Molecular Genetic Mechanisms of Gonad Development in Zebrafish

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The long-term objective of this project is to understand the genetic mechanisms that control gonad development in animals with backbones. The general morphology and endocrinology of gonad development are well conserved among vertebrates, but the genetic pathway that determines whether a gonad becomes an ovary or a testis is not well understood in any vertebrate. Understanding the mechanisms that regulate gonad growth and differentiation takes on increasing significance as environmental pollutants, such as some polychlorinated biphenyls (PCBs), may disturb gonad development and endocrine function in fish, birds, and mammals, including humans. The resulting effects on the physiology of reproduction are significant for the preservation of wildlife populations and for human reproductive health. Although the underlying pathway of gonad development may be shared among vertebrates, different vertebrate lineages have different ways of diverting the pathway towards an ovary or a testis. Some species have strong genotypic sex determination, such as mammals in which a specific gene called SRY on the Y-chromosome specifies gonad development. Other species have strong environmental sex determination, such as some turtles in which the temperature of embryo incubation regulates gonad sex. Investigations on zebrafish suggest the hypothesis that multiple genotypic factors, with a minor influence from the environment, cause an individual's gonads to express either genes for androgen synthesizing enzymes and androgen receptor, or express genes for estrogen synthesizing enzymes and estrogen receptor; these endocrine factors may then control the sex of the gonad, primary germ cells, and secondary sex characteristics. This hypothesis will be tested in three ways: Specific Aim 1. The proposed experiments will investigate the expression patterns of genes predicted by the model to play a role in early gonadal development of the zebrafish. Specific Aim 2: The proposed experiments will perturb gonad development using sex steroids and their inhibitors, and various mutations, and investigate whether gene expression patterns change in ways predicted by the model. Specific Aim 3: The proposed experiments will identify quantitative trait loci (QTLs) associated with sex differentiation in zebrafish. Map locations will be compared to those of genes isolated in Specific Aim 1 and with mammalian genomes to suggest candidate genes for gonad differentiation in zebrafish. Achieving the specific aims will result in a more robust understanding of the control mechanisms of gonad development in zebrafish, and will point to genetic loci that influence sex determination in this species. The resulting firmer understanding of the basic nature of sex determination in zebrafish will permit the future design of mutagenesis protocols to isolate novel genes important for vertebrate sex determination.