P1-110 Thursday, Jan. 4 15:30 - 17:30 Functional investigation of a rhabdomeric opsin gene in Capitella teleta NEAL, SE*; DE JONG, DM; SEAVER, EC; Whitney Lab for Marine Biosciences; Whitney Lab for Marine Biosciences, University of Florida; Whitney Lab for Marine Biosciences, University of Florida email@example.com
There is a rich diversity of photoreceptors in annelids, making them an important group for studying the evolution of light detection. The opsins are a family of genes that encode light sensitive proteins. The annelid, Capitella teleta, possesses 9 distinct opsin genes. Capitella larvae are positively phototactic, although it is not known which opsin genes are responsible for this behavior. From initial expression analysis of three opsin genes, a single rhabdomeric opsin, Opsin-119596, is expressed in the photoreceptor cells of the eyes in Capitella larvae. This study directly investigates the function of Opsin-119596 through knockout with CRISPR-Cas9, a technique that allows for targeted gene-specific knockout. Here, we established CRISPR-Cas9 technology for Capitella, and used three distinct techniques to assess gene knockout and function in larvae. First, PCR amplification and DNA sequencing was used to demonstrate efficient genomic DNA editing of the Opsin-119596 locus. Second, using in situ hybridization, loss of the Opsin-119596 transcript in the eye photoreceptor cells in Capitella was observed, presumably due to non-sense mediated decay. Finally, using a behavioral assay, there was a significant decrease in positive phototaxis relative to unmanipulated controls in Capitella larvae. This behavioral phenotype is similar to that observed when the entire larval eye is deleted. These results demonstrate the feasibility of gene editing using CRISPR-Cas9 in the annelid C. teleta, and establish a direct link between Opsin-119596 and the positive phototactic behavior of Capitella larvae.