Expression and function of the transcription factor Rx in Nematostella vectensis

Abstract

The development of the central nervous system is highly studied and a comparatively well understood field of neurogenesis. The fundamental aspects of neurogenesis have however mostly been studied in a select few bilaterian model organisms, leaving many questions regarding the evolution of this process open. The cnidarian and bilaterian lineages are sister groups that separated approximately 600 million years ago. Cnidarians occupy an informative phylogenetic position to study the early evolution of cellular and molecular aspects of neurogenesis, and to understand common principles of neural development. The Cnidaria sea anemone Nematostella vectensis has in recent years emerged as one of the most tractable cnidarian models for the study of neurogenesis and development in an evolutionary context. The later steps of neural differentiation in Nematostella are yet to be fully understood. Neuronal markers have been found for different neural sub-populations, one of which is the transcription factor NvFoxQ2d that gives rise to sensory cells in Nematostella. Although the function of NvFoxQ2d is yet to be determined, studies are being performed to further understand this transcription factor, one of which is a transcriptome of the fluorescent cells in the NvFoxQ2d::mOrange transgenic reporter line. During the analysis of this dataset, the transcription factor NvRx was found to be highly upregulated in this cell population. While the Retinal homeobox gene (Rx) has a well-described role in the formation of the retina, and the formation and proliferation of retinal progenitor cells in vertebrates, the function of NvRx in the eyeless Nematostella is in contrast yet to be determined. By performing a double fluorescence in-situ hybridization, we show that NvRx is expressed in the majority of the NvFoxQ2d cell population. We hypothesised that NvRx could play a role in the regulation of these NvFoxQ2d-expressing cells, and by performing an in-situ on embryos injected with NvRx shRNA injection, we show that NvRx acts as an upstream regulator of NvFoxQ2d. We also found NvRx-expressing cells in close proximity to NvElav1-expressing neural cells, indicating that these cells could be descended from the same progenitor cells. Finally, a CRISPR/Cas9 mediated mutant line was successfully generated to further study the function of NvRx in Nematostella development