Epigenetic Regulation of Neural Progenitor Multipotency

Abstract

Time plays a key role in the histogenesis of all tissues. Still, developmental time is doubly important in the nervous system, where the large diversity of neuronal and glial cell types are produced by neural progenitor cells that undergo step-wise competence transitions in a time-dependent manner. This is particularly evident in the developing retina, where resident neural progenitors generate complex lineages comprising diverse neuronal and glial cell types in stereotyped sequences by altering their multipotency over developmental time. However, a major question that persists is how individual multipotent progenitors dynamically regulate their developmental potential to produce specific cell types at the right time and in the correct proportions and sequence. Landmark studies have suggested that competence transitions are cell-autonomously encoded by transcriptional regulators and epigenetic processes, but these mechanisms are not well understood in vertebrate lineages. Since nucleosome remodellers interact with both transcription factors and heterochromatic complexes, we sought to address the role of nucleosome remodeling complexes in developmental timing in the mouse retina. We generated conditional knockouts (cKOs) of Chd4 – a key nucleosome remodelling enzyme in neural progenitors. Chd4 cKOs exhibited a marked expansion in early-born retinal ganglion cells. Postnatally, later-born rod photoreceptors were drastically underproduced. This was partly due to progenitors failing to differentiate on schedule and continuing to proliferate beyond their normal developmental window. This ultimately led to a striking increase in Müller glia production. Histological marker analyses suggest that these effects were independent of alterations in cell death or proliferation at perinatal stages; however, as development progressed, Chd4 cKO retinas exhibited elevated apoptosis, which might have additionally contributed to the decreased rod generation. To determine whether Chd4 regulates retinal cell-type production by altering competence windows, we performed EdU birthdating. These experiments revealed that cell fates were altered without affecting the early RPC competence window. Next, we examined the effect of Chd4 on the genome and transcriptome, focusing on the perinatal retinal progenitor pool. Multi-seq single-cell transcriptomics demonstrated that deletion of Chd4 created divergent gene expression profiles and developmental trajectories. ATAC-seq experiments performed on sorted P1 retinal progenitors revealed that chromatin accessibility was significantly increased at ~10,000 genomic loci and ~4,000 genes in the Chd4 cKO. The changes in accessibility in Chd4 cKO RPCs correlated with increases in transcription, suggesting that Chd4 restricts the genome to repress progenitor identity and promote differentiation. Thus, despite a very strong shift in the production of early-born and late-born cell types, our data suggest that Chd4-dependent nucleosome remodelling plays a crucial role in the temporal transition that governs lineage termination but does not regulate earlier temporal transitions.