A Stage-Specific OTX2 Regulatory Network and Maturation-Associated Gene Programs Are Inherent Barriers to RPE Neural Competency

Abstract

The retinal pigment epithelium (RPE) exhibits a diverse range of plasticity across vertebrates and is a potential source of cells for the regeneration of retinal neurons. Embryonic amniotes possess a transitory ability to regenerate neural retina through the reprogramming of RPE cells in an FGF-dependent manner. Chicken RPE can regenerate neural retina at embryonic day 4 (E4), but RPE neural competence is lost by embryonic day 5 (E5). To identify mechanisms that underlie loss of regenerative competence, we performed RNA and ATAC sequencing using E4 and E5 chicken RPE, as well as at both stages following retinectomy and FGF2 treatment. We find that genes associated with neural retina fate remain FGF2-inducible in the non-regenerative E5 RPE. Coinciding with fate restriction, RPE cells stably exit the cell cycle and dampen the expression of cell cycle progression genes normally expressed during regeneration, including E2F1. E5 RPE exhibits progressive activation of gene pathways associated with mature function independently of retinectomy or FGF2 treatment, including retinal metabolism, pigmentation synthesis, and ion transport. Moreover, the E5 RPE fails to efficiently repress OTX2 expression in response to FGF2. Predicted OTX2 binding motifs undergo robust accessibility increases in E5 RPE, many of which coincide with putative regulatory elements for genes known to facilitate RPE differentiation and maturation. Together, these results uncover widespread alterations in gene regulation that culminate in the loss of RPE neural competence and implicate OTX2 as a key determinant in solidifying the RPE fate. These results yield valuable insight to the basis of RPE lineage restriction during early development and will be of importance in understanding the varying capacities for RPE-derived retinal regeneration observed among vertebrates.

Distinct Metabolic States Direct Retinal Pigment Epithelium Cell Fate Decisions

“…The cellular reprogramming of RPE into neural retina is induced by FGF2 in chicken embryos, (Tangeman et al, 2022; Luz-Madrigal et al, 2014; Spence et al, 2004). However, the limited period of time and accessibility to the in vivo system has complicated the elucidation of the metabolic determinants of RPE reprogramming.…”

“…However, the limited period of time and accessibility to the in vivo system has complicated the elucidation of the metabolic determinants of RPE reprogramming. RPE can be cultured as an explant and reprogrammed into neural retina in the presence of FGF2 (Sakami et al, 2008; Tangeman et al, 2022), but the reprogramming process has not been studied in detail ex vivo. To better understand the RPE explant model, we first performed a phenotypic characterization.…”

“…S1 A), suggesting that FGF2 is a trigger of cell reprogramming, but that its sustained presence is not necessary for neuroepithelium formation. Similar to in vivo, RPE explants can reprogram when collected at E4, but lose neurocompetence at embryonic day 5 (E5) (Tangeman et al, 2022; Sakami et al, 2008). In addition, E4 RPE explants lose neurocompetence by 48h of culture, as RPE retains an ability to continue differentiating ex vivo (Sakami et al, 2008).…”

“…Despite these findings, the metabolic foundations of embryonic RPE reprogramming remain relatively unexplored. Our recent study highlighted the activation of retinol metabolism concurrent with RPE fate restriction, suggesting that the metabolic pathways associated with RPE maturation may limit reprogramming competence (Tangeman et al, 2022). However, the metabolic requirements of RPE cells as they dedifferentiate and change identity during reprogramming are less clear.…”

Effects of Lycium barbarum L. Polysaccharides on Vascular Retinopathy: An Insight Review

“…Although the mechanism of apoptosis in the pathological process of AMD has not been elucidated, the abnormal accumulation of all-trans retina is associated with caspase-3/ Gasdermin E-mediated scortosis [ 79 ]. Apoptosis begins with activation of the inflammasome, including NLRP3 and Procaspase-1 (precursors), and so on.…”