Molecular Determinants of Photoreceptor Presynaptic Terminal Morphology
Date
2018-03-13
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Abstract
Rod and cone photoreceptors are light-receptive cells in the visual system that convert photons into an electrochemical signal to be processed through the retina and transmitted into the brain. From the first visual synapse, from photoreceptor to interneurons, rod spherules and cone pedicles diverge in morphology and connectivity patterns. It is known that the transcription factor Nrl is sufficient to drive the cone-to-rod cell fate conversion and morphological change. To dissect the source of the spherule versus pedicle differences, we performed a directed RNAi screen using in vivo electroporation to knock down a select portion of the Nrl regulome to identify genes associated with morphological features.
We systematically characterized four distinct features of rod spherules and S-cone pedicles: spherule width, terminal position in the outer plexiform layer, ribbon number, and presence or absence of telodendrites. Using previously published next-generation sequencing data of the transcriptome of developing rod and cone-like photoreceptors as well as key transcription factor binding profiles, we defined a set of genes potentially associated with restricting spherule morphology from that of the default pedicle state. By knocking down genes individually, we were able to dissect the effects each gene has to restrict spherules. Our screen identified twenty-seven genes that control one or two independent features of rod photoreceptor spherule morphology, terminal width or outer plexiform layer position. Many of these were confirmed either through rescue experiments or examination of loss of function of mouse strains.
Lastly, we generated a protein interaction network to connect the seemingly random sets of genes that controlled spherule morphology. Clustering of genes in this network did not show enrichment of our positive screen targets into communities. When we created shortest network pathways between all pairs of positive targets, we discovered that there was an enrichment of pathways that utilized Ncoa2, and this gene has a direct path to Nrl. We hypothesize that we have discovered a more directly involved gene regulatory network associated with the restriction of rod photoreceptor spherules. This knowledge should help in blinding disease treatment strategies to improve proper integration into the native retinal circuitry after loss of photoreceptors.
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rod, spherule, retina, network, synapse