Amputation of a salamander tail leads to functional spinal cord regeneration through activation of endogenous stem cells. Identifying the signaling pathways that control cell proliferation in these neural stem cells will help elucidate the mechanisms underlying the salamander’s regenerative ability. Here, we show that neuregulin 1 (Nrg1)/ErbB2 signaling is an important pathway in the regulation of neural stem cell proliferation in the spinal cord of the axolotl salamander (
- Award ID(s):
- 1757443
- NSF-PAR ID:
- 10423102
- Date Published:
- Journal Name:
- Development
- Volume:
- 149
- Issue:
- 6
- ISSN:
- 0950-1991
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
More Like this
-
more » « less
Abstract Ambystoma mexicanum ). Simultaneous localization ofnrg1 mRNA and Nrg1 protein was performed by utilizing a hybridization chain reaction fluorescencein situ hybridization (FISH) methodology in tissue sections. Multiplexed FISH also permitted the phenotyping of multiple cell types on a single fixed section allowing the characterization of mRNA expression, protein expression, and tissue architecture. Pharmacological inhibition of ErbB2 showed that intact Nrg1/ErbB2 signaling is critical for adult homeostatic regeneration as well as for injury‐induced spinal cord regeneration. Overall, our results highlight the importance of the NRG1/ErbB2 signaling pathway in neural stem cell proliferation in the axolotl. -
Abstract Background The ability to regenerate body parts is a feature of metazoan organisms and the focus of intense research aiming to understand its basis. A number of mechanisms involved in regeneration, such as proliferation and tissue remodeling, affect whole tissues; however, little is known on how distinctively different constituent cell types respond to the dynamics of regenerating tissues. Preliminary studies suggest that a number of organisms alter neuronal numbers to scale with changes in body size. In some species with the ability of whole-body axis regeneration, it has additionally been observed that regenerates are smaller than their pre-amputated parent, but maintain the correct morphological proportionality, suggesting that scaling of tissue and neuronal numbers also occurs. However, the cell dynamics and responses of neuronal subtypes during nervous system regeneration, scaling, and whole-body axis regeneration are not well understood in any system. The cnidarian sea anemone Nematostella vectensis is capable of whole-body axis regeneration, with a number of observations suggesting the ability to alter its size in response to changes in feeding. We took advantage of Nematostella ’s transparent and “simple” body plan and the NvLWamide-like mCherry fluorescent reporter transgenic line to probe the response of neuron populations to variations in body size in vivo in adult animals during body scaling and regeneration. Results We utilized the previously characterized NvLWamide-like::mCherry transgenic reporter line to determine the in vivo response of neuronal subtypes during growth, degrowth, and regeneration. Nematostella alters its size in response to caloric intake, and the nervous system responds by altering neuronal number to scale as the animal changes in size. Neuronal numbers in both the endodermal and ectodermal nerve nets decreased as animals shrunk, increased as they grew, and these changes were reversible. Whole-body axis regeneration resulted in regenerates that were smaller than their pre-amputated size, and the regenerated nerve nets were reduced in neuronal number. Different neuronal subtypes had distinct responses during regeneration, including consistent, not consistent, and conditional increases in number. Conditional responses were regulated, in part, by the size of the remnant fragment and the position of the amputation site. Regenerates and adults with reduced nerve nets displayed normal behaviors, indicating that the nerve net retains functionality as it scales. Conclusion These data suggest that the Nematostella nerve net is dynamic, capable of scaling with changes in body size, and that neuronal subtypes display differential regenerative responses, which we propose may be linked to the scale state of the regenerating animals.more » « less
-
more » « less
Abstract During the early stages of limb and fin regeneration in aquatic vertebrates (i.e., fishes and amphibians), blastema undergo transcriptional rewiring of innate immune signaling pathways to promote immune cell recruitment. In mammals, a fundamental component of innate immune signaling is the cytosolic DNA sensing pathway, cGAS‐STING. However, to what extent the cGAS‐STING pathway influences regeneration in aquatic anamniotes is unknown. In jawed vertebrates, negative regulation of cGAS‐STING activity is accomplished by suppressors of cytosolic DNA such as Trex1, Pml, and PML‐like exon 9 (Plex9) exonucleases. Here, we examine the expression of these suppressors of cGAS‐STING, as well as inflammatory genes and cGAS activity during caudal fin and limb regeneration using the spotted gar (
Lepisosteus oculatus ) and axolotl (Ambystoma mexicanum ) model species, and during age‐related senescence in zebrafish (Danio rerio ). In the regenerative blastema of wounded gar and axolotl, we observe increased inflammatory gene expression, including interferon genes and interleukins 6 and 8. We also observed a decrease in axolotlTrex1 and garpml expression during the early phases of wound healing which correlates with a dramatic increase in cGAS activity. In contrast, theplex9.1 gene does not change in expression during wound healing in gar. However, we observed decreased expression ofplex9.1 in the senescing cardiac tissue of aged zebrafish, where 2′3′‐cGAMP levels are elevated. Finally, we demonstrate a similar pattern ofTrex1 ,pml , andplex9.1 gene regulation across species in response to exogenous 2′3′‐cGAMP. Thus, during the early stages of limb‐fin regeneration, Pml, Trex1, and Plex9.1 exonucleases are downregulated, presumably to allow an evolutionarily ancient cGAS‐STING activity to promote inflammation and the recruitment of immune cells. -
Salamanders and lungfishes are the only sarcopterygians (lobe-finned vertebrates) capable of paired appendage regeneration, regardless of the amputation level. Among actinopterygians (ray-finned fishes), regeneration after amputation at the fin endoskeleton has only been demonstrated in polypterid fishes (Cladistia). Whether this ability evolved independently in sarcopterygians and actinopterygians or has a common origin remains unknown. Here we combine fin regeneration assays and comparative RNA-sequencing (RNA-seq) analysis of Polypterus and axolotl blastemas to provide support for a common origin of paired appendage regeneration in Osteichthyes (bony vertebrates). We show that, in addition to polypterids, regeneration after fin endoskeleton amputation occurs in extant representatives of 2 other nonteleost actinopterygians: the American paddlefish (Chondrostei) and the spotted gar (Holostei). Furthermore, we assessed regeneration in 4 teleost species and show that, with the exception of the blue gourami (Anabantidae), 3 species were capable of regenerating fins after endoskeleton amputation: the white convict and the oscar (Cichlidae), and the goldfish (Cyprinidae). Our comparative RNA-seq analysis of regenerating blastemas of axolotl and Polypterus reveals the activation of common genetic pathways and expression profiles, consistent with a shared genetic program of appendage regeneration. Comparison of RNA-seq data from early Polypterus blastema to single-cell RNA-seq data from axolotl limb bud and limb regeneration stages shows that Polypterus and axolotl share a regeneration-specific genetic program. Collectively, our findings support a deep evolutionary origin of paired appendage regeneration in Osteichthyes and provide an evolutionary framework for studies on the genetic basis of appendage regeneration.more » « less
-
more » « less
Abstract This study investigates the response to spinal cord injury in the gray short‐tailed opossum (
Monodelphis domestica ). In opossums spinal injury early in development results in spontaneous axon growth through the injury, but this regenerative potential diminishes with maturity until it is lost entirely. The mechanisms underlying this regeneration remain unknown. RNA sequencing was used to identify differential gene expression in regenerating (SCI at postnatal Day 7, P7SCI) and nonregenerating (SCI at Day 28, P28SCI) cords +1d, +3d, and +7d after complete spinal transection, compared to age‐matched controls. Genes showing significant differential expression (log2FC ≥ 1,Padj ≤ 0.05) were used for downstream analysis. Across all time‐points 233 genes altered expression after P7SCI, and 472 genes altered expression after P28SCI. One hundred and forty‐seven genes altered expression in both injury ages (63% of P7SCI data set). The majority of changes were gene upregulations. Gene ontology overrepresentation analysis in P7SCI gene‐sets showed significant overrepresentations only in immune‐associated categories, while P28SCI gene‐sets showed overrepresentations in these same immune categories, along with other categories such as “cell proliferation,” “cell adhesion,” and “apoptosis.” Cell‐type–association analysis suggested that, regardless of injury age, injury‐associated gene transcripts were most strongly associated with microglia and endothelial cells, with strikingly fewer astrocyte, oligodendrocyte and neuron‐related genes, the notable exception being a cluster of mostly downregulated oligodendrocyte‐associated genes in the P7SCI + 7d gene‐set. Our findings demonstrate a more complex transcriptomic response in nonregenerating cords, suggesting a strong influence of non‐neuronal cells in the outcome after injury and providing the largest survey yet of the transcriptomic changes occurring after SCI in this model.
- Free Publicly Accessible Full Text
-
Accepted Manuscript1.0
- Journal Article:
- https://doi.org/10.1242/dev.199637
