Nociceptor Translational Profiling Reveals the Ragulator-Rag GTPase Complex as a Critical Generator of Neuropathic Pain

Megat, Salim; Ray, Pradipta; Moy, Jamie K.; Lou, Tzu-Fang; Barragán-Iglesias, Paulino; Li, Yan; Pradhan, Grishma; Wanghzou, Andi; Ahmad, Ayesha; Burton, Michael D.; North, Robert Y.; Dougherty, Patrick M.; Khoutorsky, Arkady; Sonenberg, Nahum; Webster, Kevin R.; Dussor, Gregory; Campbell, Zachary T.; Price, Theodore J.

doi:10.1523/jneurosci.2661-18.2018

Cited by 111 publications

(143 citation statements)

References 78 publications

(45 reference statements)

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“…Activation of MNK-eIF4E signaling by type I IFNs has been observed in other cell types where it has been linked to increased immune surveillance (Joshi et al, 2009). Multiple previous studies have demonstrated that activation of MNK-eIF4E-mediated translation events are causative in the production of chronic pain states, including neuropathic pain (Moy et al, 2017; Megat et al, 2019; Shiers et al, 2019). Since both viral infections and prolonged production of type I IFNs can cause neuropathic pain, it is possible that type I IFN receptor signaling to MNK-eIF4E may be a key pathway for production of these types of neuropathies.…”

Section: Discussionmentioning

confidence: 99%

“…A key pathway linking initial nociceptor activation to nociceptor hypersensitivity and potentially the development of chronic pain is engagement of translation regulation signaling (Obara and Hunt, 2014; Khoutorsky and Price, 2018; de la Pena et al, 2019). Importantly, eIF2α phosphorylation, mTORC1 activation and MNK-eIF4E signaling can all lead to persistent sensitization of nociceptors and all of these pathways have been linked to neuropathic pain disorders (Inceoglu et al, 2015; Khoutorsky et al, 2016; Moy et al, 2017; Megat et al, 2019; Shiers et al, 2019)…”

Section: Introductionmentioning

confidence: 99%

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Type I interferons act directly on nociceptors to produce pain sensitization: Implications for viral infection-induced pain

Barragán-Iglesias¹,

Franco-Enzástiga

Jeevakumar³

et al. 2019

Preprint

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ABSTRACTOne of the first signs of viral infection is body-wide aches and pain. While this type of pain usually subsides, at the extreme, viral infections can induce painful neuropathies that can last for decades. Neither of these types of pain sensitization are well understood. A key part of the response to viral infection is production of interferons (IFNs), which then activate their specific receptors (IFNRs) resulting in downstream activation of cellular signaling and a variety of physiological responses. We sought to understand how type I IFNs (IFN-α and IFN-β) might act directly on nociceptors in the dorsal root ganglion (DRG) to cause pain sensitization. We demonstrate that type I IFNRs are expressed in small/medium DRG neurons and that their activation produces neuronal hyper-excitability and mechanical pain in mice. Type I IFNs stimulate JAK/STAT signaling in DRG neurons but this does not apparently result in PKR-eIF2α activation that normally induces an anti-viral response by limiting mRNA translation. Rather, type I interferons stimulate MNK-mediated eIF4E phosphorylation in DRG neurons to promote pain hypersensitivity. Endogenous release of type I IFNs with the double stranded RNA mimetic poly(I:C) likewise produces pain hypersensitivity that is blunted in mice lacking MNK-eIF4E signaling. Our findings reveal mechanisms through which type I IFNs cause nociceptor sensitization with implications for understanding how viral infections promote pain and can lead to neuropathies.SIGNIFICANCE STATEMENTIt is increasingly understood that pathogens interact with nociceptors to alert organisms to infection as well as to mount early host defenses. While specific mechanisms have been discovered for diverse bacteria and fungal pathogens, mechanisms engaged by viruses have remained elusive. Here we show that type 1 interferons, one of the first mediators produced by viral infection, act directly on nociceptors to produce pain sensitization. Type I interferons act via a specific signaling pathway (MNK-eIF4E signaling) that is known to produce nociceptor sensitization in inflammatory and neuropathic pain conditions. Our work reveals a mechanism through which viral infections cause heightened pain sensitivity

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Type I interferons act directly on nociceptors to produce pain sensitization: Implications for viral infection-induced pain

Barragán-Iglesias¹,

Franco-Enzástiga

Jeevakumar³

et al. 2019

Preprint

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“…We have focused on using DRGs from uninjured mice and from human organ donors without a history of chronic pain. Many studies have demonstrated that cultured DRG neurons from mice with neuropathic pain retain some neuropathic qualities in vitro , in particular spontaneous activity in a sub-population of nociceptors [2; 33; 39; 63; 65]. This also occurs in human DRG neurons taken from people with neuropathic pain [39].…”

Section: Discussionmentioning

confidence: 99%

“…Some transcriptomic changes were found in mice but not conserved in humans. An example is caspase 6 ( Casp6 ) which has been implicated in many neuropathic pain models in mice [4; 5; 33]. This gene was over 3 fold increased in mouse DRG culture but unchanged in human DRG cultures.…”

Section: Discussionmentioning

confidence: 99%

Transcriptomic analysis of native versus cultured human and mouse dorsal root ganglia focused on pharmacological targets

Wangzhou

McIlvried²,

Paige

et al. 2019

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words)Dorsal root ganglion (DRG) neurons detect sensory inputs and are crucial for pain processing.They are often studied in vitro as dissociated cell cultures with the assumption that this reasonably represents in vivo conditions. However, to our knowledge, no study has ever directly compared genome-wide transcriptomes of DRG tissue in vivo versus in vitro, or between different labs and culturing protocols. We extracted bilateral lumbar DRG from C57BL6/J mice and human organ donors, and acutely froze one side and processed the other side as a dissociated cell culture, which was then maintained in vitro for 4 days. RNA was extracted and sequenced using the NextSeq Illumina platform. Comparing native to cultured human or mouse DRG, we found that the overall expression level of many ion channels and GPCRs specifically expressed in neurons is markedly lower in culture, but still expressed. This suggests that most pharmacological targets expressed in vivo are present in culture conditions. However, there are changes in expression levels for these genes. The reduced relative expression for neuronal genes in human DRG cultures is likely accounted for by increased expression of genes in fibroblast-like and other proliferating cells, consistent with the mitotic status of many cells in these cultures. We did find a subset of genes that are typically neuronally expressed, increased in human and mouse DRG cultures, including genes associated with nerve injury and/or inflammation in preclinical models such as BDNF, MMP9, GAL, and ATF3. We also found a striking upregulation of a number of inflammation-associated genes in DRG cultures, although many were different between mouse and human. Our findings suggest an injury-like phenotype in DRG cultures that has important implications for the use of this model system for pain drug discovery. Methods Experimental DesignBecause genetic variation can be a possible contributor to transcriptome level differences in nervous system samples from human populations [39; 41], we chose a study design wherein we cultured lumbar DRGs from one side in human donors and immediately froze the opposite side from the same donor for RNA sequencing. Although we used an inbred mouse strain (C57BL/6) for parallel mouse studies, we used a similar culturing design where cultures were done in two independent laboratories to look for variability across labs. RNA sequencing was performed at 4 days in vitro (DIV) to stay within the electrophysiologically relevant range of 1 -7 DIV for human DRG and the biochemical assay range of 4 -7 DIV for both human and mouse DRG. AnimalsPrice Lab: All procedures were approved by the Institutional Animal Care and Use Committee of University of Texas at Dallas and were in strict accordance with the US National Institute of Health (NIH) Guide for the Care and Use of Laboratory Animals. Adult C57Bl/6 mice (8-15 weeks of age) were bred in house, and were originally obtained from The Jackson Laboratory. Animals were housed in the University of Texas at Dallas animal facilities o...

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“…The TRAP approach therefore represents a complimentary approach to bulk tissue and single cell sequencing capable to reveal novel insights into tissue diversity and pathological changes. In fact, two recent studies have employed the TRAP approach to primary nociceptors and uncovered unknown differences between trigeminal and dorsal root ganglia as well as unidentified driver genes for neuropathic pain [40,41].…”

Section: Single Cell Transcriptome Profiling Versus Cell Type Specifimentioning

confidence: 99%

Neuron Type-Specific Translatomes in Spinal Cords of Naïve and Neuropathic Mice

Gupta

Scheurer

Pelczar

et al. 2020

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The spinal dorsal horn harbors a sophisticated and heterogeneous network of excitatory and inhibitory neurons that process peripheral signals encoding different sensory modalities. Although it has long been recognized that this network is crucial both for the separation and the integration of sensory signals of different modalities, the molecular identity of the underlying neurons and signaling mechanisms are still only partially understood. Here, we have used the translating ribosome affinity purification (TRAP) technique to map the translatomes of excitatory glutamatergic (VGLUT2 + ) and inhibitory GABA and/or glycinergic (VGAT + or Gad67 + ) neurons of the mouse spinal cord. Our analyses demonstrate that inhibitory and excitatory neurons are primarily set apart by the expression of genes encoding transcription factors or genes related to the production, release or re-uptake of their principal neurotransmitters (glutamate, GABA or glycine). Subsequent gene ontology (GO) term analyses revealed that neuropeptide signaling-related GO terms were highly enriched in the excitatory population. Eleven neuropeptide genes displayed largely non-overlapping expression patterns closely adhering to the laminar and hence also functional organization of the spinal cord grey matter, suggesting that they may serve as major determinants of modality-specific processing. Since this modality-specific processing of sensory input is severely compromised in chronic, especially neuropathic, pain, we also investigated whether peripheral nerve damage changes the neuron typespecific translatome. In summary, our results suggest that neuropeptides contribute to modalityspecific sensory processing in the spinal cord but also indicate that altered sensory encoding in neuropathic pain states occurs independent of major translatome changes in the spinal neurons.The ability to sense and discriminate different noxious and innocuous somatosensory stimuli is essential for all higher animals and humans in order to react adequately to external stimuli and internal conditions [1,2]. The spinal dorsal horn, i.e., the sensory part of the spinal cord, constitutes a key element in this process. It receives somatosensory signals from peripheral neurons and processes these signals together with other inputs descending from supraspinal sites in a complex network of inhibitory and excitatory interneurons before relaying these signals via projection neurons to supraspinal centers [3]. Projection neurons make up less than 10% of all dorsal horn neurons, while more than 90% of the neuronal population are interneurons of which, between 60 and 70% are excitatory glutamatergic neurons, and the rest is inhibitory (GABA and/or glycinergic).The spinal cord is organized in a laminar fashion, which has initially been proposed on the basis of differences in cell density and morphology between the different laminae [4, 5] but has later been shown to also reflect functional organization. This is especially reflected for example by the lamina specific innervation pattern by ...

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Nociceptor Translational Profiling Reveals the Ragulator-Rag GTPase Complex as a Critical Generator of Neuropathic Pain

Cited by 111 publications

References 78 publications

Type I interferons act directly on nociceptors to produce pain sensitization: Implications for viral infection-induced pain

Type I interferons act directly on nociceptors to produce pain sensitization: Implications for viral infection-induced pain

Transcriptomic analysis of native versus cultured human and mouse dorsal root ganglia focused on pharmacological targets

Neuron Type-Specific Translatomes in Spinal Cords of Naïve and Neuropathic Mice

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