Genome-wide identification, phylogeny, and gonadal expression of fox genes in Nile tilapia, Oreochromis niloticus

Abstract

The fox genes play important roles in various biological processes, including sexual development. In the present study, we isolated 65 fox genes, belonging to 18 subfamilies named A-R, from Nile tilapia through genome-wide screening. Twenty-four of them have two or three (foxm1) copies. Furthermore, 16, 25, 68, and 45 fox members were isolated from nematodes, protochordates, teleosts, and tetrapods, respectively. Phylogenetic analyses indicated fox gene family had undergone three expansions parallel to the three rounds of genome duplication during evolution. We also analyzed the clustered fox genes and found that apparent linkage duplication existed in teleosts, which further supported fish-specific genome duplication hypothesis. In addition, species- and lineage-specific duplication is another reason for fox gene family expansion. Based on the four pairs of XX and XY gonadal transcriptome data from four critical developmental stages, we analyzed the expression profile of all fox genes and identified sexually dimorphic fox genes at each stage. All fox genes were detected in gonads, with 15 of them at the background expression level (total read per kb per million reads, RPKM < 10), 29 at moderate expression level (10 < total RPKM < 100), and 21 at high expression level (total RPKM > 100). There are 27, 24, 28, and 9 sexually dimorphic fox genes at 5, 30, 90, and 180 days after hatching (dah), respectively. foxq1a, foxf1, foxr1, and foxr1 were identified as the most differentially expressed genes at each stage. foxl2 was characterized as XX-dominant gene, while foxd5, foxi3, foxn3, foxj1a, foxj3b, and foxo6b were characterized as XY-dominant genes. qPCR and in situ hybridization of foxh1 and foxj1a were performed to confirm the expression profiles and to validate the transcriptome data. Our results suggest that fox genes might play important roles in sex determination and gonadal development in teleosts.

Genome-wide identification, evolution and expression analysis of nuclear receptor superfamily in Nile tilapia, Oreochromis niloticus

“…Extensive studies on the repertoires of this family have indicated increasing NR members from cnidarians to tetrapods, with 17 NRs identified in the sea anemone (Reitzel and Tarrant, 2009), 21 in the fruit fly (Maglich et al, 2001), 33 in the lancelet (Schubert et al, 2008), 48 in the human and 49 in the mouse (Zhang et al, 2004). The increasing pattern is similar to other gene families, such as Homeobox (Hox) (Itoh and Ornitz, 2004) and Forkhead box (Fox) (Yuan et al, 2014). The whole genome duplication (WGD) is considered to be the driving force behind the expansion of gene families.…”

“…NRs with RPKM b 1.25 in each transcriptome or the total RPKM b 10 in 8 transcriptomes from four developmental stages were considered as background expression; Total RPKM N 100 were considered as high expression and total RPKM between 10 and 100 as moderate expression. The statistical criteria of "FDR ≤ 10 −2 " and "|log 2 (XX_RPKM/XY_RPKM) | ≥ 1" were used to identify XX/XY-enhanced NRs (Tao et al, 2013;Yuan et al, 2014). The NRs, which identified as XX/XY-enhanced NRs in at least three stages, were considered as XX/XY-dominant genes.…”

“…The fixation, embedding, and sectioning of dissected gonads and ISH were performed as described previously (Kobayashi et al, 2000;Yuan et al, 2014). Probes of sense and antisense digoxigenin (DIG)-labeled RNA strands were transcribed in vitro from linearized pGEM-T easy-ERα/ERβ-A/ERβ-B cDNA using a RNA labeling kit (Roche, Germany).…”

foxr1is a novel maternal-effect gene in fish that regulates embryonic cell growth viap21andrictor

“…The essentialness of foxr1 was suggested by the wide-ranging presence of this gene in most vertebrates and the retention of a single copy in most teleosts despite multiple whole genome duplication events, but its biological function is still largely unknown. Previous reports have demonstrated the predominant expression of foxr1 mRNA in the ovary of medaka, eel, and tilapia[8,11,13], but in the male germ cells and spermatids in mouse and human[24]. It was further shown to be abundantly expressed in the early cleavage and gastrula stages of Xenopus embryos, but absent in post-gastrula stages due to rapid degradation of its mRNA, indicating that it is a maternally-inherited transcript[25].…”

“…In mammals, knockouts of several fox genes, including foxa2 , foxo1 , and foxf1 , result in embryonic lethality due to defects in development of different organs ([5–7]). In reproduction, a recent transcriptomic study in the Nile tilapia, Oreochromis niloticus , showed that more than 50 fox genes were expressed in the gonads, and some of these, including foxl2 , foxo3 , and foxr1 , were specific to XX females[8]. foxl2 and its relatives are known to be key players in ovarian differentiation and oogenesis in vertebrates; it is essential for mammalian ovarian maintenance and through knockout experiments, it was demonstrated that foxl2 is a critical regulator of sex determination by regulating ovary development and maintenance also in Nile tilapia, medaka, and zebrafish[9].…”