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Fish Physiol Biochem (2014) 40:849864 DOI 10.1007/s10695-013-9891-6
Characterization of sea bass FSHb 50 anking region: transcriptional control by 17b-estradiol
Borja Muriach Manuel Carrillo Silvia Zanuy
Jos Miguel Cerd-Reverter
Received: 10 July 2013 / Accepted: 13 November 2013 / Published online: 23 November 2013 Springer Science+Business Media Dordrecht 2013
Abstract The sea bass follicle-stimulating hormone 50 anking region (sbFSHb 50 FR) was cloned and characterized in order to study the molecular mechanisms underlying transcriptional regulation of the sbFSHb gene. Analysis of the *3.5 kb of this region revealed the presence of several putative cis-acting elements, including steroid hormone response elements, cAMP response elements, pituitary-specic transcription factor response elements, activator protein-1 response elements and TATA sequence. Deleted constructs containing *3.5 kb of the sbFSHb 50 FR fused to a luciferase reporter gene were transiently transfected into human embryonic kidney (HEK 293) and mouse mature gonadotrope (LbT2)
cell lines. The sbFSHb 50 FR was efciently expressed under basal conditions in LbT2 but not in HEK 293, pointing to both positive and negative regulatory elements. In order to elucidate the estrogen-mediated sbFSHb transcriptional activity, in vitro treatments with 17b-estradiol were carried out on primary
cultures of pituitary cells and LbT2 cells transiently expressing luciferase under the control of sbFSHb 50
FR. Overall, these results demonstrate that 17bestradiol inhibits sbFSHb gene expression directly at the level of the pituitary. However, it was also shown that estrogen did not induce changes of the sbFSH promoter-directed luciferase activity, suggesting that sbFSHb 50FR (*3.5 kb) activity is cell type dependent and its estrogen regulation could require cis-acting elements located upstream of the promoter region, which is characterized in this article.
Keywords FSHb Estrogen receptor
Promoter analysis Fish Reproduction LbT2
Introduction
In sh, as in other vertebrates, the reproductive axis is mainly controlled by gonadotropin hormones, i.e., the follicle-stimulating hormone (FSH) and luteinizing hormone (LH) (Kawauchi et al. 1989; Querat 1994). Both hormones are involved in the regulation of gametogenesis. However, FSH exerts its main function during the early stages of reproduction, vitello-genesis and gonadal development, whereas LH regulates the nal maturation of the oocytes and spermiation/ovulation (Zohar et al. 2010; Levavi-Sivan et al. 2010). Both gonadotropins, together with thyroid-stimulating hormone, belong to a family of
B. Muriach M. Carrillo S. Zanuy
J. M....