Development of the blue cone bipolar cell in the mouse retina

National Eye Institute

Investigators

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Abstract

In order to compare the transcriptome of S- and M-cones, we need to isolate true S-cones. An existing transgenic mouse line that expresses EGFP driven by an S-opsin promoter was deemed unreliable, because some S-opsin negative cones also express GFP. This is likely due to the short S-opsin promoter region used in generating the mouse line. Due to the low percentage of true S-cones, this contamination from M-cones precludes successful RNAseq. In the past year, we have generated a mouse line using Bacterial Artificial Chromosome (BAC) recombination to include longer upstream and downstream regulatory sequences at the endogenous S-opsin gene locus. We successfully screened several mouse lines that express yellow fluorescent protein YFP (Venus) only in S-opsin expressing cones. We have used fluorescent sorting methods to acquire true S-cones from the dorsal retina and to prepare samples for gene profiling experiments. Part of this data has been used in a collaborative project with Anand Swaroops lab, who studies the development and evolutionary origin of rods. We will compare RNAseq data from S-cones and M-cones and identify candidate genes for their synaptic targeting. In parallel, we are developing a single cell based technique to manually pick individual photoreceptors from dissociated retinal tissues. Additionally, we have successfully labeled live S-cones in the ground squirrel retina and have been able to pick them individually. This will allow us to genetically profile S-cones and M-cones from the ground squirrel retina, which has complete separate S- and M-cone systems.