Development and function of brain photoreceptors in the annelid Platynereis dumerilii

Abstract

The aim of this thesis was the molecular, developmental and functional characterization of brain photoreceptors in the annelid Platynereis dumerilii. These results shed new light on the evolution of photoreceptors and circadian centers within Bilateria. Ciliary-type photoreceptors expressing c-opsin1 and the circadian marker bmal were previously identified in the dorsal brain of Platynereis larvae (Arendt et al., 2004), indicating that these brain photoreceptors might be involved in the entrainment of the circadian clock. To study their development, I used EdU incorporation analyses, together with time lapse imaging of embryos expressing fluorescent markers of histones and membranes. This showed that the brain circadian center (demarcated by the expression of bmal and cry1) is mostly formed by cells that belong to the same lineage. These cells are generated by asymmetric cell divisions of neural progenitors expressing the transcription factor rx (retinal homeobox), a crucial regulator of vertebrate pineal, retina and hypothalamus development. Next, I established methods to study gene function in the developing Platynereis embryos (morpholino-mediated knockdown, overexpression of mRNA, knockout with zinc finger nucleases). I applied these methods to study the role of Rx in the development of the ciliary photoreceptor lineage. Rx knockdown resulted in the loss of ciliary photoreceptors and in extensive changes in brain development, consistent with a role of Rx in the maintenance and/or multipotency of neural progenitors. The analysis of the molecular identity of these cells showed that they all express a peropsin gene, the melatonin synthesis marker hiomt and several markers of the pineal complex, the structure that in vertebrates releases the hormone melatonin. Among these cells, a new photoreceptor type was identified, which is an asymmetric serotoninergic cell expressing c-ops1 and several pineal markers. Hierarchical clustering of Platynereis and vertebrate cell types revealed that this serotoninergic cell and the “canonical” ciliary photoreceptors are the Platynereis cells more similar to the vertebrate pineal and retinal photoreceptors. To understand the function of these brain hiomt expressing cells, I investigated the expression of melatonin receptors. In the trochophore larva, the only site of ex- pression of melatonin receptors was the prototroch, a belt of ciliated cells used for locomotion. Melatonin can specifically decrease the activity of the prototroch cells. Ciliary locomotion follows a circadian rhythm (with higher speed during daytime, and lower speed at night), which can be reset by phototransduction. This suggests that melatonin is the clock output mediating nighttime decrease of locomotor activity. This study indicates that serotoninergic and melatonin releasing ciliary photoreceptors existed at the base of Bilateria. These photoreceptors have a conserved regulatory signature (rx, tbx2/3, lhx2/9, miR-7) and their ancestral function was the direct control of ciliary locomotion.