A Trans-Species Missense SNP in Amhr2 Is Associated with Sex Determination in the Tiger Pufferfish, Takifugu rubripes (Fugu)

Abstract

Heterogametic sex chromosomes have evolved independently in various lineages of vertebrates. Such sex chromosome pairs often contain nonrecombining regions, with one of the chromosomes harboring a master sex-determining (SD) gene. It is hypothesized that these sex chromosomes evolved from a pair of autosomes that diverged after acquiring the SD gene. By linkage and association mapping of the SD locus in fugu (Takifugu rubripes), we show that a SNP (C/G) in the anti-Müllerian hormone receptor type II (Amhr2) gene is the only polymorphism associated with phenotypic sex. This SNP changes an amino acid (His/Asp384) in the kinase domain. While females are homozygous (His/His384), males are heterozygous. Sex in fugu is most likely determined by a combination of the two alleles of Amhr2. Consistent with this model, the medaka hotei mutant carrying a substitution in the kinase domain of Amhr2 causes a female phenotype. The association of the Amhr2 SNP with phenotypic sex is conserved in two other species of Takifugu but not in Tetraodon. The fugu SD locus shows no sign of recombination suppression between X and Y chromosomes. Thus, fugu sex chromosomes represent an unusual example of proto–sex chromosomes. Such undifferentiated X-Y chromosomes may be more common in vertebrates than previously thought. Diverse systems of sex determination have evolved independently in the animal and plant kingdoms. In vertebrates, so far four master sex-determining (SD) genes, Sry, Dmrt1, Dmy, and Dm-W, have been identified. These genes code for transcription factors and are located on only one of the sex chromosomes surrounded by nonrecombining regions. It is hypothesized that these sex chromosomes evolved from a pair of homologous chromosomes that diverged after acquiring the SD gene. We investigated the SD locus in fugu by high-resolution genetic mapping and association mapping. We found that a SNP that changes an amino acid (His/Asp384) in the kinase domain of anti-Müllerian hormone receptor type II (Amhr2) is perfectly associated with phenotypic sex. A combination of the two alleles of the SNP (homozygous females and heterozygous males) is likely to be responsible for sex determination in fugu. While these alleles are conserved in two other species of Takifugu, they are absent in the freshwater pufferfish, Tetraodon. Furthermore, Fugu Amhr2 lies in a region that shows no evidence for recombination suppression between X and Y chromosomes. Thus, fugu sex chromosomes represent an unusual example of a pre-differentiated phase of sex chromosomes in vertebrates.