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Dan P Murphy, Andrew Hughes, Joseph C Corbo; Cis-regulatory basis of sister cell type divergence in the vertebrate retina. Invest. Ophthalmol. Vis. Sci. 2019;60(9):1001.
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© ARVO (1962-2015); The Authors (2016-present)
Photoreceptor and bipolar cells are considered ‘sister’ cell types, sharing developmental, morphological and molecular features inherited from a common ancestral cell type. While research into the cis-regulatory architecture driving gene expression in photoreceptor cells is ongoing, the cis-regulatory landscape of bipolar cells, and how it compares to that of photoreceptors, remains unknown. The goal of this study is to profile the transcriptome and chromatin accessibility of ON and OFF bipolar cells and compare these data to similar data from rod and cone photoreceptors in order to identify shared and divergent cis-regulatory mechanisms.
ON and OFF bipolar cells were isolated from the adult mouse retina by Fluorescence Activated Cell Sorting (FACS), using transgenic mouse lines expressing fluorescent marker proteins. Purified populations were subject to RNA-seq to profile the transcriptome and ATAC-seq (Assay for Transposase-Accessible Chromatin by sequencing) to map open chromatin. The role of homeodomain (HD) motifs in driving activity in each cell type was analyzed by subretinal injection and electroporation of fluorescent reporters into neonatal mouse retina.
We found that photoreceptors and bipolar cells have highly divergent transcriptional profiles, and yet share remarkably similar cis-regulatory grammars. The predominant cis-regulatory motifs in the enhancers of both cell types are K50 homeodomain binding sites, which are required for activity. These motifs are bound by two transcription factors (TFs), CRX and OTX2, that are co-expressed in both photoreceptors and bipolar cells. In contrast, photoreceptor-specific open chromatin regions have an abundance of Q50 homeodomain motifs which are lacking from bipolar-specific open chromatin. We show that these Q50 motifs, which are bound by the bipolar-specific TF VSX2, mediate repression of photoreceptor-specific genes in bipolar cells.
The evolutionary appearance of Q50 motifs in the enhancers of photoreceptor genes, perhaps via mutation of pre-existing K50 motifs, was a critical step in the divergence of the bipolar transcriptome from that of photoreceptors. In this way, subtle changes in cis-regulatory grammar paved the way for the emergence of the interneuronal circuitry found in modern vertebrate retinas.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.
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