Molecular genetic characterization of NR4A nuclear receptor activities regulating organogenesis in vivo

NR4A nuclear receptors (NRs) are a conserved, functionally diverse group of transcription factors that regulate many metabolic and developmental processes including organogenesis. Determining the specific functions of NR4A NRs during proliferation and differentiation is crucial to the understanding of the regulatory networks involved in these processes. We are studying the functions of the NR4A nuclear receptor, nhr-6, in the genetic model organism C. elegans. Previous studies have determined that NHR-6 is required for both cell proliferation and differentiation during development of the spermatheca organ. NHR-6 localizes to the nucleus of spermathecal cells during proliferation and differentiation of these cells and has been shown to regulate these functions through a DNA binding mechanism. I utilized microscopy and genetic techniques to uncover a specific role for NHR-6 during G 1 /S transition of the cell cycle. I also uncovered that this cell cycle progression role is independent of its function during differentiation, further suggesting a dualistic role for NHR-6 during development.;Additionally, I utilized the technique of chromatin immunoprecipitation followed by high throughput DNA sequencing (ChIP-seq) to uncover NHR-6 target genes. ChIP-seq results revealed 3,222 unique sites that were enriched for NHR-6 binding during the proliferation and differentiation stages of spermatheca development. I incorporated a functional bias to select high priority NHR-6 target genes that were validated by Real-time quantitative (RT-qPCR) and used in an RNAi screen to analyze spermatheca phenotypes. These results have shown a specific function for NHR-6 during proliferation as well as revealed a number of candidate NHR-6 target genes that display functions in spermatheca development. This data reveals an extensive transcription factor regulatory network that will help elucidate the precise function of NHR-6 during spermatheca organogenesis.