Genomic basis of sex change in fish

Abstract

Sexual dimorphism is an ancient feature of most sexually reproducing species: males and females differ not just in their gonadal structure and functions, but in many aspects of their morphology, physiology, and behavior. In most multi- cellular organisms this fundamental dichotomy is established during early embryonic development; individuals become either female or male and maintain these roles for their entire life. However, in some plant and animal species, notably fishes, individuals begin life as one sex, but change sometime later to the other, a process termed sequential hermaphroditism. Sex change in sequential hermaphrodites entails radical restructuring of the gonads, alteration in morphology, and modifications to behavior. The molecular basis of this stunning transformation is poorly understood, but is of considerable interest, as knowledge of this process would greatly enhance our understanding of sex determination, cellular differentiation and developmental commitment.

The aim of this thesis is to investigate how sex change is regulated in fish at the molecular level. Teleost fishes display remarkably diverse and plastic sexual development patterns. Functional sex change is widespread in marine fishes, spanning at least 27 families, and including many species of commercial importance. The bluehead wrasse (Thalassoma bifasciatum), a small Caribbean reef fish, exhibits female-to-male (protogynous) sex change based on social context. Removing the dominant male from a social group can induce sex change in the largest females. The speed of this process is astonishing: the behavioral changes can begin within minutes, followed by gonadal sex change that occurs in 8 to 10 days, while changes in the external morphology can take up to 21 days. Significant progress has been made in understanding the ecology and the neuroendocrine bases of sex change in this species, but detailed mechanisms still remain elusive, especially at the molecular level.

This thesis explores the genetic basis of sex change using the bluehead wrasse as a model. RNA-sequencing technology was used to capture the transcriptomic profiles in the brain and gonadal tissues of bluehead wrasses at a series of time points across the entire process of protogynous sex change. To identify the primary trigger and subsequent genetic cascade that re-engineer a female into a male, the transcriptional changes in the brain and gonad during sex change were examined using advanced bioinformatic tools. A de novo transcriptome assembly was generated for read mapping and subsequent gene expression analysis. Functional annotation and enrichment analysis were carried out for the differentially expressed genes to detect global differences in genetic pathways. A large number of differentially expressed transcripts (n=59590) were identified in the gonad of bluehead wrasses, but a much smaller set (n=313) was identified in the brain. Several potential triggers of gonadal sex change and a few genetic pathways, such as the Jak-STAT signaling pathway, that likely contribute to the transformation of ovary to testis were detected in the gonadal transcriptomes of bluehead wrasses. Surprisingly, genes implicated in female sexual development, e.g. foxl2, rspo1, and wnt4b, showed less conserved expressional patterns in the gonads of bluehead wrasses when compared to their well established, and highly constrained, roles in mammalian models. These findings suggest that bluehead wrasses predominantly use an evolutionarily conserved genetic toolkit, but that subtle variation in the standard sex-determination regulatory network may contribute to the sexual plasticity characteristic of these fish. In addition, although transcriptional changes in the brain were much less pronounced than those detected in the gonads, a few promising candidates, including genes associated with biosynthesis of neurosteroids and isotocin, were suggested as important regulatory factors in the brain during sex change. Finally, based on these novel data, a hypothetical mechanism by which gonadal sex change may be initiated and processed in protogynous fish was proposed. Overall, this thesis provides the first genomic data for a sex changing fish, and a framework for better understanding the molecular basis of phenotypic plasticity and tissue re-engineering in response to environmental influences. In a general sense, it also sheds some light on the evolution of diverse sex determination and differentiation systems in vertebrates.