A single base mutation in the androgen receptor gene causes androgen insensitivity in the testicular feminized rat.

The complete form of androgen insensitivity is an inherited X-linked syndrome in which genetic males fail to undergo masculinization in utero due to defective functioning of the androgen receptor (AR). The molecular basis of androgen insensitivity was investigated in the testicular feminized (Tfm) rat with this syndrome. AR mRNA size and amount, as well as nuclear AR protein revealed by immunocytochemistry, suggested normal expression of the AR gene in the Tfm rat. Sequence analysis of the AR coding region from Tfm and wild-type littermate male rats revealed a single transition mutation, guanine to adenine, within exon E, changing arginine 734 to glutamine within the steroid-binding domain of the AR. This arginine is highly conserved among the family of nuclear receptors and may be part of a phosphorylation recognition site. A recreated mutant AR (Arg734----Gln) expressed in COS cells had only 10-15% of the androgen-binding capacity of wild-type AR; the reduced androgen-binding capacity was similar to that of AR in tissue extracts of the Tfm rat. Stimulation of transcriptional activity by the recreated mutant AR was reduced relative to wild-type AR in cotransfection assays in CV1 cells using as reporter plasmid the mouse mammary tumor virus promoter linked to the chloramphenicol acetyltransferase gene. Thus, arginine 734 appears essential for normal AR function both in androgen binding and transcriptional activation. Absence of these functions results in androgen insensitivity and lack of male sexual development

Androgen insensitivity syndrome (AIS)

SummaryAndrogen insensitivity syndrome in its complete form is a disorder of hormone resistance characterised by a female phenotype in an individual with an XY karyotype and testes producing age-appropriate normal concentrations of androgens. Pathogenesis is the result of mutations in the X-linked androgen receptor gene, which encodes for the ligand-activated androgen receptor—a transcription factor and member of the nuclear receptor superfamily. This Seminar describes the clinical manifestations of androgen insensitivity syndrome from infancy to adulthood, reviews the mechanism of androgen action, and shows examples of how mutations of the androgen receptor gene cause the syndrome. Management of androgen insensitivity syndrome should be undertaken by a multidisciplinary team and include gonadectomy to avoid gonad tumours in later life, appropriate sex-hormone replacement at puberty and beyond, and an emphasis on openness in disclosure

Androgen receptor genotyping in a large Australasian cohort with androgen insensitivity syndrome; identification of four novel mutations

We genotyped the androgen receptor (AR) gene in 31 Australasian patients with androgen insensitivity syndrome (AIS). The entire coding region of AR was examined including analysis of polymorphic CAG and GGN repeats in all patients. AR defects were found in 66.7% (6/9) of patients with complete AIS (CAIS) and 13.6% (3/22) of patients with partial AIS (PAIS). A novel deletion (N858delG) leading to a premature stop codon was found in CAIS patient P1. CAIS patient P2 has a novel deletion (N2676delGAGT) resulting in a stop at codon 787. These mutations would result in inactivation of AR protein. A novel insertion of a cysteine residue in the first zinc finger of the AR DNA-binding domain (N2045_2047dupCTG) was found in CAIS patient P3. PAIS patient P4 has a novel amino acid substitution (Arg760Ser) in the AR ligand binding domain, which may impair ligand binding. Five patients were found to have previously reported AR mutations and no mutations were identified in the remaining patients

Exposure To Elevated Prenatal Testosterone Metabolites Induce Autism-Like Behavior In Rats : Evidence For The Extreme Male Brain And Implications For Human Health

Neurodevelopmental disorders such as autism spectrum disorder (ASD) originate during early brain development and are the result of complex interactions between genetic and environmental factors. Disruptions in the prenatal hormone environment have been associated with increasing the risk for autism. The extreme male brain theory of autism states that ASD results from abnormally high levels of prenatal testosterone that result in a hypermasculinized (autistic) brain. Prenatal androgen signaling programs behavior through the actions of androgen or estrogen receptors throughout diverse brain regions. The balance of this sex hormone signaling is critical for neuronal organization, and disruptions in normal prenatal hormone levels lead to aberrations in critical behaviors such as juvenile social play, which is important for normal cognitive and social development. This research demonstrates that exposure to excess testosterone metabolites during development induces autism-like behaviors in the rat, including reduced social interactions, abnormal stress response, and enhanced spatial ability, which support the extreme male brain theory.M.S

Role of Androgen Receptor in Hypothalamic Regulation of Metabolism and Reproduction

Androgens are steroid hormones that have sex-specific effects on regulation of metabolic and reproductive physiology. Androgens primarily act upon nuclear hormone receptors, including androgen receptors (AR), or estrogen receptors (ERs) after conversion to estradiol by aromatase. Adult males typically have higher levels of circulating androgens. However, imbalance of androgens outside the homeostatic range has both reproductive and metabolic deficits for both sexes. For example, hypoandrogenism in males and hyperandrogenism in females can result in infertility, obesity, and increased risk of diabetes. While many tissues express AR and are sensitive to the effects of androgens, the brain is a key androgen responsive organ. In adults, AR is expressed in many brain regions, including those that are involved in regulation of reproduction, metabolism, behavior, cognition, mood, and autonomic processes. Yet, characterization of AR expression has been lacking in the female and prepubertal brain. Here, we present a comprehensive neuroanatomical characterization of Ar mRNA expression in the mouse brain of both sexes, and compare adult and prepubertal expression. We found that expression of Ar undergoes dynamic change during a critical window of prepubertal development in male and female mice. Furthermore, we describe brain regions that may preferentially respond to androgens, rather than estrogens. For example, the ventral premammillary nucleus (PMv) shows dense expression of AR-immunoreactivity in both sexes, yet it is relatively low in ERs and aromatase. To identify neuronal populations that are targets of androgen action, we compared areas of dense AR expression, including the PMv and arcuate nucleus (ARH), with known neuronal populations in these nuclei. AR was found to be coexpressed in PMv and ARH leptin receptor (LepRb) neurons, which are crucial in regulation of energy homeostasis and exert permissive effects on fertility. We hypothesized that androgens acting via AR in LepRb cells contribute to the regulation of reproduction and metabolism at the hypothalamic level, and that loss of AR from LepRb cells would disrupt reproductive and metabolic homeostasis. We found that deletion of AR from LepRb cells (LepRbΔAR) results in sex-specific changes in the neuroendocrine reproductive axis, locomotor activity, and body composition. We observed that loss of negative feedback actions of sex steroids induces an exaggerated rise in luteinizing hormone in LepRbΔAR male mice and in follicle stimulating hormone in LepRbΔAR female mice. Furthermore, female LepRbΔAR mice show increased lean mass, while male LepRbΔAR mice display increased ambulatory activity. Subsequently, we tested if deletion of AR from LepRb neurons would protect female mice from hyperandrogenism-induced reproductive deficits. Female mice exposed to androgen excess during late prenatal development exhibit disrupted estrous cycles, infertility, and mild metabolic changes during adulthood. This phenotype replicates many features of polycystic ovary syndrome (PCOS), which is also partly characterized by androgen excess. We found that female mice with deletion of AR in LepRb neurons had improved estrous cycles with prenatal androgenization compared to their AR-intact littermates. Our findings highlight that LepRb neurons represent an important target of androgen action in the brain, and contribute to sex-specific differences in the neuroendocrine reproductive axis and some aspects of metabolic regulation. Furthermore, LepRb neurons may be involved in the pathogenesis of PCOS. These studies further elucidate the specific targets of androgens in the brain, and open the possibility of additional mechanistic study into the physiologic actions of androgens, especially in females.PHDMolecular and Integrative PhysiologyUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/169958/1/acara_1.pd

Sex differences in movement organization II : the organization of sex differences in movement during food protection, contact righting, skilled reaching and vertical exploration in the rat : the role of gonadal steroids, body morphology, and the central nervous system

xvi, 249 leaves : ill. ; 28 cm.Whether there are sex differences in the kinematic organization of non-reproductive behaviors is rarely addressed. In this thesis, evidence is presented that male and female rats organize their posture and stepping differently during a food protection task, contact righting, skilled reaching, and vertical rearing. Neonatal gonadal steroid exposure can alter sex-typical patterns of movement organization. Whether these differences are due to sex differences in body morphology or central nervous system (CNS) was also addressed using gravid females and tfm males. The results reveal that sex differences in movement are CNS based. Furthermore, the expression and choice of sex-typical patterns of movement can be altered by CNS injury. Finally, evidence is presented that sex differences in movement organization are also present in marsupials and insects. The implications of these results for our understanding of the evolution of sex differences in CNS anatomy and behavior will be discussed

Modèle in vitro pour l'étude de l'expression d'INSL3 en présence de perturbateurs endocriniens

Plusieurs études ont démontré l’existence d’une corrélation directe entre l’exposition fœtale à des contaminants environnementaux (comme le DEHP ou les estrogènes) et la perturbation de la différenciation sexuelle chez les fœtus mâles. Ces études ont notamment mis en évidence l’existence d’une fenêtre de susceptibilité; une période pendant laquelle l’exposition des cellules de Leydig aux contaminants perturbe leurs activités et entraine une diminution de leurs sécrétions des hormones testostérone (T) et Insulin-like 3 (INSL3). Ces hormones sécrétées par les cellules de Leydig sont requises pour la différenciation sexuelle du fœtus mâle, l’adoption des caractéristiques masculines primaires et secondaires (revue de Robert Courrier, 1962). Ces hormones influencent aussi la formation des spermatozoïdes et le comportement sexuel. Les études démontrent qu’une production insuffisante de ces hormones est corrélée avec une fertilité réduite (de modérée à très grave), une sous-masculinisation, la non-descente des testicules dans le scrotum (cryptorchidie) et parfois même une féminisation des organes sexuels primaires. Selon plusieurs spécialistes, ces symptômes seraient le résultat d’une perturbation de l’ontogenèse des testicules durant le développement fœtal. Syndrome de dysgénésie testiculaire est le nom qui a été donné à cette théorie. La cryptorchidie congénitale découlerait donc d’une insuffisance d’activités hormonales et révélerait par le fait même un problème de dysgénésie testiculaire. Bien que la prévalence varie à travers le monde, il semblerait, qu’en moyenne, au moins 4 % des nouveau-nés naissent avec cette condition. La localisation des testicules cryptorchides peut être corrigée chirurgicalement par orchiopexie (orchidopexie). Néanmoins, la correction chirurgicale ne semble pas être suffisante puisque les individus cryptorchides à la naissance demeurent plus à risque de développer un cancer du testicule et de souffrir de problèmes reliés à la fertilité, et ce, même après correction chirurgicale. Bien que les preuves provenant des études épidémiologiques et animales soient de plus en plus explicites, les preuves moléculaires demeurent partielles ou peu concluantes. Des preuves de nature moléculaire pourraient permettre de mieux définir le risque réel que représente le DEHP pour la santé humaine et la fertilité masculine. Le principal objectif des recherches effectuées était d’identifier et de valider un modèle in vitro permettant l’étude de composés potentiellement toxiques en minimisant le sacrifice d’animaux. À travers ces travaux, nous démontrons que la testostérone (T) augmente les niveaux d’ARNm d’Insl3 dans une lignée de cellules de Leydig et dans des cellules de Leydig primaires. Nous démontrons aussi que la testostérone induit l’activité des promoteurs d’Insl3 de différentes espèces. De plus, nous mettons en évidence que l’effet inducteur de la T sur l’activité transcriptionnelle et l’ARNm d’Insl3 nécessite le récepteur aux androgènes (AR). Nous avons localisé l’élément de réponse à la T sur le promoteur proximal d’INSL3. Cette région est toutefois dépourvue d’un élément de réponse aux androgènes consensus, ce qui suggère un mécanisme d’action indirect. Enfin, nous démontrons que le mono-(2-éthylhexyl) phtalate, le métabolite actif du perturbateur endocrinien DEHP, réprime aussi la transcription d’Insl3 dans les cellules de Leydig en antagonisant l’action de la dyade T/AR. Nous démontrons que l’estradiol (E2) réduit le niveau d’ARNm d’Insl3 dans les cellules de Leydig MA-10. Nos résultats démontrent qu’E2 réduit l’activité des promoteurs humain et murin d’Insl3/INSL3. Nous avons circonscrit la région répondant à l’E2 sur le promoteur proximal du promoteur humain d’INSL3. Cette région ne contient pas d’élément consensus de réponse aux estrogènes. Ceci indique que le mécanisme de répression est indirect. Conformément à cela, nous avons noté que la réponse à l’E2 est perdue lorsqu’une mutation est introduite au niveau des sites de liaisons des récepteurs nucléaires NUR77 et SF1. Enfin, nous démontrons que l’effet de l’exposition à l’E2 peut être renversé par un traitement conjoint avec de la testostérone, un activateur de la transcription d’Insl3. Malheureusement, les travaux n’ont pu être complétés dans le cadre de la présente thèse puisque la lignée cellulaire utilisée comme modèle, la lignée MA-10, a cessé de répondre aux divers stimuli. Néanmoins, les données présentées procurent d’importantes nouvelles perspectives sur la régulation de la transcription d’Insl3/INSL3 dans les cellules de Leydig, sur le mode d’action des phtalates et suggèrent des pistes de recherche à venir. La présente thèse contribue ainsi à la somme des savoirs qui permettra de comprendre comment le DEHP vient inhiber la sécrétion de T et d’INSL3