Transcriptome profiling of developing photoreceptor subtypes reveals candidate genes involved in avian photoreceptor diversification

Abstract

Avian photoreceptors are a diverse class of neurons, comprised of four single cones, the two members of the double cone, and rods. The signaling events and transcriptional regulators driving the differentiation of these diverse photoreceptors are currently unknown. In addition, many distinctive features of photoreceptor subtypes, including spectral tuning, oil droplet size and pigmentation, synaptic targets and spatial patterning, have been well characterized, but the molecular mechanisms underlying these attributes have not been explored. To identify genes specifically expressed in distinct chicken (Gallus gallus) photoreceptor subtypes, we developed fluorescent reporters that label photoreceptor subpopulations, isolated these subpopulations using fluorescence-activated cell sorting and subjected them to next-generation sequencing. By comparing the expression profiles of photoreceptors labeled with rhodopsin, red opsin, green opsin, and violet opsin reporters, we have identified hundreds of differentially expressed genes that may underlie the distinctive features of these photoreceptor subtypes. These genes are involved in a variety of processes, including phototransduction, transcriptional regulation, cell adhesion, maintenance of intra- and extra-cellular structure, and metabolism. Of particular note are a variety of differentially expressed transcription factors, which may drive and maintain photoreceptor diversity, and cell adhesion molecules that may mediate spatial patterning of photoreceptors and act to establish retinal circuitry. These analyses provide a framework for future studies that will dissect the role of these various factors in the differentiation of avian photoreceptor subtypes.

A single‐cell transcriptome atlas of the adult human retina

“…Notably, there are two known receptor isoforms for THRB (TRβ1 and TRβ2) and further research to determine the precise roles of THRB isoforms in subtype specification of human cones would be of great interest. Moreover, the transcription factor TBX2 has been implicated in subtype specification of Sws1 ‐cones in zebrafish and chicken (Alvarez‐Delfin et al , ; Enright et al , ). In support of these studies, our data showed that TBX2 marks the S‐cones in humans, which is also conserved in macaque (Peng et al , ).…”

Otx2 and Oc1 directly regulate the transcriptional program of cone photoreceptor development

“…While cells rarely showed expression driven only by ThrbCRM1, a large number of cells showed expression driven only by Rxrg208. This pattern suggests that these cells might represent early HCs, which may express Rxrg but not Thrb, and/or different subtypes of cones 62 .…”

“…The activity of Neurod1 CRM A showed a strong co-localization with the activity of ThrbCRM1, likely in cones and HCs (Figure 6B). In addition, we electroporated the Sall1 CRM A, as Sall1 was shown to be regulated by thyroid hormone and to be expressed in chick cones 50,62 . Sall1 CRM A was active in cones, seen by the morphology of the GFP-positive cells and IHC for the Rxrg protein (Figure 6C).…”

RNA sequencing analysis of the developing chicken retina

“…Only recently have transcriptome experiments began to be employed as a tool to dissect global changes in gene expression during chick retinal development. A recent study using an elegant reporter system followed by cell capture and RNA-seq analysis, characterized differential gene expression of rod and cone photoreceptor differentiation during chick retinal development 7 . Focusing solely on this one important class of retinal neuron, Enright and colleagues were able to identify hundreds of differentially expressed genes involved in diverse cellular processes contributing to the birth of chicken photoreceptors 7 .…”

“…A recent study using an elegant reporter system followed by cell capture and RNA-seq analysis, characterized differential gene expression of rod and cone photoreceptor differentiation during chick retinal development 7 . Focusing solely on this one important class of retinal neuron, Enright and colleagues were able to identify hundreds of differentially expressed genes involved in diverse cellular processes contributing to the birth of chicken photoreceptors 7 . Currently, there are no publicly available data sets characterizing the whole chicken retina transcriptome during development.…”