Browsing by Author "Socorro, S."
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- A cDNA for European sea bass (Dicentrachus labrax) 11 beta-hydroxylase: Gene expression during the thermosensitive period and gonadogenesisPublication . Socorro, S.; Martins, Rute S. T.; Deloffre, Laurence A. M.; Mylonas, Constantinos C.; Canario, Adelino V. M.Steroid P450 11 -hydroxylase, encoded by the CYP11B gene, is a key mitochondrial enzyme for the production of 11-oxygenated androgens, which have been shown to be potent masculinising steroids in several Wsh species. In this study we have isolated a CYP11B cDNA of 1903 base pairs from the testis of European sea bass (Dicentrarchus labrax) encoding a predicted protein of 552 amino acids. The amino acid identities to other vertebrate 11 -hydroxylase proteins ranged from 66% to 72% to other Wsh; 45% to amphibian and 35–39% to mammalian. Southern blot indicated that a single CYP11B gene is present. Northern blot analysis detected two transcripts in testis and head kidney, one of the same size of the isolated clone and the other of 3.9 kb. Reverse transcriptase-polymerase chain reaction showed abundant mRNA expression only in testis and head kidney, being residual in a range of other tissues. Expression of CYP11B and CYP19A (which encodes for ovarian aromatase) was detected from at least 4 days post-hatching and did not appear to be aVected by rearing temperature (15 and 20 °C) during the Wrst 60 days, a period in which high temperatures promote masculinisation in European sea bass. Throughout, gonadogenesis (60–300 dph), a highly dimorphic pattern of CYP11B expression was consistent with a role of this gene in testicular development.
- Identification of androgen receptor variants in testis from humans and other vertebratesPublication . Laurentino, S. S.; Pinto, Patricia IS; Tomas, J.; Cavaco, J. E. B.; Sousa, M.; Barros, A.; Power, Deborah; Canario, Adelino V. M.; Socorro, S.The androgen receptor (AR) is a ligand-activated transcription factor member of the nuclear receptor superfamily. The existence of alternatively spliced variants is well recognised for several members of this superfamily, most of them having functional importance. For example, several testicular oestrogen receptor variants have been suggested to play a role in the regulation of spermatogenesis. However, information on AR variants is mostly related to cancer and androgen insensitivity syndrome (AIS) cases. The objective of this study was to investigate the expression of AR variants in the testis from humans and other vertebrates. Four AR variants [ARD2Stop, ARD223Stop, ARD3 and ARD4(120)] were identified in human testis. ARD2Stop and ARD3, with exon 2 or 3 deleted, respectively, were also expressed in human liver, lung, kidney and heart. In addition, ARD2Stop was expressed in rat and gilthead seabream testis, while an ARD3 was detected in African clawed frog testis. This is the first report revealing the existence of AR variants in the testis of evolutionarily distant vertebrate species and in nonpathological tissues. These data suggest the functional importance of these novel AR forms and demonstrate a complexity in AR signalling that is not exclusive of pathological conditions.
- Immunohistochemical detection of estrogen receptors in fish scalesPublication . Pinto, Patricia IS; Estêvão, M. Dulce; Redruello, Begoña; Socorro, S.; Canario, Adelino V. M.; Power, DeborahCalcium mobilization from internal stores, such as scales, induced by 17b-estradiol during sexual maturation in salmonids is well documented. This calcium mobilization from scales is proposed to be mediated by the estrogen receptor (ER). However, the ER subtypes involved and signaling mechanisms responsible for this effect remain to be fully characterized. In the present study, we have localized ERa, ERba and ERbb proteins in juvenile and adult sea bream (Sparus auratus) and Mozambique tilapia (Oreochromis mossambicus) scales by immunohistochemistry with sea bream ER subtype specific antibodies. The three ERs were detected in isolated or small groups of round cells, in the basal layer of the scales of both juvenile and adult fish and the localization and signal intensity varied with the species and age of the animals. The ERs may be co-localized in cells of the scale posterior region that expressed tartrateresistant acid phosphatase (TRAP), a marker for osteoclasts. These results suggest that the calcium mobilizing action of 17b-estradiol on fish scales is via its direct action on ERs localized in osteoclasts.
- Structure, tissue distribution and estrogen regulation of splice variants of the sea bream estrogen receptor alpha genePublication . Pinto, Patricia IS; Teodósio, H. R.; Socorro, S.; Power, Deborah; Canario, Adelino V. M.Estrogen actions are mainly mediated by specific nuclear estrogen receptors (ERs), for which different genes and a diversity of transcript variants have been identified, mainly in mammals. In this study, we investigated the presence of ER splice variants in the teleost fish gilthead sea bream (Sparus auratus), by comparison with the genomic organization of the related species Takifugu rubripes. Two exon2-deleted ERα transcript variants were isolated from liver cDNA of estradiol-treated fish. The ΔE2 variant lacks ERα exon 2, generating a premature termination codon and a putative C-terminal truncated receptor, while the ΔE2,3* variant contains an in-frame deletion of exon 2 and part of exon 3 and codes for a putative ERα protein variant lacking most of the DNA-binding domain. Both variants were expressed at very low levels in several female and male sea bream tissues, and their expression was highly inducible in liver by estradiol-17β treatment with a strong positive correlation with the typical wild-type (wt) ERα response in this tissue. These findings identify novel estrogen responsive splice variants of fish ERα, and provide the basis for future studies to investigate possible modulation of wt-ER actions by splice variants.