Achieving the long-term goal of formulating a comprehensive understanding of how naturally-occurring genetic changes produce diversity will require the comparative study in many taxa of tractable developmental traits that were acquired as a result of inter-species divergence in developmental genetic pathways. Because many genetic pathways regulating developmental traits are broadly conserved, traits prone to change are ideal traits to study. Surprisingly, sex determination pathways can evolve rapidly, despite the fundamental role of these pathways in instructing an organism to develop as a male or female, a choice having numerous developmental consequences. Changes in sex determination pathways between even closely related species can produce striking developmental differences. A conserved sex determination pathway in Caenorhabditis elegans and C. briggsae regulates sexual development, but different genetic alterations of the pathway independently produced the same sexual developmental trait, hermaphroditism, in both species. These genetic modifications cause females to produce male gametes (sperm) prior to producing female gametes (eggs), resulting in individual self-fertility. The observation that self-fertility independently evolved twice in Caenorhabditis, coupled with the vast resources available for experimental developmental and genetic approaches in Caenorhabditis, makes hermaphroditism an ideal model trait with which to conduct a thorough genetic analysis of the evolutionary genetic architecture underlying developmental change. In order to achieve this objective, one experimental approach of the proposed research is to determine whether a gene known to be necessary for hermaphroditism, fog-2, is sufficient to produce self-fertility when moved as a transgene into true females. In addition, whether a second gene, gld-1, known to be sperm- promoting in C. elegans but egg-promoting in C. briggsae, was involved in gamete sex determination prior to the evolution of hermaphroditism will be identified by eliminating gld-1 function in multiple species of the genus Caenorhabditis using RNA interference and visually assessing resulting germline phenotypes. Together, these approaches will reveal whether one or multiple evolutionary changes were required in C. elegans and C. briggsae to produce hermaphroditism. In anticipation that additional genes necessary for hermaphroditism exist in C. briggsae, novel genes controlling gamete sexual fate in C. briggsae will be identified using a mutagenesis screen for self-sterile worms. Describing the suites of genetic changes responsible for the independent acquisition of hermaphroditism in C. elegans and C. briggsae will contribute to the long-term goal of understanding how genetic pathways evolve to produce developmental change. Additionally, identifying novel genetic factors regulating germ cell sexual fate might improve our understanding of causes of human hermaphroditism and infertility.