Abstract
During vertebrate retinal development, subsets of progenitor cells generate progeny in a non-stochastic manner, suggesting that these decisions are tightly regulated. However, the gene-regulatory network components that are functionally important in these progenitor cells are largely unknown. Here we identify a functional role for the OTX2 transcription factor in this process. CRISPR/Cas9 gene editing was used to produce somatic mutations of OTX2 in the chick retina and identified similar phenotypes to those observed in human patients. Single cell RNA sequencing was used to determine the functional consequences OTX2 gene editing on the population of cells derived from OTX2-expressing retinal progenitor cells. This confirmed that OTX2 is required for the generation of photoreceptors, but also for repression of specific retinal fates and alternative gene regulatory networks. These include specific subtypes of retinal ganglion and horizontal cells, suggesting that in this context, OTX2 functions to repress sister cell fate choices.
Highlights
The vertebrate retina is a highly structured tissue comprising six neuronal classes and several types of glia
Elucidation of the mechanisms by which each cell type is generated from multipotent retinal progenitor cells (RPCs) is crucial for the development of successful cell replacement therapies for patients that suffer from retinal degeneration
These data show the strict requirement of Orthodenticle Homeobox 2 (OTX2) for the specification of PR cells, while specific subtypes of retinal ganglion cells (RGCs) and horizontal cells (HCs) are generated in the OTX2 mutant retinas, suggesting that alternate fates are not randomly chosen, but are selected from those normally associated with OTX2-positive RPCs
Summary
The vertebrate retina is a highly structured tissue comprising six neuronal classes and several types of glia. The effects of OTX2 loss on OTX2-positive RPCs were analyzed using a reporter active in this cell population combined with the examination of known markers and characterization using single cell RNA sequencing These data show the strict requirement of OTX2 for the specification of PR cells, while specific subtypes of RGCs and HCs are generated in the OTX2 mutant retinas, suggesting that alternate fates are not randomly chosen, but are selected from those normally associated with OTX2-positive RPCs. In addition, these data show the strict requirement of OTX2 for the specification of PR cells, while specific subtypes of RGCs and HCs are generated in the OTX2 mutant retinas, suggesting that alternate fates are not randomly chosen, but are selected from those normally associated with OTX2-positive RPCs This high-resolution analysis of OTX2 function provides new insights into the process of PR formation in the vertebrate retina and the mechanisms of retinal cell fate choice
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