[show abstract][hide abstract] ABSTRACT: Breast cancer development and progression is regulated by growth factors and steroid hormones. Although the majority of human breast cancers expresses androgen receptor (AR), the role of androgens in breast tumorigenesis remains largely unexplored. Here we demonstrate that an AR ligand, 5-alpha-dihydrotestosterone (DHT), inhibits MCF-7 breast cancer cell growth induced by insulin like growth factor 1 (IGF-I). Our results show that DHT induces association of AR with IRS-1, the major IGF-1 receptor signaling molecule. The AR/IRS-1 complex translocates to the nucleus and is recruited to gene promoters containing androgen responsive elements causing an increase of AR transcriptional activity. Moreover, IRS-1 knockdown suggests that IRS-1/AR interaction decreases the ubiquitin/proteasome dependent degradation of AR, increasing its stability. Taken together, these data indicate that nuclear IRS-1 is a novel AR regulator required to sustain AR activity and demonstrate, for the first time in breast cancer cells, the existence of a functional interplay between the IGF system and AR. This interplay may represent the molecular basis of mechanisms through which androgens exert their inhibitory role on the proliferation of breast cancer cells.
Breast Cancer Research and Treatment 05/2009; 115(2):297-306. · 4.47 Impact Factor
[show abstract][hide abstract] ABSTRACT: Overexpression of androgen receptor (AR) decreases estrogen receptor alpha (ERalpha) transactivation, which plays a basic role in hormone-dependent breast cancer. This transcriptional interference can be due to shared coactivators. Here we demonstrated that in MCF-7 cells ARA70, an AR-specific coactivator, interacted with endogenous ERalpha, increasing its transcriptional activity, and it was recruited to the pS2 gene promoter. Moreover, a dominant negative ARA70 down-regulated ERalpha transcriptional activity as well as pS2 mRNA. ARA70 overexpression reversed the AR down-regulatory effect on ERalpha signaling. However, in the presence of a progressive increase of transfected AR, ARA70 switched into enhancing the inhibitory effect of AR on ERalpha signaling. These opposite effects of ARA70 were further evidenced by coimmunoprecipitation assay in MCF-7wt, MCF-7-overexpressing AR, and HeLa cells, exogenously expressing an excess of ERalpha with respect to AR or an excess of AR with respect to ERalpha. Thus, ARA70 is a coactivator for ERalpha and may represent a functional link between ERalpha/AR modulating their cross-talk in models of estrogen signaling in MCF-7 and HeLa cells.
Journal of Biological Chemistry 06/2005; 280(21):20421-30. · 4.65 Impact Factor
[show abstract][hide abstract] ABSTRACT: Huntington's and Kennedy's disease are autosomal dominant neurodegenerative diseases caused by pathogenic expansion of polyglutamine tracts. Expansion of glutamine repeats must in some way confer a gain of pathological function that disrupts an essential cellular process and leads to loss of affected neurons. Association of huntingtin with vesicular structures raised the possibility that axonal transport might be altered. Here we show that polypeptides containing expanded polyglutamine tracts, but not normal N-terminal huntingtin or androgen receptor, directly inhibit both fast axonal transport in isolated axoplasm and elongation of neuritic processes in intact cells. Effects were greater with truncated polypeptides and occurred without detectable morphological aggregates.
[show abstract][hide abstract] ABSTRACT: Estrogens and thyroid hormones play a significant role in regulating functions and development of the testis. The synthesis of estrogens from androgens is catalyzed by the enzyme complex termed aromatase, which in the testis displays an age-related cellular compartmentalization, primarily in Sertoli cells in immature animals, whereas in adults it is expressed in Leydig and germ cells. T3 induces a precocious terminal differentiation of prepubertal Sertoli cells together with a dramatic decrease of their aromatase activity. In the present work, we have examined the mechanism by which T3 exerts this inhibitory action on aromatase expression. As an experimental model, we used the mouse Sertoli cell line TM4, which conserves a large spectrum of functional features present in immature Sertoli cells. For instance, after revealing the presence of aromatase by immunocytochemistry and measuring its enzymatic activity, we confirmed in this cell line the functional events previously characterized in primary cultures of immature rat Sertoli cells: 1) a strong stimulation of aromatase activity by dibutyryl-cAMP [(Bu)2cAMP] (simulating FSH action); and 2) the inhibition of aromatase activity by incubation with T3 under basal condition and after (Bu)2cAMP stimulation. After identifying promoter II as the regulatory region located immediately upstream of the transcriptional initiation site in the TM4 cell line by rapid amplification of cDNA ends analysis, we conducted experiments to examine the molecular mechanism by which thyroid hormones modulate aromatase gene expression in this cell line. TM4 cells were transfected with plasmids containing different segments of the rat promoter II sequence ligated to a luciferase reporter gene. Analysis of the activities of these promoter fusions demonstrated that T3 inhibits basal and (Bu)2cAMP-stimulated activity of the aromatase promoter. This effect was not revealed in T3-treated cells transfected with construct in which the steroidogenic factor-1 (SF-1) response element was mutated. These results indicate that the inhibitory effect of T3 requires the integrity of the SF-1 response element and are further supported in the EMSA. The EMSA experiments demonstrated that thyroid hormone/thyroid receptor alpha1 complex (TH/TRalpha1) is able to compete with SF-1 in binding to oligonucleotides containing an SF-1 motif, an element essential for the activity of the PII aromatase promoter. The findings suggest that the binding of the thyroid hormone/thyroid receptor alpha1 complex to the SF-1 motif is the molecular mechanism by which T3 exerts an inhibitory effect on aromatase gene expression in the TM4 cell line.
[show abstract][hide abstract] ABSTRACT: The androgen receptor (AR) is a high affinity receptor protein encoded on the human X-chromosome that mediates the actions of androgens during development and in the adult. Defects in this receptor protein result in a wide range of abnormalities of male sexual development. Studies in a number of different laboratories have identified mutations of the AR gene in subjects with androgen resistance syndromes. Defects that interrupt the AR open-reading frame have been traced to a number of distinct types of genetic alterations, have been identified in widely separated segments of the AR gene, and are invariably associated with the phenotype of complete androgen insensitivity. By contrast, mutations that cause single amino acid substitutions within the AR are localized to the DNA- or ligand-binding domains of the receptor protein and have been associated with the full range of androgen resistant phenotypes. The diversity of mutations that have been identified has prompted a consideration of the relationship between AR mutation and phenotype. Analyses of AR abundance and function suggest that the phenotypic abnormalities that result from mutation of the AR reflect the extent to which AR activity is impaired in target tissues. Such decreases in AR function may be the result of the diminished receptor function, decreases in receptor concentration, or a combination of these two effects.
Molecular and Cellular Endocrinology 01/2003; 198(1-2):61-7. · 4.04 Impact Factor
[show abstract][hide abstract] ABSTRACT: Defects of the AR cause a wide range of abnormalities of male development, ranging from individuals with mild defects of virilization to those with complete female phenotypes. In parallel with this phenotypic spectrum, a large number of different mutations have been identified that alter the synthesis or functional activity of the receptor protein. This report aims to categorize the alterations of immunoreactive AR (IRAR) expression and the underlying genetic changes in a single category of patient: those in whom ligand binding is undetectable in genital skin fibroblasts. Our study found a wide range in the levels of IRAR that are detectable in fibroblast strains established from 27 such individuals. A large proportion (19 of 27) express significant amounts of AR protein, as detected using a sensitive Western blot technique. In a smaller number (8/27), AR expression was undetectable. Intact IRAR was identified in16 of the 19 fibroblast strains in which AR expression could be detected. The AR gene was analyzed in 14 strains from this group. In 13 instances, single amino acid substitutions were identified within the ligand-binding domain of the receptor protein. In three of the remaining patients (3 of 19), truncation of the receptor protein was suggested by the rapid migration of the IRAR in SDS-polyacrylamide gels. In those three patients, production of the shortened immunoreactive receptor was traced to mutations that interrupted the AR open reading frame. By contrast, only one of the eight patient samples with no detectable IRAR carried a mutation that resulted in a single amino acid substitution. An interruption of the AR open reading frame was identified in six of the eight strains in which immunoreactive receptor was absent. In the remaining strain, no mutation was present within or surrounding the eight coding exons. This study serves to define the effects that mutations of the AR have on the levels of expressed immunoreactive receptor protein. In addition, it demonstrates the type of information that can be obtained if an immunoblot assay were to be used as a component of a screening method to analyze samples from patients with defects of the AR. Finally, the study suggests that in some androgen-resistant patients, defects outside the AR open reading frame may result in major alterations of AR expression.
[show abstract][hide abstract] ABSTRACT: Mutations in the androgen receptor (AR) gene cause a range of phenotypic abnormalities of male sexual development. At one end of the spectrum are individuals with complete androgen insensitivity (complete testicular feminization) who exhibit normal breast development and female external genitalia. At the other extreme are individuals with male phenotypes that are characterized by either subtle undervirilization or infertility. Studies in a number of different laboratories have identified mutations of the AR gene in subjects with androgen resistance syndromes. Defects that interrupt the AR open-reading frame have been traced to a number of distinct types of genetic alterations, have been identified in widely separated segments of the AR gene, and are invariably associated with the phenotype of complete androgen insensitivity. By contrast, mutations that cause single amino acid substitutions within the AR are localized to the DNA- or ligand-binding domains of the receptor protein and have been associated with the full range of androgen-resistant phenotypes. Regardless of the nature of the mutation, functional studies and assays of AR abundance suggest that the phenotypic abnormalities that result from mutation of the AR are the result of the impairment of receptor function, decreases in receptor concentration, or both.
Recent Progress in Hormone Research 02/2002; 57:181-94.
[show abstract][hide abstract] ABSTRACT: ndrogens exert their effects in mediating the develop- ment of the normal male phenotype via a single re- ceptor protein, the androgen receptor (AR), which is encoded on the X chromosome. Abnormalities that alter the function of this receptor result in a range of abnormalities of male phenotypic development. These phenotypes range from that of normal females (complete testicular feminization, com- plete androgen insensitivity) to those that are characterized by only minor degrees of undervirilization and/or infertility. The defects of receptor function that have been character- ized fall into two major categories. The first are those that disrupt the primary sequence of the AR. These mutations can be due to the introduction of premature termination codons, frameshift mutations, deletions or insertions, or alterations of RNA splicing. The ARs that are produced as a result of these genetic alterations are uniformly associated with complete androgen resistance. The second, and most common type of AR mutation, is that which is caused by single amino acid substitutions within the AR protein. In contrast to the preceding category, mutations of this type may result in the full spectrum of androgen resistant phenotypes. These mutations cluster in two impor- tant domains of the receptor protein: the DNA- and the hormone- binding domains. Substitutions in the DNA-bind- ingdomain act to impair the ability of the AR to recognize target sequences within or adjacent to androgen-responsive genes. The degree of impairment of DNA binding is directly related to the degree of impairment of receptor function. Amino acid replacements within the hormone-binding do- main can have a range of effects on the binding of ligand by the AR. In some instances, the capacity to bind hormone is completely abolished, while in other instances, only subtle qualitative abnormalities of ligand binding can be detected. In all mutants containing amino acid substitutions within the hormone-binding domain, the ability to form stable AR- hormone complexes determines the amount of receptor func- tion that remains. The phenotype that is observed does not appear to cor- relate with the identity of the residue that is mutated or its replacement. Instead, the phenotype appears to be a reflec- tion of the degree to which androgen action is impaired. As the level of mutant AR protein expressed is similar to that observed in normal subjects, this usually reflects the degree to which AR function is altered. The male hormones, androgens, control wide range of processes in the male during embryonic development and in postnatal life. During embryogenesis, the concerted actions of testosterone (T) and 5a-dihydrotestosterone (DHT) are critical to the virilization of the Wolffian duct structures and the external genitalia. Genetic defects that impair androgen synthesis or action will result in abnormalities of male phe- notypic development in 46, XY individuals with testes (male pseudohermaphroditism). Among the potential etiologies, defects of the AR are among the most common.