Regulation Of Axon Regeneration Research Articles

A cytidine deaminase regulates axon regeneration by modulating the functions of the Caenorhabditis elegans HGF/plasminogen family protein SVH-1.

The pathway for axon regeneration in Caenorhabditis elegans is activated by SVH-1, a growth factor belonging to the HGF/plasminogen family. SVH-1 is a dual-function factor that acts as an HGF-like growth factor to promote axon regeneration and as a protease to regulate early development. It is important to understand how SVH-1 is converted from a protease to a growth factor for axon regeneration. In this study, we demonstrate that cytidine deaminase (CDD) SVH-17/CDD-2 plays a role in the functional conversion of SVH-1. We find that the codon exchange of His-755 to Tyr in the Asp-His-Ser catalytic triad of SVH-1 can suppress the cdd-2 defect in axon regeneration. Furthermore, the stem hairpin structure around the His-755 site in svh-1 mRNA is required for the activation of axon regeneration by SVH-1. These results suggest that CDD-2 promotes axon regeneration by transforming the function of SVH-1 from a protease to a growth factor through modification of svh-1 mRNA.

Mechanism research progress of acupuncture for regulating axon regeneration after stroke.

Stroke brings the pathological changes of brain tissues such as hematoma formation or ischemia and hypoxia, which leads to neuronal death and axon degeneration. Axon regeneration after its injury is mainly dependent on the surrounding microenvironment and the related proteins, including glial scar, myelin associated inhibitory factors, axon guidance molecules, and neurotrophic factors. All of them affect axon growth by regulating the morphology and orientation of growth cones, the synaptic stability, and the proliferation and differentiation of neural stem cells. This article summarizes the mechanism of acupuncture in regulating axon regeneration after stroke. Acupuncture inhibits glial scar formation, alleviates the inhibitory effects of its physical and chemical barriers on axon growth, reverses the inhibitory effects of myelin-related inhibitory factors on axon growth, and adjusts the level of axon guidance molecules to promote the proliferation and differentiation of neural stem cells and the regeneration of injured axons, and up-regulates neurotrophic factors. Eventually, post-stroke nerve injury can be ameliorated.

MicroRNA-2184 orchestrates Mauthner-cell axon regeneration in zebrafish via syt3 modulation

MicroRNAs (miRNAs) play a significant role in axon regeneration following spinal cord injury. However, the functions of numerous miRNAs in axon regeneration within the central nervous system (CNS) remain largely unexplored. Here, we elucidate the positive role of microRNA-2184 (miR-2184) in axon regeneration within zebrafish Mauthner cells (M-cells). The upregulation of miR-2184 in a single M-cell can facilitate axon regeneration, while the specific sponge-induced silencing of miR-2184 leads to impeded regeneration. We show that syt3, a downstream target of miR-2184, negatively regulates axon regeneration, and the regeneration suppression modulated by syt3 depends on its binding to Ca2+. Furthermore, pharmacological stimulation of the cAMP/PKA pathway suggests that changes in the readily releasable pool may affect axon regeneration. Our data indicate that miR-2184 promotes axon regeneration of M-cells within the CNS by modulating the downstream target syt3, providing valuable insights into potential therapeutic strategies.

MircoRNA-25-3p in skin precursor cell-induced Schwann cell-derived extracellular vesicles promotes axon regeneration by targeting Tgif1

Nerve injury often leads to severe dysfunction because of the lack of axon regeneration in adult mammal. Intriguingly a series of extracellular vesicles (EVs) have the obvious ability to accelerate the nerve repair. However, the detailed molecular mechanisms to describe that EVs switch neuron from a transmitter to a regenerative state have not been elucidated. This study elucidated the microRNA (miRNA) expression profiles of two types of EVs that promote nerve regeneration. The functions of these miRNAs were screened in vitro. Among the 12 overlapping miRNAs, miR-25-3p was selected for further analysis as it markedly promoted axon regeneration both in vivo and in vitro. Furthermore, knockdown experiments confirmed that PTEN and Klf4, which are the major inhibitors of axon regeneration, were the direct targets of miR-25-3p in dorsal root ganglion (DRG) neurons. The utilization of luciferase reporter assays and functional tests provided evidence that miR-25-3p enhances axon regeneration by targeting Tgif1. Additionally, miR-25-3p upregulated the phosphorylation of Erk. Furthermore, Rapamycin modulated the expression of miR-25-3p in DRG neurons. Finally, the pro-axon regeneration effects of EVs were confirmed by overexpressing miR-25-3p and Tgif1 knockdown in the optic nerve crush model. Thus, the enrichment of miR-25-3p in EVs suggests that it regulates axon regeneration, proving a potential cell-free treatment strategy for nerve injury.

Rhotekin regulates axon regeneration through the talin-Vinculin-Vinexin axis in Caenorhabditis elegans.

Axon regeneration requires actomyosin interaction, which generates contractile force and pulls the regenerating axon forward. In Caenorhabditis elegans, TLN-1/talin promotes axon regeneration through multiple down-stream events. One is the activation of the PAT-3/integrin-RHO-1/RhoA GTPase-LET-502/ROCK (Rho-associated coiled-coil kinase)-regulatory non-muscle myosin light-chain (MLC) phosphorylation signaling pathway, which is dependent on the MLC scaffolding protein ALP-1/ALP-Enigma. The other is mediated by the F-actin-binding protein DEB-1/vinculin and is independent of the MLC phosphorylation pathway. In this study, we identified the svh-7/rtkn-1 gene, encoding a homolog of the RhoA-binding protein Rhotekin, as a regulator of axon regeneration in motor neurons. However, we found that RTKN-1 does not function in the RhoA-ROCK-MLC phosphorylation pathway in the regulation of axon regeneration. We show that RTKN-1 interacts with ALP-1 and the vinculin-binding protein SORB-1/vinexin, and that SORB-1 acts with DEB-1 to promote axon regeneration. Thus, RTKN-1 links the DEB-1-SORB-1 complex to ALP-1 and physically connects phosphorylated MLC on ALP-1 to the actin cytoskeleton. These results suggest that TLN-1 signaling pathways coordinate MLC phosphorylation and recruitment of phosphorylated MLC to the actin cytoskeleton during axon regeneration.

Glycogen synthase kinase 3 signaling in neural regeneration in vivo.

Glycogen synthase kinase 3 (GSK3) signaling plays important and broad roles in regulating neural development in vitro and in vivo. Here, we reviewed recent findings of GSK3-regulated axon regeneration in vivo in both the peripheral and central nervous systems and discussed a few controversial findings in the field. Overall, current evidence indicates that GSK3β signaling serves as an important downstream mediator of the PI3K-AKT pathway to regulate axon regeneration in parallel with the mTORC1 pathway. Specifically, the mTORC1 pathway supports axon regeneration mainly through its role in regulating cap-dependent protein translation, whereas GSK3β signaling might be involved in regulating N6-methyladenosine mRNA methylation-mediated, cap-independent protein translation. In addition, GSK3 signaling also plays a key role in reshaping the neuronal transcriptomic landscape during neural regeneration. Finally, we proposed some research directions to further elucidate the molecular mechanisms underlying the regulatory function of GSK3 signaling and discover novel GSK3 signaling-related therapeutic targets. Together, we hope to provide an updated and insightful overview of how GSK3 signaling regulates neural regeneration in vivo.

Glia instruct axon regeneration via a ternary modulation of neuronal calcium channels in Drosophila

A neuron’s regenerative capacity is governed by its intrinsic and extrinsic environment. Both peripheral and central neurons exhibit cell-type-dependent axon regeneration, but the underlying mechanism is unclear. Glia provide a milieu essential for regeneration. However, the routes of glia-neuron signaling remain underexplored. Here, we show that regeneration specificity is determined by the axotomy-induced Ca2+ transients only in the fly regenerative neurons, which is mediated by L-type calcium channels, constituting the core intrinsic machinery. Peripheral glia regulate axon regeneration via a three-layered and balanced modulation. Glia-derived tumor necrosis factor acts through its neuronal receptor to maintain calcium channel expression after injury. Glia sustain calcium channel opening by enhancing membrane hyperpolarization via the inwardly-rectifying potassium channel (Irk1). Glia also release adenosine which signals through neuronal adenosine receptor (AdoR) to activate HCN channels (Ih) and dampen Ca2+ transients. Together, we identify a multifaceted glia-neuron coupling which can be hijacked to promote neural repair.

βPix Guanine Nucleotide Exchange Factor Regulates Regeneration of Injured Peripheral Axons.

Axon regeneration is essential for successful recovery after peripheral nerve injury. Although growth cone reformation and axonal extension are crucial steps in axonal regeneration, the regulatory mechanisms underlying these dynamic processes are poorly understood. Here, we identify βPix (Arhgef7), the guanine nucleotide exchange factor for Rac1 GTPase, as a regulator of axonal regeneration. After sciatic nerve injury in mice, the expression levels of βPix increase significantly in nerve segments containing regenerating axons. In regrowing axons, βPix is localized in the peripheral domain of the growth cone. Using βPix neuronal isoform knockout (NIKO) mice in which the neuronal isoforms of βPix are specifically removed, we demonstrate that βPix promotes neurite outgrowth in cultured dorsal root ganglion neurons and in vivo axon regeneration after sciatic nerve crush injury. Activation of cJun and STAT3 in the cell bodies is not affected in βPix NIKO mice, supporting the local action of βPix in regenerating axons. Finally, inhibiting Src, a kinase previously identified as an activator of the βPix neuronal isoform, causes axon outgrowth defects in vitro, like those found in the βPix NIKO neurons. Altogether, these data indicate that βPix plays an important role in axonal regrowth during peripheral nerve regeneration.

Promoting axon regeneration by inhibiting RNA N6-methyladenosine demethylase ALKBH5.

A key limiting factor of successful axon regeneration is the intrinsic regenerative ability in both the peripheral nervous system (PNS) and central nervous system (CNS). Previous studies have identified intrinsic regenerative ability regulators that act on gene expression in injured neurons. However, it is less known whether RNA modifications play a role in this process. Here, we systematically screened the functions of all common m6A modification-related enzymes in axon regeneration and report ALKBH5, an evolutionarily conserved RNA m6A demethylase, as a regulator of axonal regeneration in rodents. In PNS, knockdown of ALKBH5 enhanced sensory axonal regeneration, whereas overexpressing ALKBH5 impaired axonal regeneration in an m6A-dependent manner. Mechanistically, ALKBH5 increased the stability of Lpin2 mRNA and thus limited regenerative growth associated lipid metabolism in dorsal root ganglion neurons. Moreover, in CNS, knockdown of ALKBH5 enhanced the survival and axonal regeneration of retinal ganglion cells after optic nerve injury. Together, our results suggest a novel mechanism regulating axon regeneration and point ALKBH5 as a potential target for promoting axon regeneration in both PNS and CNS.

TIR-1/SARM1 inhibits axon regeneration and promotes axon degeneration

Growth and destruction are central components of the neuronal injury response. Injured axons that are capable of repair, including axons in the mammalian peripheral nervous system and in many invertebrate animals, often regenerate and degenerate on either side of the injury. Here we show that TIR-1/dSarm/SARM1, a key regulator of axon degeneration, also inhibits regeneration of injured motor axons. The increased regeneration in tir-1 mutants is not a secondary consequence of its effects on degeneration, nor is it determined by the NADase activity of TIR-1. Rather, we found that TIR-1 functions cell-autonomously to regulate each of the seemingly opposite processes through distinct interactions with two MAP kinase pathways. On one side of the injury, TIR-1 inhibits axon regeneration by activating the NSY-1/ASK1 MAPK signaling cascade, while on the other side of the injury, TIR-1 simultaneously promotes axon degeneration by interacting with the DLK-1 mitogen-activated protein kinase (MAPK) signaling cascade. In parallel, we found that the ability to cell-intrinsically inhibit axon regeneration is conserved in human SARM1. Our finding that TIR-1/SARM1 regulates axon regeneration provides critical insight into how axons coordinate a multidimensional response to injury, consequently informing approaches to manipulate the response toward repair.

Regulation of axonal regeneration after mammalian spinal cord injury.

One hundred years ago, Ramón y Cajal, considered by many as the founder of modern neuroscience, stated that neurons of the adult central nervous system (CNS) are incapable of regenerating. Yet, recent years have seen a tremendous expansion of knowledge in the molecular control of axon regeneration after CNS injury. We now understand that regeneration in the adult CNS is limited by (1) a failure to form cellular or molecular substrates for axon attachment and elongation through the lesion site; (2) environmental factors, including inhibitors of axon growth associated with myelin and the extracellular matrix; (3)astrocyte responses, which can both limit and support axon growth; and (4) intraneuronal mechanisms controlling the establishment of an active cellular growth programme. We discuss these topics together with newly emerging hypotheses, including the surprising finding from transcriptomic analyses of the corticospinal system in mice that neurons revert to an embryonic state after spinal cord injury, which can be sustained to promote regeneration with neural stem cell transplantation. These gains in knowledge are steadily advancing efforts to develop effective treatment strategies for spinal cord injury inhumans.

Analysis of neuronal injury transcriptional response identifies CTCF and YY1 as co-operating factors regulating axon regeneration.

Injured sensory neurons activate a transcriptional program necessary for robust axon regeneration and eventual target reinnervation. Understanding the transcriptional regulators that govern this axon regenerative response may guide therapeutic strategies to promote axon regeneration in the injured nervous system. Here, we used cultured dorsal root ganglia neurons to identify pro-regenerative transcription factors. Using RNA sequencing, we first characterized this neuronal culture and determined that embryonic day 13.5 DRG (eDRG) neurons cultured for 7 days are similar to e15.5 DRG neurons in vivo and that all neuronal subtypes are represented. This eDRG neuronal culture does not contain other non-neuronal cell types. Next, we performed RNA sequencing at different time points after in vitro axotomy. Analysis of differentially expressed genes revealed upregulation of known regeneration associated transcription factors, including Jun, Atf3 and Rest, paralleling the axon injury response in vivo. Analysis of transcription factor binding sites in differentially expressed genes revealed other known transcription factors promoting axon regeneration, such as Myc, Hif1α, Pparγ, Ascl1a, Srf, and Ctcf, as well as other transcription factors not yet characterized in axon regeneration. We next tested if overexpression of novel candidate transcription factors alone or in combination promotes axon regeneration in vitro. Our results demonstrate that expression of Ctcf with Yy1 or E2f2 enhances in vitro axon regeneration. Our analysis highlights that transcription factor interaction and chromatin architecture play important roles as a regulator of axon regeneration.

Non-canonical role of the ATR pathway in axon regeneration as a mechanosensitive brake.

Non-canonical role of the ATR pathway in axon regeneration as a mechanosensitive brake.

Morpholino studies shed light on the signaling pathways regulating axon regeneration in lampreys.

Morpholino studies shed light on the signaling pathways regulating axon regeneration in lampreys.

Rab-27 acts in an intestinal pathway to inhibit axon regeneration in C. elegans.

Injured axons must regenerate to restore nervous system function, and regeneration is regulated in part by external factors from non-neuronal tissues. Many of these extrinsic factors act in the immediate cellular environment of the axon to promote or restrict regeneration, but the existence of long-distance signals regulating axon regeneration has not been clear. Here we show that the Rab GTPase rab-27 inhibits regeneration of GABAergic motor neurons in C. elegans through activity in the intestine. Re-expression of RAB-27, but not the closely related RAB-3, in the intestine of rab-27 mutant animals is sufficient to rescue normal regeneration. Several additional components of an intestinal neuropeptide secretion pathway also inhibit axon regeneration, including NPDC1/cab-1, SNAP25/aex-4, KPC3/aex-5, and the neuropeptide NLP-40, and re-expression of these genes in the intestine of mutant animals is sufficient to restore normal regeneration success. Additionally, NPDC1/cab-1 and SNAP25/aex-4 genetically interact with rab-27 in the context of axon regeneration inhibition. Together these data indicate that RAB-27-dependent neuropeptide secretion from the intestine inhibits axon regeneration, and point to distal tissues as potent extrinsic regulators of regeneration.

RhoA drives actin compaction to restrict axon regeneration and astrocyte reactivity after CNS injury

An inhibitory extracellular milieu and neuron-intrinsic processes prevent axons from regenerating in the adult central nervous system (CNS). Here we show how the two aspects are interwoven. Genetic loss-of-function experiments determine that the small GTPase RhoA relays extracellular inhibitory signals to the cytoskeleton by adapting mechanisms set in place during neuronal polarization. In response to extracellular inhibitors, neuronal RhoA restricts axon regeneration by activating myosin II to compact actin and, thereby, restrain microtubule protrusion. However, astrocytic RhoA restricts injury-induced astrogliosis through myosin II independent of microtubules by activating Yes-activated protein (YAP) signaling. Cell-type-specific deletion in spinal-cord-injured mice shows that neuronal RhoA activation prevents axon regeneration, whereas astrocytic RhoA is beneficial for regenerating axons. These data demonstrate how extracellular inhibitors regulate axon regeneration, shed light on the capacity of reactive astrocytes to be growth inhibitory after CNS injury, and reveal cell-specific RhoA targeting as a promising therapeutic avenue.

Genome-wide chromatin accessibility analyses provide a map for enhancing optic nerve regeneration

Retinal Ganglion Cells (RGCs) lose their ability to grow axons during development. Adult RGCs thus fail to regenerate their axons after injury, leading to vision loss. To uncover mechanisms that promote regeneration of RGC axons, we identified transcription factors (TF) and open chromatin regions that are enriched in rat embryonic RGCs (high axon growth capacity) compared to postnatal RGCs (low axon growth capacity). We found that developmental stage-specific gene expression changes correlated with changes in promoter chromatin accessibility. Binding motifs for TFs such as CREB, CTCF, JUN and YY1 were enriched in the regions of the chromatin that were more accessible in embryonic RGCs. Proteomic analysis of purified rat RGC nuclei confirmed the expression of TFs with potential role in axon growth such as CREB, CTCF, YY1, and JUND. The CREB/ATF binding motif was widespread at the open chromatin region of known pro-regenerative TFs, supporting a role of CREB in regulating axon regeneration. Consistently, overexpression of CREB fused to the VP64 transactivation domain in mouse RGCs promoted axon regeneration after optic nerve injury. Our study provides a map of the chromatin accessibility during RGC development and highlights that TF associated with developmental axon growth can stimulate axon regeneration in mature RGC.

Regulation of Axonal Regeneration and Its Inhibition by Glycan Ligands

Regulation of Axonal Regeneration and Its Inhibition by Glycan Ligands

Regulation of Axonal Regeneration and Its Inhibition by Glycan Ligands

Regulation of Axonal Regeneration and Its Inhibition by Glycan Ligands

The Integrin Signaling Network Promotes Axon Regeneration via the Src-Ephexin-RhoA GTPase Signaling Axis.

Axon regeneration is an evolutionarily conserved process essential for restoring the function of damaged neurons. In Caenorhabditis elegans hermaphrodites, initiation of axon regeneration is regulated by the RhoA GTPase–ROCK (Rho-associated coiled-coil kinase)–regulatory nonmuscle myosin light-chain phosphorylation signaling pathway. However, the upstream mechanism that activates the RhoA pathway remains unknown. Here, we show that axon injury activates TLN-1/talin via the cAMP–Epac (exchange protein directly activated by cAMP)–Rap GTPase cascade and that TLN-1 induces multiple downstream events, one of which is integrin inside-out activation, leading to the activation of the RhoA–ROCK signaling pathway. We found that the nonreceptor tyrosine kinase Src, a key mediator of integrin signaling, activates the Rho guanine nucleotide exchange factor EPHX-1/ephexin by phosphorylating the Tyr-568 residue in the autoinhibitory domain. Our results suggest that the C. elegans integrin signaling network regulates axon regeneration via the Src–RhoGEF–RhoA axis.SIGNIFICANCE STATEMENT The ability of axons to regenerate after injury is governed by cell-intrinsic regeneration pathways. We have previously demonstrated that the Caenorhabditis elegans RhoA GTPase–ROCK (Rho-associated coiled-coil kinase) pathway promotes axon regeneration by inducing MLC-4 phosphorylation. In this study, we found that axon injury activates TLN-1/talin through the cAMP–Epac (exchange protein directly activated by cAMP)–Rap GTPase cascade, leading to integrin inside-out activation, which promotes axonal regeneration by activating the RhoA signaling pathway. In this pathway, SRC-1/Src acts downstream of integrin activation and subsequently activates EPHX-1/ephexin RhoGEF by phosphorylating the Tyr-568 residue in the autoinhibitory domain. Our results suggest that the C. elegans integrin signaling network regulates axon regeneration via the Src–RhoGEF–RhoA axis.