Chromatin accessibility dynamics and single cell RNA-Seq reveal new regulators of regeneration in neural progenitors

Kakebeen, Anneke Dixie; Chitsazan, Alexander Daniel; Williams, Madison C; Saunders, Lauren; Wills, Andrea E.

doi:10.7554/elife.52648

Cited by 51 publications

(63 citation statements)

References 87 publications

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“…Contrary to the expected, recent high-throughput analysis of the response of pax6-expressing neural progenitor cells to tail amputation showed that before proliferation a set of neural progenitors is directly differentiated into neurons through the activation of a neurogenic program during the first 24 hpa. To replenish the regenerated spinal cord of neural progenitors its proliferation is only triggered at 48–72 hpa [ 64 ]. This immediate differentiation, without a previous proliferative step, probably allows the formation of new neurons very rapidly supporting a successful regenerative process.…”

Section: Discussionmentioning

confidence: 99%

Cellular response to spinal cord injury in regenerative and non-regenerative stages in Xenopus laevis

et al. 2021

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Background The efficient regenerative abilities at larvae stages followed by a non-regenerative response after metamorphosis in froglets makes Xenopus an ideal model organism to understand the cellular responses leading to spinal cord regeneration. Methods We compared the cellular response to spinal cord injury between the regenerative and non-regenerative stages of Xenopus laevis. For this analysis, we used electron microscopy, immunofluorescence and histological staining of the extracellular matrix. We generated two transgenic lines: i) the reporter line with the zebrafish GFAP regulatory regions driving the expression of EGFP, and ii) a cell specific inducible ablation line with the same GFAP regulatory regions. In addition, we used FACS to isolate EGFP+ cells for RNAseq analysis. Results In regenerative stage animals, spinal cord regeneration triggers a rapid sealing of the injured stumps, followed by proliferation of cells lining the central canal, and formation of rosette-like structures in the ablation gap. In addition, the central canal is filled by cells with similar morphology to the cells lining the central canal, neurons, axons, and even synaptic structures. Regeneration is almost completed after 20 days post injury. In non-regenerative stage animals, mostly damaged tissue was observed, without clear closure of the stumps. The ablation gap was filled with fibroblast-like cells, and deposition of extracellular matrix components. No reconstruction of the spinal cord was observed even after 40 days post injury. Cellular markers analysis confirmed these histological differences, a transient increase of vimentin, fibronectin and collagen was detected in regenerative stages, contrary to a sustained accumulation of most of these markers, including chondroitin sulfate proteoglycans in the NR-stage. The zebrafish GFAP transgenic line was validated, and we have demonstrated that is a very reliable and new tool to study the role of neural stem progenitor cells (NSPCs). RNASeq of GFAP::EGFP cells has allowed us to clearly demonstrate that indeed these cells are NSPCs. On the contrary, the GFAP::EGFP transgene is mainly expressed in astrocytes in non-regenerative stages. During regenerative stages, spinal cord injury activates proliferation of NSPCs, and we found that are mainly differentiated into neurons and glial cells. Specific ablation of these cells abolished proper regeneration, confirming that NSPCs cells are necessary for functional regeneration of the spinal cord. Conclusions The cellular response to spinal cord injury in regenerative and non-regenerative stages is profoundly different between both stages. A key hallmark of the regenerative response is the activation of NSPCs, which massively proliferate, and are differentiated into neurons to reconstruct the spinal cord. Also very notably, no glial scar formation is observed in regenerative stages, but a transient, glial scar-like structure is formed in non-regenerative stage animals.

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Section: Discussionmentioning

confidence: 99%

Cellular response to spinal cord injury in regenerative and non-regenerative stages in Xenopus laevis

et al. 2021

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“…Instead of interpreting a snapshot of a cell's gene expression, a chromatin signature can provide information about the cell's prospective state and reveal important insights into the gene regulatory networks at play. Chromatin signatures have already been determined for many types of stem cells [81][82][83][84], and ATACseq is broadly applicable across systems [85].…”

Section: Caveatsmentioning

confidence: 99%

One stem cell program to rule them all?

Wiggans

Pearson

2020

The FEBS Journal

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Many species of animals have stem cells that they maintain throughout their lives, which suggests that stem cells are an ancestral feature of all animals. From this, we take the viewpoint that cells with the biological properties of 'stemness'-self-renewal and multipotency-may share ancestral genetic circuitry. However, in practice is it very difficult to identify and compare stemness gene signatures across diverse animals and large evolutionary distances? First, it is critical to experimentally demonstrate self-renewal and potency. Second, genomic methods must be used to determine specific gene expression in stem cell types compared with non-stem cell types to determine stem cell gene enrichment. Third, gene homology must be mapped between diverse animals across large evolutionary distances. Finally, conserved genes that fulfill these criteria must be tested for role in stem cell function. It is our viewpoint that by comparing stem cell-specific gene signatures across evolution, ancestral programs of stemness can be uncovered, and ultimately, the dysregulation of stemness programs drives the state of cancer stem cells.

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“…With the exception of an mRNA-seq study in lamprey, which demonstrated that the response of brain and spinal cord to SCI indeed differ [ 48 ], previous genome-wide expression studies on spinal cord regeneration have focused on either the spinal cord itself, with the lesion at the epicenter ( Xenopus laevis tadpole and frog [ 70 ], turtle [ 142 ], zebrafish [ 50 ]), or on a regenerating tail (e.g., salamander [ 87 ], Xenopus tropicalis tadpole [ 21 , 55 , 80 , 102 ]). Such regeneration involves not only axon regeneration, but also considerable wound repair and tissue restoration.…”

Section: Introductionmentioning

confidence: 99%

Comparative gene expression profiling between optic nerve and spinal cord injury in Xenopus laevis reveals a core set of genes inherent in successful regeneration of vertebrate central nervous system axons

et al. 2020

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Background: The South African claw-toed frog, Xenopus laevis, is uniquely suited for studying differences between regenerative and non-regenerative responses to CNS injury within the same organism, because some CNS neurons (e.g., retinal ganglion cells after optic nerve crush (ONC)) regenerate axons throughout life, whereas others (e.g., hindbrain neurons after spinal cord injury (SCI)) lose this capacity as tadpoles metamorphose into frogs. Tissues from these CNS regions (frog ONC eye, tadpole SCI hindbrain, frog SCI hindbrain) were used in a three-way RNAseq study of axotomized CNS axons to identify potential core gene expression programs for successful CNS axon regeneration. Results: Despite tissue-specific changes in expression dominating the injury responses of each tissue, injuryinduced changes in gene expression were nonetheless shared between the two axon-regenerative CNS regions that were not shared with the non-regenerative region. These included similar temporal patterns of gene expression and over 300 injury-responsive genes. Many of these genes and their associated cellular functions had previously been associated with injury responses of multiple tissues, both neural and non-neural, from different species, thereby demonstrating deep phylogenetically conserved commonalities between successful CNS axon regeneration and tissue regeneration in general. Further analyses implicated the KEGG adipocytokine signaling pathway, which links leptin with metabolic and gene regulatory pathways, and a novel gene regulatory network with genes regulating chromatin accessibility at its core, as important hubs in the larger network of injury response genes involved in successful CNS axon regeneration.

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Chromatin accessibility dynamics and single cell RNA-Seq reveal new regulators of regeneration in neural progenitors

Cited by 51 publications

References 87 publications

Cellular response to spinal cord injury in regenerative and non-regenerative stages in Xenopus laevis

Cellular response to spinal cord injury in regenerative and non-regenerative stages in Xenopus laevis

One stem cell program to rule them all?

Comparative gene expression profiling between optic nerve and spinal cord injury in Xenopus laevis reveals a core set of genes inherent in successful regeneration of vertebrate central nervous system axons

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