Tracing the Emergence of a Novel Sex-Determining Gene in Medaka,Oryzias luzonensis

Abstract

Three sex-determining (SD) genes, SRY (mammals), Dmy (medaka), and DM-W (Xenopus laevis), have been identified to date in vertebrates. However, how and why a new sex-determining gene appears remains unknown, as do the switching mechanisms of the master sex-determining gene. Here, we used positional cloning to search for the sex-determining gene in Oryzias luzonensis and found that Gsdf Y (gonadal soma derived growth factor on the Y chromosome) has replaced Dmy as the master sex-determining gene in this species. We found that Gsdf Y showed high expression specifically in males during sex differentiation. Furthermore, the presence of a genomic fragment that included Gsdf Y converts XX individuals into fertile XX males. Luciferase assays demonstrated that the upstream sequence of Gsdf Y contributes to the male-specific high expression. Gsdf is downstream of Dmy in the sex-determining cascade of O. latipes, suggesting that emergence of the Dmy-independent Gsdf allele led to the appearance of this novel sexdetermining gene in O. luzonensis. IN most vertebrates, sex is determined genetically. Mammals and birds with cytogenetically well-differentiated sex chromosomes have sex determination systems that differ between the taxonomic classes but not within them (Solari 1994). In mammals, for example, the sex-determining (SD) gene SRY/Sry on the Y chromosome has a universal role in sex determination (Gubbay et al. 1990;Sinclair et al. 1990;Koopman et al. 1991;Foster et al. 1992). By contrast, some fish groups, such as salmonids, sticklebacks, and Oryzias fishes, have sex chromosomes that differ among closely related species (Devlin and Nagahama 2002;Woram et al. 2003;Takehana et al. 2007a;Ross et al. 2009).A DM-domain gene, Dmy, was the first SD gene identified in a nonmammalian vertebrate, the fish medaka Oryzias latipes (Matsuda et al. 2002(Matsuda et al. , 2007. In this species, the term Y chromosome is employed to refer to a recombining chromosome that carries the male-determining gene Dmy, and X is used for the homologous chromosome; these are not a heteromorphic pair. This gene is conserved among all wild populations of O. latipes examined to date . The closely related species O. curvinotus also has Dmy on its Y chromosome, which is orthologous to the O. latipes Y chromosome (Matsuda et al. 2003). However, Dmy has not been detected in any other type of fish, including other Oryzias fishes (Kondo et al. 2003). Analysis of the Y-specific region of the O. latipes sex chromosome has demonstrated that Dmy arose from duplication of the autosomal Dmrt1 gene (Nanda et al. 2002;Kondo et al. 2006). This Dmrt1 duplication is estimated to have occurred within the last 10 million years in a common ancestor of O. latipes, O. curvinotus, and O. luzonensis. In O. luzonensis, however, no functional duplicated copy of Dmrt1 has been detected (Kondo et al. 2003) (Figure 5A).O. luzonensis possesses an XX-XY system, which is homologous to an autosomal linkage group (LG 12) and Uwa 1985). In the d-rR strain, the wild-type alle...

Co-option of Sox3 as the male-determining factor on the Y chromosome in the fish Oryzias dancena

“…Scale bars, 1 mm. found in O. latipes 13 and the other is Gsdf in O. luzonensis 15 ( Supplementary Fig. 8).…”

“…Sex chromosome turnover is widespread in non-mammalian vertebrates, especially in fish 9,10 , suggesting that many different genes can act as the sexdetermining signal. Several different sex-determining genes or candidates have been identified on the sex chromosomes of chicken 11 , frog 12 and fish [13][14][15][16][17][18] , but only rarely have they been formally demonstrated or their downstream pathways analysed. It is uncertain whether these genes are the major contributors to sex chromosome turnover, and it is unclear how these novel signals are integrated into the ancestral molecular pathway of sex determination.…”

Gonadal soma-derived factor (gsdf), a TGF-beta superfamily gene, induces testis differentiation in the teleost fish Oreochromis niloticus

“…In the present study, the 460 tilapia gsdf mRNA was dominantly detected in the male gonads in advance of dmrt1 detection, and both gsdf and dmrt1 were localized specifically in germ cell-surrounding cells in morphologically undifferentiated gonads. In medaka, the expression profiles of sex differentiation-related genes suggest that gsdf is downstream of the sex-determining gene dmy/dmrt1Yb during sex determination, but it is expressed earlier than dmrt1 in the Sertoli cell 465 lineage (Kikuchi and Hamaguchi, 2013;Myosho et al, 2012;Shibata et al, 2010;Takehana et al, In the tilapia, upregulation of other genes involving testis differentiation, such as anti-Mullerian hormone (after 15 dph) and sox9 (after 35 dph) were delayed far behind gsdf (5 dph) and dmrt1 (6 dph) upregulation in XY gonads (Ijiri et al, 2008), suggesting important roles of gsdf and 470 dmrt1 as initial switches to promote testis differentiation.…”

“…A knock-out analysis for the gsdf gene must be conducted to provide a conclusive answer to this question. In O. luzonensis, a subspecies of medaka that lacks dmy/dmrt1Yb (Kondo et al, 2004;Matsuda et al, 2003), gsdfY (gonadal soma-derived factor on the Y chromosome) was recently identified as a 500 sex-determining gene (Myosho et al, 2012). Further functional analysis indicated that a transgene with a region including the gsdfY genomic fragment in XX O. luzonensis and O.…”

“…In addition, dmy/dmrt1Yb is only found in two subspecies among more than 20 related species of medaka (Matsuda et al, 2003). Over the past several years, sex-determining genes 25 have been identified in numerous lower vertebrates, including dm-W in the amphibian Xenopus laevis (Yoshimoto et al, 2008) and gsdfY, amhY, sdY, amhr2 and sox3 in a suite of species of teleost fish, i.e., a medaka-related fish, (Oryzias luzonensis) (Myosho et al, 2012), Patagonian pejerrey (Odontesthes bonariensis) (Hattori et al, 2012), rainbow trout (Onchorhynchus mykiss) (Yano et al, 2012), tiger pufferfish (Takifugu rubripes) (Kamiya et al, 2012), and another 30 medaka-related fish (Oryzias dancena) (Takehara et al, 2014), respectively. Thus, in contrast to many other developmental processes, evolutionary diversity exists in the initial switches of sex determination.…”