Characterisation and expression of Sox9 in the leopard gecko, Eublepharis macularius

Since the discovery of the sex-determining gene, Sry, a number of genes hav e been identified which are involved in sex determination and gonadogenesis in mammals. Although Sry is known to be the testis-determining factor in m ammals, this is not the case in non-mammalian vertebrates. Sox9 is another gene that has been shown to have a male-specific role in sex determination, but, unlike Sry, Sox9 has been shown to be involved in sex determination i n mammals, birds, and reptiles. This is the first gene to be described that has a conserved role in sex determination in species with either chromosom al or environmental sex-determining mechanisms. Many reptiles do not have s ex chromosomes but exhibit temperature-dependent sex determination (TSD). S ox9 has been shown to be expressed in both turtle and alligator during gona dogenesis. To determine if Sox9 also has a role in a gecko species with TSD , we studied gonadal expression of Sox9 during embryonic development of the Leopard gecko (Eublepharis macularius). Gecko Sox9 was found to be highly conserved at the nucleotide level when compared to other vertebrate species including human, chick, alligator, and turtle. Sox9 was found to be expres sed in embryos incubated at the male-producing temperature (32.5 degreesC) as well as in embryos incubated at the female-producing temperatures (26 an d 34 degreesC), Northern blot analysis showed that Sox9 was expressed at bo th temperatures from morphological stages 31 to 37. mRNA in situ hybridisat ion on isolated urogenital systems showed expression at both female- and ma le-producing temperatures up to stage 36. After this stage, no expression w as seen in the female gonads but expression remained in the male. These dat a provide further evidence that Sox9 is an essential component of a testis- determining pathway that is conserved in species with differing sex-determi ning mechanisms. (C) 2001 Wiley-Liss, Inc.