Sensory neurons with cell bodies in dorsal root ganglia (DRG) represent a useful model to study axon regeneration. Whereas regeneration and functional recovery occurs after peripheral nerve injury, spinal cord injury or dorsal root injury is not followed by regenerative outcomes. Regeneration of sensory axons in peripheral nerves is not entirely cell autonomous. Whether the DRG microenvironment influences the different regenerative capacities after injury to peripheral or central axons remains largely unknown. To answer this question, we performed a single-cell transcriptional profiling of mouse DRG in response to peripheral (sciatic nerve crush) and central axon injuries (dorsal root crush and spinal cord injury). Each cell type responded differently to the three types of injuries. All injuries increased the proportion of a cell type that shares features of both immune cells and glial cells. A distinct subset of satellite glial cells (SGC) appeared specifically in response to peripheral nerve injury. Activation of the PPARα signaling pathway in SGC, which promotes axon regeneration after peripheral nerve injury, failed to occur after central axon injuries. Treatment with the FDA-approved PPARα agonist fenofibrate increased axon regeneration after dorsal root injury. This study provides a map of the distinct DRG microenvironment responses to peripheral and central injuries at the single-cell level and highlights that manipulating non-neuronal cells could lead to avenues to promote functional recovery after CNS injuries or disease.

Profiling sensory neuron microenvironment after peripheral and central axon injury reveals key pathways for neural repair

The vertebrate nervous system exhibits dramatic variability in regenerative capacity across species and neuronal populations. For example, while the mammalian central nervous system (CNS) is limited in its regenerative capacity, the CNS of many other vertebrates readily regenerates after injury, as does the peripheral nervous system (PNS) of mammals. Comparing molecular responses across species and tissues can therefore provide valuable insights into both conserved and distinct mechanisms of successful regeneration. One gene that is emerging as a conserved pro-regenerative factor across vertebrates is activating transcription factor 3 (ATF3), which has long been associated with tissue trauma. A growing number of studies indicate that ATF3 may actively promote neuronal axon regrowth and regeneration in species ranging from lampreys to mammals. Here, we review data on the structural and functional conservation of ATF3 protein across species. Comparing RNA expression data across species that exhibit different abilities to regenerate their nervous system following traumatic nerve injury reveals that ATF3 is consistently induced in neurons within the first few days after injury. Genetic deletion or knockdown of ATF3 expression has been shown in mouse and zebrafish, respectively, to reduce axon regeneration, while inducing ATF3 promotes axon sprouting, regrowth, or regeneration. Thus, we propose that ATF3 may be an evolutionarily conserved regulator of neuronal regeneration. Identifying downstream effectors of ATF3 will be a critical next step in understanding the molecular basis of vertebrate CNS regeneration.

/pdf/activating-transcription-factor-3-atf3-is-a-highly-conserved-3btxqkmq.pdf

Activating Transcription Factor 3 (ATF3) is a Highly Conserved Pro-regenerative Transcription Factor in the Vertebrate Nervous System

Axons in the adult mammalian nervous system can extend over formidable distances, up to one meter or more in humans. During development, axonal and dendritic growth requires continuous addition of new membrane. Of the three major kinds of membrane lipids, phospholipids are the most abundant in all cell membranes, including neurons. Not only immature axons, but also severed axons in the adult require large amounts of lipids for axon regeneration to occur. Lipids also serve as energy storage, signaling molecules and they contribute to tissue physiology, as demonstrated by a variety of metabolic disorders in which harmful amounts of lipids accumulate in various tissues through the body. Detrimental changes in lipid metabolism and excess accumulation of lipids contribute to a lack of axon regeneration, poor neurological outcome and complications after a variety of central nervous system (CNS) trauma including brain and spinal cord injury. Recent evidence indicates that rewiring lipid metabolism can be manipulated for therapeutic gain, as it favors conditions for axon regeneration and CNS repair. Here, we review the role of lipids, lipid metabolism and ectopic lipid accumulation in axon growth, regeneration and CNS repair. In addition, we outline molecular and pharmacological strategies to fine-tune lipid composition and energy metabolism in neurons and non-neuronal cells that can be exploited to improve neurological recovery after CNS trauma and disease.

The Role of Lipids, Lipid Metabolism and Ectopic Lipid Accumulation in Axon Growth, Regeneration and Repair after CNS Injury and Disease

Sensory neurons with cell bodies in dorsal root ganglia (DRG) represent a useful model to study axon regeneration. Whereas regeneration and functional recovery occurs after peripheral nerve injury, spinal cord injury or dorsal root injury is not followed by regenerative outcomes. This results in part from a failure of central injury to elicit a pro-regenerative response in sensory neurons. However, regeneration of sensory axons in peripheral nerves is not entirely cell autonomous. Whether the different regenerative capacities after peripheral or central injury result in part from a lack of response of macrophages, satellite glial cells (SGC) or other non-neuronal cells in the DRG microenvironment remains largely unknown. To answer this question, we performed a single cell transcriptional profiling of DRG in response to peripheral (sciatic nerve crush) and central injuries (dorsal root crush and spinal cord injury). Each cell type responded differently to peripheral and central injuries. Activation of the PPAR signaling pathway in SGC, which promotes axon regeneration after nerve injury, did not occur after central injuries. Treatment with the FDA-approved PPARα agonist fenofibrate, increased axon regeneration after dorsal root injury. This study provides a map of the distinct DRG microenvironment responses to peripheral and central injuries at the single cell level and highlights that manipulating non-neuronal cells could lead to avenues to promote functional recovery after CNS injuries.

https://www.biorxiv.org/content/biorxiv/early/2020/11/27/2020.11.25.398537.full.pdf

Profiling sensory neuron microenvironment after peripheral and central axon injury reveals key pathways for axon regeneration

Abstract The dynamic changes in chromatin conformation alter the organization and structure of the genome and further regulate gene transcription. Basically, the chromatin structure is controlled by reversible, enzyme-catalyzed covalent modifications to chromatin components and by noncovalent ATP-dependent modifications via chromatin remodeling complexes, including switch/sucrose nonfermentable (SWI/SNF), inositol-requiring 80 (INO80), imitation switch (ISWI) and chromodomain-helicase DNA-binding protein (CHD) complexes. Recent studies have shown that chromatin remodeling is essential in different stages of postnatal and adult neurogenesis. Chromatin deregulation, which leads to defects in epigenetic gene regulation and further pathological gene expression programs, often causes a wide range of pathologies. This review first gives an overview of the regulatory mechanisms of chromatin remodeling. We then focus mainly on discussing the physiological functions of chromatin remodeling, particularly histone and DNA modifications and the four classes of ATP-dependent chromatin-remodeling enzymes, in the central and peripheral nervous systems under healthy and pathological conditions, that is, in neurodegenerative disorders. Finally, we provide an update on the development of potent and selective small molecule modulators targeting various chromatin-modifying proteins commonly associated with neurodegenerative diseases and their potential clinical applications. 

/pdf/small-molecule-modulators-of-chromatin-remodeling-from-1skcztw6.pdf

Small molecule modulators of chromatin remodeling: from neurodevelopment to neurodegeneration

Regeneration failure after spinal cord injury (SCI) results in part from the lack of a pro-regenerative response in injured neurons, but the response to SCI has not been examined specifically in injured sensory neurons. Using RNA sequencing of dorsal root ganglion, we determined that thoracic SCI elicits a transcriptional response distinct from sciatic nerve injury (SNI). Both SNI and SCI induced upregulation of ATF3 and Jun, yet this response failed to promote growth in sensory neurons after SCI. RNA sequencing of purified sensory neurons one and three days after injury revealed that unlike SNI, the SCI response is not sustained. Both SCI and SNI elicited the expression of ATF3 target genes, with very little overlap between conditions. Pathway analysis of differentially expressed ATF3 target genes revealed that fatty acid biosynthesis and terpenoid backbone synthesis were downregulated after SCI but not SNI. Pharmacologic inhibition of fatty acid synthase, the enzyme generating palmitic acid, decreased axon growth and regeneration in vitro. These results support the notion that decreased expression of lipid metabolism-related genes after SCI, including fatty acid synthase, may restrict axon regenerative capacity after SCI.

/pdf/ascending-dorsal-column-sensory-neurons-respond-to-spinal-42oze2ox8n.pdf

Ascending dorsal column sensory neurons respond to spinal cord injury and downregulate genes related to lipid metabolism.

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.

/pdf/genome-wide-chromatin-accessibility-analyses-provide-a-map-2f3rwmnw6b.pdf

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

Regulation of chromatin plays fundamental roles in the development of the brain. Haploinsufficiency of the chromatin remodeling enzyme CHD7 causes CHARGE syndrome, a genetic disorder that affects the development of the cerebellum. However, how CHD7 controls chromatin states in the cerebellum remains incompletely understood. Using conditional knockout of CHD7 in granule cell precursors in the mouse cerebellum, we find that CHD7 robustly promotes chromatin accessibility, active histone modifications, and RNA polymerase recruitment at enhancers. In vivo profiling of genome architecture reveals that CHD7 concordantly regulates epigenomic modifications associated with enhancer activation and gene expression of topologically-interacting genes. Genome and gene ontology studies show that CHD7-regulated enhancers are associated with genes that control brain tissue morphogenesis. Accordingly, conditional knockout of CHD7 triggers a striking phenotype of cerebellar polymicrogyria, which we have also found in a case of CHARGE syndrome. Finally, we uncover a CHD7-dependent switch in the preferred orientation of granule cell precursor division in the developing cerebellum, providing a potential cellular basis for the cerebellar polymicrogyria phenotype upon loss of CHD7. Collectively, our findings define epigenomic regulation by CHD7 in granule cell precursors and identify abnormal cerebellar patterning upon CHD7 depletion, with potential implications for our understanding of CHARGE syndrome.

/pdf/charge-syndrome-protein-chd7-regulates-epigenomic-activation-zpkfsvnyee.pdf

CHARGE syndrome protein CHD7 regulates epigenomic activation of enhancers in granule cell precursors and gyrification of the cerebellum.

Regeneration failure after spinal cord injury (SCI) results in part from the lack of a pro-regenerative response in injured neurons, but the response to SCI has not been examined specifically in injured sensory neurons. Using RNA sequencing of dorsal root ganglion, we determined that thoracic SCI elicits a transcriptional response distinct from sciatic nerve injury (SNI). Both SNI and SCI induced upregulation of ATF3 and Jun, yet this response failed to promote growth in sensory neurons after SCI. RNA sequencing of purified sensory neurons one and three days after injury revealed that unlike SNI, the SCI response is not sustained. Analysis of differentially expressed genes revealed that lipid biosynthetic pathways, including fatty acid biosynthesis, were differentially regulated after SCI and SNI. Pharmacologic inhibition of fatty acid synthase, the enzyme generating fatty acid, decreased axon growth in vitro. These findings suggest that decreased fatty acid synthesis inhibits axon regeneration after SCI.

/pdf/ascending-dorsal-column-sensory-neurons-respond-to-spinal-4ibb6slyey.pdf

Tassia Mangetti Gonçalves

Papers

Ascending dorsal column sensory neurons respond to spinal cord injury and downregulate genes related to lipid metabolism.

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

CHARGE syndrome protein CHD7 regulates epigenomic activation of enhancers in granule cell precursors and gyrification of the cerebellum.

Ascending dorsal column sensory neurons respond to spinal cord injury and downregulate genes related to lipid metabolism