Epigenomic profiling of retinal progenitors reveals LHX2 is required for developmental regulation of open chromatin

Zibetti, Cristina; Liu, Sheng; Wan, Jun; Qian, Jiang; Blackshaw, Seth

doi:10.1038/s42003-019-0375-9

Cited by 39 publications

(87 citation statements)

References 46 publications

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“…Several genes have been attributed to neuroretinal specification as well as the proliferative and multipotent ability of retinal progenitor cells, including Vsx2 (also known as Chx10), Pax6, Lhx2, Rax, Six3, and Six6 [71][72][73][74][75][76][77][78][79]. Many of the genes are also implicated in retinal abnormalities; for instance, Pax6 mutations can lead to foveal hypoplasia, while Rax mutations can cause microphthalmia leading to retinal dysplasia [80,81].…”

Section: The Genetics Of Retinal Developmentmentioning

confidence: 99%

Retinogenesis of the Human Fetal Retina: An Apical Polarity Perspective

Quinn

Wijnholds

2019

Genes

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The Crumbs complex has prominent roles in the control of apical cell polarity, in the coupling of cell density sensing to downstream cell signaling pathways, and in regulating junctional structures and cell adhesion. The Crumbs complex acts as a conductor orchestrating multiple downstream signaling pathways in epithelial and neuronal tissue development. These pathways lead to the regulation of cell size, cell fate, cell self-renewal, proliferation, differentiation, migration, mitosis, and apoptosis. In retinogenesis, these are all pivotal processes with important roles for the Crumbs complex to maintain proper spatiotemporal cell processes. Loss of Crumbs function in the retina results in loss of the stratified appearance resulting in retinal degeneration and loss of visual function. In this review, we begin by discussing the physiology of vision. We continue by outlining the processes of retinogenesis and how well this is recapitulated between the human fetal retina and human embryonic stem cell (ESC) or induced pluripotent stem cell (iPSC)-derived retinal organoids. Additionally, we discuss the functionality of in utero and preterm human fetal retina and the current level of functionality as detected in human stem cell-derived organoids. We discuss the roles of apical-basal cell polarity in retinogenesis with a focus on Leber congenital amaurosis which leads to blindness shortly after birth. Finally, we discuss Crumbs homolog (CRB)-based gene augmentation.

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Section: The Genetics Of Retinal Developmentmentioning

confidence: 99%

Retinogenesis of the Human Fetal Retina: An Apical Polarity Perspective

Quinn

Wijnholds

2019

Genes

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“…ATAC-Seq DNA was amplified, and the number of PCR cycles were calculated by following previously described protocol (63). PCR products (10UL) were run on a 1.5% agarose gel for expected DNA pattern.…”

Section: Atac-seqmentioning

confidence: 99%

Cross-species transcriptomic and epigenomic analysis reveals key regulators of injury response and neuronal regeneration in vertebrate retinas

Hoang

Wang

Boyd

et al. 2019

Preprint

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sentence: This study identifies gene regulatory networks controlling proliferative and neurogenic competence in retinal Müller glia. AbstractInjury induces retinal Müller glia of cold-blooded, but not mammalian, vertebrates to regenerate neurons. To identify gene regulatory networks that control neuronal reprogramming in retinal glia, we comprehensively profiled injury-dependent changes in gene expression and chromatin accessibility in Müller glia from zebrafish, chick and mice using bulk RNA-Seq and ATAC-Seq, as well as single-cell RNA-Seq. Crossspecies integrative analysis of these data, together with functional validation, identified evolutionarily conserved and species-specific gene networks controlling glial quiescence, gliosis and neurogenesis. In zebrafish and chick, transition from the resting state to gliosis is essential for initiation of retinal regeneration, while in mice a dedicated network suppresses neurogenic competence and restores quiescence. Selective disruption of NFI family transcription factors, which maintain and restore quiescence, enables Müller glia to proliferate and generate neurons in adult mice following retinal injury. These findings may aid in the design of cell-based therapies aimed at restoring retinal neurons lost to degenerative disease. previously shown to induce MG reprogramming independent of injury (20). In the mouse, we also tested FGF2 and insulin treatment, which induces limited MG proliferation, but not neurogenic competence (21).For bulk RNA-Seq and ATAC-Seq, in zebrafish, we used the reporter line Tg[gfap:EGFP]nt11 (22) and cell sorting to enrich MG cells, purifying 9.8% of total input retinal cells (Fig. S1A). In mouse, we performed intraperitoneal injection of tamoxifen at postnatal day (P) 21 to induce MG-specific Sun1-GFP expression in GlastCreERT2;CAG-lsl-Sun1-GFP mice (23), purifying 3.5% of total input cells ( Fig. S1A). Based on RT-qPCR, we had an overall enrichment of 30-fold of canonical MG marker genes such as rlbp1a/Rlbp1 and Glul in the GFP-positive (GFP+) fraction in both species (Fig. S1B). Bulk RNA-Seq samples were generated from both the GFP+ MG and GFP-neuronal fractions in zebrafish and mouse, while ATAC-Seq samples were generated from the GFP+ cells. Each sample had a minimum of two biological replicates. In both species, we analyzed retinas with a time course following either inner retinal injury induced by NMDA excitotoxicity or outer retinal injury induced by light damage. In total, we generated 100 bulk RNA-Seq libraries and 40 bulk ATAC-Seq libraries ( Fig. 1A, Table S1, Supplementary Data 1).In parallel, we conducted scRNA-Seq analysis of whole retinas at the same time points as used for the bulk RNA-Seq. We profiled retinas following either NMDA or light damage in zebrafish and mouse, as well as following NMDA damage in P10 chick. To distinguish injury-responsive genes from injury-independent reprogramming genes, we profiled zebrafish retinas at multiple time points following T+R treatment, and a single time point in chick and mouse retinas...

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“…In addition, NFIa/b/x are expressed in retinal progenitors and MG, and conditional knockouts of these transcription factors in the developing retina are associated with defects in gliogenesis and the production of bipolar neurons (Clark et al 2019). As progenitors develop, there is an acquired association of NFI motifs with Lhx2 binding sites, which may be related to guiding the neurogenic potency of late retinal progenitors (Zibetti et al 2019).…”

Section: Discussionmentioning

confidence: 99%

Cis-Regulatory Accessibility Directs Muller Glial Development and Regenerative Capacity

VandenBosch

Wohl

Wilken

et al. 2019

Preprint

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19Diseases and damage to the retina lead to losses in retinal neurons and eventual visual 20 impairment. Although the mammalian retina has no inherent regenerative capabilities, fish have 21 robust regeneration from Müller glia (MG). Recently, we have shown that driving expression of 22 Ascl1 in adult mouse MG stimulates neurogenesis similar to fish regeneration. The regeneration 23 observed in the mouse is limited in the variety of neurons that can be derived from MG; Ascl1-24 expressing MG primarily generate bipolar cells. To better understand the limits of MG-based 25 regeneration in mouse retinas, we used ATAC-and RNA-seq to compare newborn progenitors 26 with MG. Our analysis demonstrated striking similarities between MG and progenitors, with 27 losses in regulatory motifs for neurogenesis genes. Young MG were found to have intermediate 28 expression profiles and accessible DNA, which is mirrored in the ability of Ascl1 to direct 29 bipolar neurogenesis in young MG. When comparing what makes bipolar and photoreceptor 30 cells distinct from glial cells, we find that bipolar-specific accessible regions are more frequently 31 linked to bHLH motifs and Ascl1 binding, indicating that Ascl1 preferentially binds to bipolar 32 regions. Overall, our analysis indicates a loss of neurogenic gene expression and motif 33 accessibility during glial maturation that may prevent efficient reprogramming. 34 35 36 Neuron loss caused by disease and damage to the mammalian retina can lead to 37 permanent vision loss. While some species are readily capable of regenerating lost neurons, 38 mammalian retinas are not regenerative. In the mammalian retina, neuron loss caused by direct 39 damage to the retina leads to reactive gliosis of the Müller glia (MG), similar to that of astrocytes 40 in the brain (Bringmann et al. 2009).41 Teleost fish, by contrast, are capable of regenerating retinal neurons, including 42 photoreceptors and ganglion cells, after damage. This regeneration is carried out by the MG, 43 which respond to damage by generating progenitor-like cells, similar to those in the developing 44 retina (Goldman 2014; Gemberling et al. 2013). This regeneration is accompanied by waves of 45 gene expression and morphological changes to the MG, regulated by epigenomic changes 46 directing regeneration (for review see VandenBosch and Reh 2019). The murine retina also 47 undergoes epigenomic changes after damage, but neurogenic programs are not re-expressed, and 48 neuronal regeneration does not occur (VandenBosch and Reh 2019). 49A critical difference between the fish MG and the mammalian MG in their response to 50 damage is in their expression of the proneural transcription factor Ascl1. In fish, Ascl1 is quickly 51 upregulated after damage, and is necessary for regeneration of new neurons (Ramachandran et 52 al. 2010; Fausett et al. 2008). In the murine retina, Ascl1 is expressed in retinal progenitors and 53 necessary for development of rods and bipolar cells (Ohsawa and Kageyama 2008); however it is 54 not expressed in...

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Epigenomic profiling of retinal progenitors reveals LHX2 is required for developmental regulation of open chromatin

Cited by 39 publications

References 46 publications

Retinogenesis of the Human Fetal Retina: An Apical Polarity Perspective

Retinogenesis of the Human Fetal Retina: An Apical Polarity Perspective

Cross-species transcriptomic and epigenomic analysis reveals key regulators of injury response and neuronal regeneration in vertebrate retinas

Cis-Regulatory Accessibility Directs Muller Glial Development and Regenerative Capacity

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