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See detailGene knock down via antisense oligonucleotides to the steroid receptor coactivator SRC-1 modulates testosterone-dependent male sexual behavior and neural gene expression
Charlier, Thierry ULg; Ball, Gregory F; Balthazart, Jacques ULg

in Hormones & Behavior (2004), 46

Studies of eukaryotic genome expression demonstrate the importance of steroid receptor coactivators in mediating efficient gene transcription. Little is know about the physiological role of these ... [more ▼]

Studies of eukaryotic genome expression demonstrate the importance of steroid receptor coactivators in mediating efficient gene transcription. Little is know about the physiological role of these coactivators in vivo. We recently showed that the Steroid Receptor Coactivator SRC-1 is densely expressed in steroid-sensitive brain areas in birds and its expression is steroid-dependent and sexually differentiated. We tested the role of SRC-1 in the activation by testosterone of male sexual behavior in quail. Daily injections of LNA antisense oligonucleotides in the third ventricle (AS group) significantly reduced the expression of male copulatory behavior in response to exogenous testosterone compared to control animals (Ctrl group) receiving the vehicle alone or scrambled LNA. Sexual behavior was restored and even enhanced within 48 hours after interruption of LNA injection (ASSC group). Western blot analysis confirmed the decrease of SRC-1 expression in AS animals and suggested an over-expression of the coactivator in ASSC animals. The effect of SRC-1 knock down on behavior was correlated with a reduced volume of the medial preoptic nucleus (POM) defined by Nissl-staining and aromatase immunohistochemistry. In addition, the amount of aromatase-immunoreactive material in POM, defined as the relative optical density of the aromatase immunoreactivity multiplied by the percentage of surface covered within the nucleus and by the total POM volume of the POM, was decreased in the AS compared to the Ctrl group, suggesting a blockade of aromatase transcription. Together, these data indicate that SRC-1 functions as a critical regulatory molecule in the brain that modulates steroid-dependent gene transcription and behavior. [less ▲]

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See detailSex differences in the distribution of the steroid receptor coactivator SRC-1 in the song control nuclei of male and female canaries
Charlier, Thierry ULg; Balthazart, Jacques ULg; Ball, Gregory F

in Brain Research (2003), 959(2), 263-274

The steroid receptor coactivator SRC-1 modulates ligand-dependent transactivation of several nuclear receptors, including the receptors for sex steroid hormones. The distribution of SRC-1 transcripts was ... [more ▼]

The steroid receptor coactivator SRC-1 modulates ligand-dependent transactivation of several nuclear receptors, including the receptors for sex steroid hormones. The distribution of SRC-1 transcripts was analyzed here by in situ hybridization in coronal sections through the brain of male and female canaries. A broad but heterogeneous distribution of SRC-1 transcripts was observed with high numbers of densely labeled cells being present in many steroid-sensitive areas including the medial preoptic nucleus, several hypothalamic nuclei, five song control nuclei (HVc, the lateral and medial portion of the magnocellular nucleus of the anterior neostriatum, area X and the nucleus uvaeformis) and several catecholaminergic areas (area ventralis of Tsai, substantia nigra, locus coeruleus). The volume of two song control nuclei, HVc and area X were reconstructed based on the boundaries of the cell groups exhibiting a denser SRC-1 expression as compared to the surrounding areas. Sex differences in the expression of SRC-1 were also detected in several song control nuclei. In particular, the volume of HVc based on the high density of SRC-1 expression was significantly larger in males than in females. The effect of steroids on the song control system could be, at least in part, indirect and result from a modulation by steroids of the catecholaminergic inputs to the song control nuclei. The presence of the steroid receptor coactivator SRC-1 in the telencephalic song control nuclei and in the catecholaminergic cell groups that innervate the song system supports the idea that SRC-1 expression could play an active role in the control of singing behavior by modulating estrogen and androgen receptor action at both locations. (C) 2002 Elsevier Science B.V. All rights reserved. [less ▲]

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See detailModulation of steroid activity by transcription coactivators in songbirds
Charlier, Thierry ULg; Auger, Catherine J; Balthazart, Jacques ULg et al

in Hormones & Behavior (2003), 44

Songbirds have developed a specialized, steroid-dependent telencephalic vocal control system for the production of learned vocalization. Recent progress in the study of the mechanisms by which steroid ... [more ▼]

Songbirds have developed a specialized, steroid-dependent telencephalic vocal control system for the production of learned vocalization. Recent progress in the study of the mechanisms by which steroid receptors act on the eukaryotic genome has highlighted the role of a newly discovered protein family, the Nuclear Receptor Coactivators. More specifically, the CREB-binding protein (CBP) and the Steroid Receptor Coactivator-1 (SRC-1) have been shown to be actively involved in mediating steroid hormone action in the developing rat brain. The distribution of the coactivator SRC-1 was analyzed in canaries by in situ hybridization. A very broad but heterogeneous distribution of the transcript was observed, mainly in steroid-sensitive areas of the hypothalamus, the song control system and several catecholaminergic areas. The presence of SRC-1 in these regions was also confirmed by immunocytochemistry. A similar very high concentration of the coactivator CBP protein was also found in steroid-sensitive areas of the hypothalamus and in the song system. Sex differences in SRC-1 mRNA concentration were detected in HVC and in area X. Moreover, preliminary data obtained independently in starlings (CBP) and in quail (SRC-1) suggest that the expression of coactivators is regulated by steroids as well as by photoperiod. The presence of these steroid receptor coactivators in the telencephalic song control nuclei and in catecholaminergic cell groups that innervate the song system and their possible regulation by photoperiod and/or steroids support the idea that SRC-1 and CBP could play an important role in the control of singing behavior by modulating estrogen and androgen receptor action. [less ▲]

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See detailCloning and identification of functional domains in quail Brain aromatase
Charlier, Thierry ULg; Baillien, Michelle; Ball, Gregory F. et al

Poster (2003)

Recent evidence indicates that aromatase activity (AA) in the hypothalamus is not only modulated by slow (hours to days) genomic actions but also through fast (seconds to minutes) non-genomic mechanisms ... [more ▼]

Recent evidence indicates that aromatase activity (AA) in the hypothalamus is not only modulated by slow (hours to days) genomic actions but also through fast (seconds to minutes) non-genomic mechanisms. We recently showed that Calcium (Ca2+)-dependent phosphorylations catalyzed by multiple protein kinases including PKC, and possibly PKA and CAMK, rapidly down-regulate AA in quail hypothalamic homogenates. Western blotting experiments also indicated that phosphorylations affect the aromatase molecule itself but it was impossible to fully characterize the putative phosphorylation sites on the quail enzyme because its sequence was unknown. We therefore cloned and sequenced the quail brain aromatase. We identified a 1541-bp open reading frame that encodes a predicted 490-amino acid protein containing all functional domains previously described in mammalian and other avian aromatases. Multiple motifs match consensus sequences corresponding to several protein kinases including those that were shown to affect AA during pharmacological experiments with specific kinase inhibitors (e.g., PKC, PKA, MAPK, Myosine light chain kinase, Tyr. kinase). Another potential control pathway of AA, independent from phosphorylations, could involve a direct control by Ca2+-dependent calmodulin (CAM), as suggested by the identification in Western blots of CAM on purified aromatase from quail hypothalamic homogenates. Accordingly, two Ca2+-dependent calmodulin binding motifs (1-8-14b) as defined by Rhoads and Friedberg (FASEB, 1997, 11:331-340) are present and conserved in most vertebrates including quail aromatase. These results suggest that the phosphorylation of some residues as well as the direct binding of calmodulin on the aromatase protein represent part of the mechanism(s) underlying the rapid changes in AA. [less ▲]

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See detailSeasonal changes in the songbird brain are modulated by song performance via testosterone-dependent and independent action
Sartor, Jennifer J; Charlier, Thierry ULg; Pytte, Caroline L et al

Poster (2002)

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See detailSex differences in the distribution of the steroid receptor-coactivator-1 in the canary brain
Charlier, Thierry ULg; Ball, Gregory F; Balthazart, Jacques ULg

in Hormones & Behavior (2002), 41

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See detailSteroid receptor coactivator SRC-1 exhibits high expression in steroid-sensitive brain areas regulating reproductive behaviors in the quail brain.
Charlier, Thierry ULg; Lakaye, Bernard ULg; Ball, Gregory F et al

in Neuroendocrinology (2002), 76(5), 297-315

The steroid receptor coactivator SRC-1 modulates ligand-dependent transactivation of several nuclear receptors, including the receptors for sex steroid hormones. Reducing the expression of SRC-1 by ... [more ▼]

The steroid receptor coactivator SRC-1 modulates ligand-dependent transactivation of several nuclear receptors, including the receptors for sex steroid hormones. Reducing the expression of SRC-1 by injection of specific antisense oligonucleotides markedly inhibits the effects of estrogens of the sexual differentiation of brain and behavior in rats and inhibits the activation of female sexual behavior in adult female rats. SRC-1 thus appears to be involved in both the development and activation of sexual behavior. In the Japanese quail brain, we amplified by RT-PCR a 3,411-bp fragment extending from the HLH domain to the activating domain-2 of the protein. The quail SRC-1 is closely related to the mammalian (m) SRC-1 and contains a high proportion of GC nucleotides (62.5%). Its amino acid sequence presents 70% identity with mammalian SRC-1 and contains the three conserved LXXLL boxes involved in the interaction with nuclear receptors. In both males and females, RT-PCR demonstrates a similarly high level of expression in the telencephalon, diencephalon, optic lobes, brain stem, spinal cord, pituitary, liver, kidney, adrenal gland, heart, lung, gonads and gonoducts. Males express significantly higher levels of SRC-1 in the preoptic area-hypothalamus than females. In both sexes, lower levels of expression are observed in the cerebellum and muscles. In situ hybridization utilizing a mixture of four digoxigenin-labeled oligonucleotides confirms at the cellular level the widespread distribution of SRC-1 mRNA in the brain and a particularly dense expression in steroid-sensitive areas that play a key role in the control of male sexual behavior. These data confirm the presence and describe for the first time the SRC-1 distribution in the brain of an avian species. They confirm its broad, nearly ubiquitous, distribution in the entire body including the brain as could be expected for a coactivator that regulates to the action of many nuclear receptors. However this distribution is heterogeneous in the brain and sexually differentiated in at least some areas. The very dense expression of SRC-1 in limbic and mesencephalic nuclei that are associated with the control of male sexual behavior is consistent with the notion that this coactivator plays a significant role in the activation of this behavior. [less ▲]

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See detailCloning and distribution of steroid receptor coactivator SRC-1 in quail.
Charlier, Thierry ULg; Lakaye, Bernard ULg; Ball, Gregory F. et al

Poster (2001)

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