Additionally we used the predicted

Additionally, we used the predicted Cyp19a1b amino MRS 2768 tetrasodium salt mg sequence from a number of bony fish species to infer their genetic relatedness and found that R. quelen occupies an ancient position within the Siluriformes group. A phylogenetic tree was constructed and it included both full-length and partial fish Cyp19a1b protein sequences available in GenBank. The highest sequence similarity of R. quelen Cyp19a1b was with the Siluriformes, Ictalurus punctatus (88%), Silurus meridionalis (87%), and Pelteobagrus fulvidraco (86%), consistent with the known phylogenetic relationships of the catfish (Fig. 2). The mRNA of cyp19a1b was detected in all studied tissues including head and posterior kidney, liver and gonads, in addition to the expected expression in pituitary and brain regions in both male and female (Fig. 3). Females and males presented similar levels of mRNA cyp19a1b in pituitary, hypothalamus, telencephalon, anterior and posterior kidney tissues, while levels in liver and gonad were significantly lower than pituitary and hypothalamus. Males presented higher levels of cyp19a1b than females in pituitary and posterior kidney. We observed sexually-dimorphic effects of E2 injection. In E2-treated males the expression of cyp19a1b was down-regulated in the head kidney (Fig. 4A), while in E2-treated females the expression in the pituitary and head kidney were up-regulated (Fig. 4B).
Discussion To the best of our knowledge, the present study is the first to report the nucleotide sequences of R. quelen brain aromatase cyp19a1b. R. quelen cyp19a1b has high identity (~88%) to three other catfish species, Ictalurus punctatus, Silurus meridionalis and Pelteobagrus fulvidraco. The R. quelen deduced amino acid sequence, with 347 aa of length, corresponds to 60–70% of the total Cyp19a1b protein for several teleosts (Blázquez and Piferrer, 2004; Chaube et al., 2015; Gelinas et al., 1998; Strobl-Mazzulla et al., 2005). Our predictions suggest that R. quelen cyp19a1b presents I-helix, Ozol's peptide, aromatase-specific conserved and heme-binding regions, which are the supposed functional regions of aromatase, confirming that cyp19a1b is a highly-conserved gene. The presence of these conserved regions may indicate the conservation of the enzymatic activity. We also examined potential sex differences in cyp19a1b mRNA levels in pituitary, telencephalon, hypothalamus, liver, gonads and head and posterior kidneys. The presence of cyp19a1b in brain regions and pituitary is well documented for other fish species, for example, Carassius auratus (Gelinas et al., 1998), Oncorhynchus mykiss (Menuet et al., 2003), Danio rerio (Menuet et al., 2005; Pellegrini et al., 2007), Hippoglossus hippoglossus (Matsuoka et al., 2006), Salaria pavo (Gonçalves et al., 2008), Heteropneutes fossilis (Chaube et al., 2015). In the teleost brain aromatase is only expressed in radial glial cells (Coumailleau et al., 2015; Diotel et al., 2010). Sexually dimorphic expression in telencephalon and hypothalamus has been observed in Danio rerio (Goto-Kazeto et al., 2004) and Odontesthes bonariensis (Strobl-Mazzulla et al., 2005) with males showing higher expression than females. It is known that teleost have high sexually dimorphic neurogenic capacity, in part due to the high aromatase activity in the different brain regions (Ampatzis et al., 2012; Coumailleau et al., 2015). The expression of cyp19a1b in several peripheral tissues (e.g., liver, head kidney, testis, ovary, gill, etc.) was reported for several species of fish (Chang et al., 2005; Chaube et al., 2015; Kwon et al., 2001; Shanthanagouda et al., 2012; Strobl-Mazzulla et al., 2005; Tang et al., 2010; Van Nes et al., 2005), although this normally occurs in lower levels than in brain or pituitary. However, there is no consensus among the species, sex or tissues where brain aromatase is expressed, which leads to an unclear role to cyp19a1b expression in these tissues. It is suggested that the estrogen produced in peripheral organs can act in local targets modulating mechanisms not related directly to reproduction, as growth and differentiation of tissues (Blázquez and Piferrer, 2004; Piferrer and Blázquez, 2005). Likewise, is suggested that the presence of estrogen receptors in different organs is associated with different functions or tissue-specific mechanisms of regulation (Marlatt et al., 2012; Strobl-Mazzulla et al., 2008). In this way, the presence of cyp19a1b in peripheral tissues can modulate, in a tissue-specific way, the transcription of several genes by the action of estrogen receptors.