-terminal extension, which contains residues critical for androgen function.
Asunto(s)
Síndrome de Resistencia Androgénica/genética , Mutación Missense , Conformación Proteica , Receptores Androgénicos , Animales , Células COS , Chlorocebus aethiops , Humanos , Masculino , Modelos Moleculares , Técnicas de Diagnóstico Molecular , Estructura Molecular , Receptores Androgénicos/química , Receptores Androgénicos/genética , Receptores Androgénicos/metabolismoRESUMEN
We previously described an androgen receptor (AR) point mutation located in the DNA-binding domain (DBD), adjacent to another AR substitution. Both were observed in two unrelated families with male breast cancer (MBC) and partial androgen insensitivity syndrome. This work was designed to determine the potential role of these two residues by in vitro study of the consequences of these two substitutions on biological functions and their structural impact at the atomic level. Mutant ARs revealed normal androgen-binding affinities and weaker DNA binding to an isolated androgen-responsive element. In cotransfection assays the mutant ARs displayed a reduced transactivation efficiency at 0.3 x 10(-10) M. Neither binding to an estrogen-responsive element nor transactivation efficiency of an ERE reporter gene was observed. Molecular modeling revealed that Arg607 and Arg608 were partially surface-exposed and located in adjacent areas in the AR-DBD complex with DNA. This is in favor of a protein-protein interaction. It is conceivable that such an interaction could be affected by mutation of one of these two arginines.