Epigenomic signatures underpin the axonal regenerative ability of dorsal root ganglia sensory neurons

Palmisano, Ilaria; Danzi, Matt C.; Hutson, Thomas E.; Zhou, Luming; McLachlan, Eilidh; Serger, Elisabeth; Shkura, Kirill; Srivastava, Prashant K.; Hervera, Arnau; Neill, Nick O’; Liu, Tong; Dhrif, Hassen; Wang, Zheng; Kubát, Miroslav; Wuchty, Stefan; Merkenschlager, Matthias; Levi, Liron; Elliott, Evan; Bixby, John L.; Giovanni, Simone Di

doi:10.1038/s41593-019-0490-4

Cited by 82 publications

(105 citation statements)

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“…A key observation from the cell culture experiments is that none of the hit TFs were predicted to co-occupy solely in promoter regions; rather, all hit TFs were characterized by predicted co-occupancy in both promoters and enhancers. This suggests a key role for enhancer elements in regulating pro-growth gene transcription, in line with the recent finding in DRG neurons that modulation of enhancers may be critical for transcriptional activation of growth genes 13,42 . Finally, guided by network analysis of the screening results, a systematic campaign of combinatorial testing in vivo identified highly consistent enhancement of CST axon growth by combined expression of Klf6 and Nr5a2.…”

Section: Discussionsupporting

confidence: 85%

“…It has, however, been linked to the differentiation of neural stem cells and to cellular reprogramming, and can replace Oct4 in the reprogramming of murine somatic cells to pluripotent cells 38,39 . Our present work adds to the growing body of evidence that TFs involved in neuronal differentiation may positively influence axon growth 9,13 .…”

Section: Discussionmentioning

confidence: 54%

“…One likely possibility is that because multiple TFs generally act in a coordinated manner to regulate transcription, a more complete restoration of axon growth will depend on multiple TFs. Consistent with this, regeneration competent neurons recruit groups of interacting TFs in the hours to days post-injury, which results in transcriptional remodeling leading to reactivation of growth gene networks and functional recovery [13][14][15] . The need for multi-TF interventions in CNS neurons is recognized conceptually 5,16 , but a major challenge has been to develop a systematic pipeline aimed at discovering growth-relevant TF combinations.…”

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confidence: 60%

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Co-occupancy analysis reveals novel transcriptional synergies for axon growth

Venkatesh

Mehra

Wang

et al. 2020

Preprint

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Transcription factors (TFs) act as powerful levers to regulate neural physiology and can be targeted to improve cellular responses to injury or disease. Because TFs often depend on cooperative activity, a major challenge is to identify and deploy optimal sets. Here we developed a novel bioinformatics pipeline, centered on TF co-occupancy of regulatory DNA, and used it to predict factors that improve axon growth in corticospinal tract (CST) axons when combined with a known pro-regenerative TF, Klf6. Assays of neurite outgrowth confirmed cooperative activity by 12 candidates, and in vivo testing showed strong promotion of CST growth upon combined expression of Klf6 and Nr5a2. Transcriptional profiling of CST neurons identified Klf6/Nr5a2-responsive gene networks involved in macromolecule biosynthesis and DNA repair. These data identify novel TF combinations that promote enhanced CST growth, clarify the transcriptional correlates, and provide a bioinformatics roadmap to detect TF synergy. Introduction :As they mature, neurons in the central nervous system (CNS) decline in their capacity for robust axon growth, which broadly limits recovery from injury 1-4 . Axon growth depends on the transcription of large networks of regeneration-associated genes (RAGs), and coaxing activation of these networks in adult CNS neurons is a major unmet goal 2,5 . During periods of developmental axon growth, RAG expression is supported by pro-growth transcription factors (TFs) that bind to relevant promoter and enhancer regions to activate transcription 3,6 . One factor that limits RAG expression in mature neurons is the developmental downregulation of these pro-growth TFs 2,5 . Thus identifying TFs that act developmentally to enable axon growth and supplying them to mature neurons is a promising approach to improve regenerative outcomes in the injured nervous system.An ongoing challenge, however, is to decode the optimal set of factors for axon growth. To date, progress has centered on identifying individual TFs whose ectopic expression in mature neurons leads to improved axon growth. For example, we showed previously that the TF Klf6 is developmentally downregulated and that viral re-expression drives improved axon growth in mature corticospinal neurons, which are critical mediators of fine movement 7 . The restoration of axon growth from this and other single-factor studies remains partial, however, indicating the need for additional intervention. 7-12 . One likely possibility is that because multiple TFs generally act in a coordinated manner to regulate transcription, a more complete restoration of axon growth will depend on multiple TFs. Consistent with this, regeneration competent neurons recruit groups of interacting TFs in the hours to days post-injury, which results in transcriptional remodeling leading to reactivation of growth gene networks and functional recovery [13][14][15] . The need for multi-TF interventions in CNS neurons is recognized conceptually 5,16 , but a major challenge has been to develop a systematic pipeline ...

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Section: Discussionsupporting

confidence: 85%

Section: Discussionmentioning

confidence: 54%

mentioning

confidence: 60%

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Co-occupancy analysis reveals novel transcriptional synergies for axon growth

Venkatesh

Mehra

Wang

et al. 2020

Preprint

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“…Interestingly, a recent study (Palmisano et al, 2019) suggested that sensory axon regeneration shared similar molecular pathways with that during developmental axon growth. Another study (Poplawski et al, 2020) also showed that CNS neurons undergoing successful axon regeneration re-express many developmental related genes.…”

Section: Discussionmentioning

confidence: 99%

N6-methyladenine DNA demethylase ALKBH1 regulates mammalian axon regeneration

Qian

Feng

et al. 2020

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Recent studies have shown that DNA N6-methyladenine (N6-mA) modification is emerging to be a novel and important epigenetic regulator of mammalian gene transcription. Several studies demonstrated DNA N6-mA in human or rodents was regulated by methyltransferase N6AMT1 and demethylase ALKBH1. Moreover, studies in mouse brain or human glioblastoma cells showed that reduced level of N6-mA or higher level of ALKBH1 was correlated with up regulated levels of genes associated with neuronal development. We thus investigated the functional roles of ALKBH1 in sensory axon regeneration. Our results showed that ALKBH1 regulated the level of N6-mA in sensory neurons, and upon peripheral nerve injury ALKBH1 was up regulated in mouse sensory neurons. Functionally, knocking down ALKBH1 in sensory neurons resulted in reduced axon regeneration in vitro and in vivo, which could be rescued by simultaneously knocking down N6AMT1. Moreover, knocking down ALKBH1 led to decreased levels of many neurodevelopment regulatory genes, including neuritin that is well known to enhance axon growth and regeneration. Our study not only revealed a novel physiological function of DNA N6-mA, but also identified a new epigenetic mechanism regulating mammalian axon regeneration.

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“…Peripheral nerve injury triggers dynamic chromatin accessibility in DRG neurons associated with changes in gene expression and gene regions enriched with CTCF binding motifs (Palmisano et al, 2019) possibility also influencing epigenetic modifications. DNMT3a is a strong candidate for mediating increased levels of methylation of Cacna1b e37a locus in Trpv1 -lineage neurons.…”

Section: Discussionmentioning

confidence: 99%

Cell-specific exon methylation and CTCF binding in neurons regulates calcium ion channel splicing and function

Soto

Lipscombe

2019

Preprint

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Cell-specific alternative splicing modulates myriad cell functions and this process is disrupted in disease. The mechanisms governing alternative splicing are known for relatively few genes and typically focus on RNA splicing factors. In sensory neurons, cell-specific alternative splicing of the presynaptic voltage-gated calcium channel Cacna1b gene modulates opioid sensitivity. How this splicing is regulated has remained unknown. We find that cell-specific exon DNA hypomethylation permits binding of CTCF, the master regulator of chromatin structure in mammals, which, in turn, controls splicing in noxious heat-sensing nociceptors.Hypomethylation of an alternative exon specifically in nociceptors allows for CTCF binding, and expression of CaV2.2 channels with increased opioid sensitivity. Following nerve injury, exon methylation is increased, and splicing is disrupted. Our studies define the molecular mechanisms of cell-specific alternative splicing of a functionally validated exon in normal and disease statesand reveal a potential target for the treatment of chronic pain.Highlights (each bullet not > 85 characters including spaces)• The molecular basis of cell-specific splicing of a synaptic calcium channel gene.• Splicing controlled by cell-specific exon hypomethylation and CTCF binding.• Peripheral nerve injury disrupts exon hypomethylation and splicing.• Targeted demethylation of exon by dCAS9-TET modifies alternative splicing.

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Epigenomic signatures underpin the axonal regenerative ability of dorsal root ganglia sensory neurons

Cited by 82 publications

References 50 publications

Co-occupancy analysis reveals novel transcriptional synergies for axon growth

Co-occupancy analysis reveals novel transcriptional synergies for axon growth

N6-methyladenine DNA demethylase ALKBH1 regulates mammalian axon regeneration

Cell-specific exon methylation and CTCF binding in neurons regulates calcium ion channel splicing and function

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