Gene Expression In Sensory Neurons Research Articles

Social experience and pheromone receptor activity reprogram gene expression in sensory neurons.

Social experience and pheromone signaling in olfactory neurons affect neuronal responses and male courtship behaviors in Drosophila. We previously showed that social experience and pheromone signaling modulate chromatin around behavioral switch gene fruitless, which encodes a transcription factor necessary and sufficient for male sexual behaviors. Fruitless drives social experience-dependent modulation of courtship behaviors and physiological sensory neuron responses to pheromone; however, the molecular mechanisms underlying this modulation of neural responses remain less clear. To identify the molecular mechanisms driving social experience-dependent changes in neuronal responses, we performed RNA-seq from antennal samples of mutants in pheromone receptors and fruitless, as well as grouped or isolated wild-type males. Genes affecting neuronal physiology and function, such as neurotransmitter receptors, ion channels, ion and membrane transporters, and odorant binding proteins are differentially regulated by social context and pheromone signaling. While we found that loss of pheromone detection only has small effects on differential promoter and exon usage within fruitless gene, many of the differentially regulated genes have Fruitless-binding sites or are bound by Fruitless in the nervous system. Recent studies showed that social experience and juvenile hormone signaling co-regulate fruitless chromatin to modify pheromone responses in olfactory neurons. Interestingly, genes involved in juvenile hormone metabolism are also misregulated in different social contexts and mutant backgrounds. Our results suggest that modulation of neuronal activity and behaviors in response to social experience and pheromone signaling likely arise due to large-scale changes in transcriptional programs for neuronal function downstream of behavioral switch gene function.

Painful intercellular connections

Pain Chronic pain impairs quality of life and is challenging to treat. Wangzhou et al. measured ligand and receptor gene expression in sensory neurons and various peripheral cell types from humans and mice. Using these data, the authors constructed a connectome of potential cell type–specific interaction points between sensory nerves and innervated tissues, which suggested that blocking heparin-binding epidermal growth factor (HBEGF) signaling may be an effective way to alleviate chronic pain in multiple diseases. This target was validated in a mouse model of mechanical pain. Sci. Signal. 14 , abe1648 (2021).

The Role of DNA Methylation in Transcriptional Regulation of Pro-Nociceptive Genes in Rat Trigeminal Ganglia.

Epigenetic modulation by DNA methylation is associated with aberrant gene expression in sensory neurons, which consequently leads to pathological pain responses. In this study, we sought to investigate whether peripheral inflammation alters global DNA methylation in trigeminal ganglia (TG) and results in abnormal expression of pro-nociceptive genes. Our results show that peripheral inflammation remotely reduced the level of global DNA methylation in rat TG with a concurrent reduction in DNMT1 and DNMT3a expression. Using unbiased steps, we selected the following pro-nociceptive candidate genes that are potentially regulated by DNA methylation: TRPV1, TRPA1, P2X3, and PIEZO2. Inhibition of DNMT with 5-Aza-dC in dissociated TG cells produced dose-dependent upregulation of TRPV1, TRPA1, and P2X3. Systemic treatment of animals with 5-Aza-dC significantly increased the expression of TRPV1, TRPA1, and PIEZO2 in TG. Furthermore, the overexpression of DNMT3a, as delivered by a lentiviral vector, significantly downregulated TRPV1 and PIEZO2 expression and also reliably decreased TRPA1 and P2X3 transcripts. MeDIP revealed that this overexpression also significantly enhanced methylation of CGIs associated with TRPV1 and TRPA1. In addition, bisulfite sequencing data indicated that the CGI associated with TRPA1 was methylated in a pattern catalyzed by DNMT3a. Taken together, our results show that all 4 pro-nociceptive genes are subject to epigenetic modulation via DNA methylation, likely via DNMT3a under inflammatory conditions. These findings provide the first evidence for the functional importance of DNA methylation as an epigenetic factor in the transcription of pro-nociceptive genes in TG that are implicated in pathological orofacial pain responses.

Combinatorial Effects of the Glucocorticoid Receptor and Krüppel-Like Transcription Factor 15 on Bovine Herpesvirus 1 Transcription and Productive Infection.

Bovine herpesvirus 1 (BoHV-1), an important bovine pathogen, establishes lifelong latency in sensory neurons. Latently infected calves consistently reactivate from latency following a single intravenous injection of the synthetic corticosteroid dexamethasone. The immediate early transcription unit 1 (IEtu1) promoter, which drives bovine ICP0 (bICP0) and bICP4 expression, is stimulated by dexamethasone because it contains two glucocorticoid receptor (GR) response elements (GREs). Several Krüppel-like transcription factors (KLF), including KLF15, are induced during reactivation from latency, and they stimulate certain viral promoters and productive infection. In this study, we demonstrate that the GR and KLF15 were frequently expressed in the same trigeminal ganglion (TG) neuron during reactivation and cooperatively stimulated productive infection and IEtu1 GREs in mouse neuroblastoma cells (Neuro-2A). We further hypothesized that additional regions in the BoHV-1 genome are transactivated by the GR or stress-induced transcription factors. To test this hypothesis, BoHV-1 DNA fragments (less than 400 bp) containing potential GR and KLF binding sites were identified and examined for transcriptional activation by stress-induced transcription factors. Intergenic regions within the unique long 52 gene (UL52; a component of the DNA primase/helicase complex), bICP4, IEtu2, and the unique short region were stimulated by KLF15 and the GR. Chromatin immunoprecipitation studies revealed that the GR and KLF15 interacted with sequences within IEtu1 GREs and the UL52 fragment. Coimmunoprecipitation studies demonstrated that KLF15 and the GR were associated with each other in transfected cells. Since the GR stimulates KLF15 expression, we suggest that these two transcription factors form a feed-forward loop that stimulates viral gene expression and productive infection following stressful stimuli.IMPORTANCE Bovine herpesvirus 1 (BoHV-1) is an important viral pathogen that causes respiratory disease and suppresses immune responses in cattle; consequently, life-threatening bacterial pneumonia can occur. Following acute infection, BoHV-1 establishes lifelong latency in sensory neurons. Reactivation from latency is initiated by the synthetic corticosteroid dexamethasone. Dexamethasone stimulates lytic cycle viral gene expression in sensory neurons of calves latently infected with BoHV-1, culminating in virus shedding and transmission. Two stress-induced cellular transcription factors, Krüppel-like transcription factor 15 (KLF15) and the glucocorticoid receptor (GR), cooperate to stimulate productive infection and viral transcription. Additional studies demonstrated that KLF15 and the GR form a stable complex and that these stress-induced transcription factors bind to viral DNA sequences, which correlates with transcriptional activation. The ability of the GR and KLF15 to synergistically stimulate viral gene expression and productive infection may be critical for the ability of BoHV-1 to reactivate from latency following stressful stimuli.

Sexually dimorphic control of gene expression in sensory neurons regulates decision-making behavior in C. elegans

Animal behavior is directed by the integration of sensory information from internal states and the environment. Neuroendocrine regulation of diverse behaviors of Caenorhabditis elegans is under the control of the DAF-7/TGF-β ligand that is secreted from sensory neurons. Here, we show that C. elegans males exhibit an altered, male-specific expression pattern of daf-7 in the ASJ sensory neuron pair with the onset of reproductive maturity, which functions to promote male-specific mate-searching behavior. Molecular genetic analysis of the switch-like regulation of daf-7 expression in the ASJ neuron pair reveals a hierarchy of regulation among multiple inputs-sex, age, nutritional status, and microbial environment-which function in the modulation of behavior. Our results suggest that regulation of gene expression in sensory neurons can function in the integration of a wide array of sensory information and facilitate decision-making behaviors in C. elegans.

Epigenetic regulation of chronic pain.

Chronic pain arising from peripheral inflammation and tissue or nerve injury is a common clinical symptom. Although intensive research on the neurobiological mechanisms of chronic pain has been carried out during previous decades, this disorder is still poorly managed by current drugs such as opioids and nonsteroidal anti-inflammatory drugs. Inflammation, tissue injury and/or nerve injury-induced changes in gene expression in sensory neurons of the dorsal root ganglion, spinal cord dorsal horn and pain-associated brain regions are thought to participate in chronic pain genesis; however, how these changes occur is still elusive. Epigenetic modifications including DNA methylation and covalent histone modifications control gene expression. Recent studies have shown that peripheral noxious stimulation changes DNA methylation and histone modifications and that these changes may be related to the induction of pain hypersensitivity under chronic pain conditions. This review summarizes the current knowledge and progress in epigenetic research in chronic pain and discusses the potential role of epigenetic modifications as therapeutic antinociceptive targets in this disorder.

The Ion Channel TRPA1 Is Required for Chronic Itch

Chronic itch is a debilitating condition that affects one in 10 people. Little is known about the molecules that mediate chronic itch in primary sensory neurons and skin. We demonstrate that the ion channel TRPA1 is required for chronic itch. Using a mouse model of chronic itch, we show that scratching evoked by impaired skin barrier is abolished in TRPA1-deficient animals. This model recapitulates many of the pathophysiological hallmarks of chronic itch that are observed in prevalent human diseases such as atopic dermatitis and psoriasis, including robust scratching, extensive epidermal hyperplasia, and dramatic changes in gene expression in sensory neurons and skin. Remarkably, TRPA1 is required for both transduction of chronic itch signals to the CNS and for the dramatic skin changes triggered by dry-skin-evoked itch and scratching. These data suggest that TRPA1 regulates both itch transduction and pathophysiological changes in the skin that promote chronic itch.

LAT Region Factors Mediating Differential Neuronal Tropism of HSV-1 and HSV-2 Do Not Act in Trans

After HSV infection, some trigeminal ganglion neurons support productive cycle gene expression, while in other neurons the virus establishes a latent infection. We previously demonstrated that HSV-1 and HSV-2 preferentially establish latent infection in A5+ and KH10+ sensory neurons, respectively, and that exchanging the latency-associated transcript (LAT) between HSV-1 and HSV-2 also exchanges the neuronal preference. Since many viral genes besides the LAT are functionally interchangeable between HSV-1 and HSV-2, we co-infected HSV-1 and HSV-2, both in vivo and in vitro, to determine if trans-acting viral factors regulate whether HSV infection follows a productive or latent pattern of gene expression in sensory neurons. The pattern of HSV-1 and HSV-2 latent infection in trigeminal neurons was no different following co-infection than with either virus alone, consistent with the hypothesis that a trans-acting viral factor is not responsible for the different patterns of latent infection of HSV-1 and HSV-2 in A5+ and KH10+ neurons. Since exchanging the LAT regions between the viruses also exchanges neuronal preferences, we infected transgenic mice that constitutively express 2.8 kb of the LAT region with the heterologous viral serotype. Endogenous expression of LAT did not alter the pattern of latent infection after inoculation with the heterologous serotype virus, demonstrating that the LAT region does not act in trans to direct preferential establishment of latency of HSV-1 and HSV-2. Using HSV1-RFP and HSV2-GFP in adult trigeminal ganglion neurons in vitro, we determined that HSV-1 and HSV-2 do not exert trans-acting effects during acute infection to regulate neuron specificity. Although some neurons were productively infected with both HSV-1 and HSV-2, no A5+ or KH10+ neurons were productively infected with both viruses. Thus, trans-acting viral factors do not regulate preferential permissiveness of A5+ and KH10+ neurons for productive HSV infection and preferential establishment of latent infection.

Retrograde BMP Signaling Regulates Trigeminal Sensory Neuron Identities and the Formation of Precise Face Maps

Somatosensory information from the face is transmitted to the brain by trigeminal sensory neurons. It was previously unknown whether neurons innervating distinct areas of the face possess molecular differences. We have identified a set of genes differentially expressed along the dorsoventral axis of the embryonic mouse trigeminal ganglion and thus can be considered trigeminal positional identity markers. Interestingly, establishing some of the spatial patterns requires signals from the developing face. We identified bone morphogenetic protein 4 (BMP4) as one of these target-derived factors and showed that spatially defined retrograde BMP signaling controls the differential gene expressions in trigeminal neurons through both Smad4-independent and Smad4-dependent pathways. Mice lacking one of the BMP4-regulated transcription factors, Onecut2 (OC2), have defects in the trigeminal central projections representing the whiskers. Our results provide molecular evidence for both spatial patterning and retrograde regulation of gene expression in sensory neurons during the development of the somatosensory map.

NGF Up-regulates TRPA1: Implications for Orofacial Pain

The transient receptor potential ankyrin repeat 1 (TRPA1) channel is believed to be involved in many forms of acute and chronic hyperalgesia. Nerve Growth Factor (NGF) regulates chronic inflammatory hyperalgesia by controlling gene expression in sensory neurons, including genes involved in inflammatory hyperalgesia in the dental pulp. We hypothesized that NGF increases functional activities of the TRPA1 channel in trigeminal ganglion neurons. Here, we show that NGF induced a concentration- and time-dependent up-regulation of TRPA1 mRNA in trigeminal ganglia neurons, as detected by real-time RT-PCR and in situ hybridization. In addition, NGF evoked a time-dependent increase of mustard oil (MO)-evoked TRPA1 activation in trigeminal ganglia neurons. Collectively, these findings demonstrate that NGF participates in the functional up-regulation of TRPA1 in trigeminal ganglia neurons. These enhanced activities of TRPA1 could play an important role in the development of hyperalgesia following nerve injury and inflammation in the orofacial region.

Groucho/transducin-like Enhancer-of-split (TLE)-dependent and -independent transcriptional regulation by Runx3

Regulation of gene expression by tissue-specific transcription factors involves both turning on and turning off transcription of target genes. Runx3, a runt-domain transcription factor, regulates cell-intrinsic functions by activating and repressing gene expression in sensory neurons, dendritic cells (DC), and T cells. To investigate the mechanism of Runx3-mediated repression in an in vivo context, we generated mice expressing a mutant Runx3 lacking the C-terminal VWRPY, a motif required for Runx3 interaction with the corepressor Groucho/transducin-like Enhancer-of-split (TLE). In contrast with Runx3(-/-) mice, which displayed ataxia due to the death of dorsal root ganglia TrkC neurons, Runx3(VWRPY-/-) mice were not ataxic and had intact dorsal root ganglia neurons, indicating that ability of Runx3 to tether Groucho/TLE is not essential for neurogenesis. In the DC compartment, the mutant protein Runx3(VWRPY-) promoted normally developed skin Langerhans cells but failed to restrain DC spontaneous maturation, indicating that this latter process involves Runx3-mediated repression through recruitment of Groucho/TLE. Moreover, in CD8(+) thymocytes, Runx3(VWRPY-) up-regulated alphaE/CD103-like WT Runx3, whereas unlike wild type, it failed to repress alphaE/CD103 in CD8(+) splenocytes. Thus, in CD8-lineage T cells, Runx3 regulates alphaE/CD103 in opposing regulatory modes and recruits Groucho/TLE to facilitate the transition from activation to repression. Runx3(VWRPY-) also failed to mediate the epigenetic silencing of CD4 gene in CD8(+) T cells, but normally regulated other pan-CD8(+) T cell genes. These data provide evidence for the requirement of Groucho/TLE for Runx3-mediated epigenetic silencing of CD4 and pertain to the mechanism through which other Runx3-regulated genes are epigenetically silenced.

Comparative functional genomics revealed conservation and diversification of three enhancers of the isl1 gene for motor and sensory neuron-specific expression

Islet-1 (Isl1) is a member of the Isl1 family of LIM-homeodomain transcription factors (LIM-HD) that is expressed in a defined subset of motor and sensory neurons during vertebrate embryogenesis. To investigate how this specific expression of isl1 is regulated, we searched for enhancers of the isl1 gene that are conserved in vertebrate evolution. Initially, two enhancer elements, CREST1 and CREST2, were identified downstream of the isl1 locus in the genomes of fugu, chick, mouse, and human by BLAST searching for highly similar elements to those originally identified as motor and sensory neuron-specific enhancers in the zebrafish genome. The combined action of these elements is sufficient for completely recapitulating the subtype-specific expression of the isl1 gene in motor neurons of the mouse spinal cord. Furthermore, by direct comparison of the upstream flanking regions of the zebrafish and human isl1 genes, we identified another highly conserved noncoding element, CREST3, and subsequently C3R, a similar element to CREST3 with two CDP CR1 recognition motifs, in the upstream regions of all other isl1 family members. In mouse and human, CRESTs are located as far as more than 300 kb away from the isl1 locus, while they are much closer to the isl1 locus in zebrafish. Although all of zebrafish CREST2, CREST3, and C3R activate gene expression in the sensory neurons of zebrafish, CREST2 of mouse and human does not have the sequence necessary for sensory neuron-specific expression. Our results revealed both a remarkable conservation of the regulatory elements regulating subtype-specific gene expression in motor and sensory neurons and the dynamic process of reorganization of these elements whereby each element increases the level of cell-type specificity by losing redundant functions with the other elements during vertebrate evolution.

A novel replication-defective HSV-1 vector for regulatable gene delivery to wounds

Abstract Introduction: Gene delivery of growth factors to wounds can accelerate healing but has so far not been achieved with any control over the level of transgene expression. Herpes viral vectors are efficient vehicles for in vivo gene delivery and have the advantage of being able to establish long-term gene expression in sensory neurons that are known to interact with the healing wound. To increase safety and control of in vivo gene-transfer we constructed a novel replication-deficient viral vector that for the first time allows efficient regulation of transgene expression in a dose-dependent manner by a HSV-1 recombinant system. Methods: QR9TOhEGF is a novel replication-defective virus that encodes hEGF under control of the tetO-containing CMV promoter (T-REx, Invitrogen) and the tetracycline repressor protein tetR. Without tetracycline, tetR binds to tetO, leading to effective repression of hEGF expression, while in the presence of tetracycline this repression is released. Vero cells were infected with QR9TOhEGF in the presence and absence of tetracycline. Porcine full-thickness wounds sealed with a polyvinyl chamber were infected with QR9TOhEGF in the presence and absence of tetracycline. Wound fluid was assayed at 24 and 48h using hEGF-ELISA. Results: hEGF concentration (pg/ml) at 24 and 48hrs following infection shows up to at least 400-fold regulation. Sensitivity limit: 3pg/ml Vero cellsT−T+Fold regulationp Value (unpaired t-test)24 h613602260.009648 h3.514004000.0003In vivo2.06 × 10 7 PFU/wound24 h841120.028148 h6365.80.0241EGF-ELISA of conditioned medium (Vero cells) and wound fluid (In vivo) at 24 and 48 hours, in the presence and absence of tetracycline. Lower limit of sensitivity: 3pg/ml. Conclusions: We have constructed an HSV-1 vector that allows the highest degree of regulation of gene expression in a single vector system. The feasibility of regulatable gene transfer to wounds has been demonstrated. Our method of in vivo delivery is currently being optimized to achieve higher expression.

Coupling gene chip analyses and rat genetic variances in identifying potential target genes that may contribute to neuropathic allodynia development.

Genetic factors and nerve injury-induced changes of gene expression in sensory neurons are potential contributors to tactile allodynia, a neuropathic pain state manifested as hypersensitivity to innocuous mechanical stimulation. To uncover genes relevant to neuropathic allodynia, we analyzed gene expression profiles in dorsal root ganglia (DRG) of spinal nerve-ligated Harlan and Holtzman Sprague Dawley rats, strains with different susceptibilities to neuropathic allodynia. Using Affymetrix gene chips, we identified genes showing differential basal-level expression in these strains without injury-induced regulation. Of more than 8000 genes analyzed, less than 180 genes in each strain were regulated after injury, and 19-22% of that was regulated in a strain-specific manner. Importantly, we identified functionally related genes that were co-regulated post injury in one or both strains. In situ hybridization and real-time PCR analyses of a subset of identified genes confirmed the patterns of the microarray data, and the former also demonstrated that injury-induced changes occurred, not only in neurons, but also in non-neuronal cells. Together, our studies provide a global view of injury plasticity in DRG of these rat stains and support a plasticity-based mechanism mediating variations in allodynia susceptibility, thus providing a source for further characterization of neuropathic pain-relevant genes and potential pathways.

Changes in gene expression in sensory neurons in acute pancreatitis

Changes in gene expression in sensory neurons in acute pancreatitis

Interferon gamma gene expression in sensory neurons: evidence for autocrine gene regulation.

We explored expression and possible function of interferon-gamma (IFN-gamma) in cultured fetal (E15) rat dorsal root ganglion neurons combining whole cell patch-clamp electrophysiology with single cell reverse transcriptase polymerase chain reaction and confocal laser immunocytochemistry. Morphologically, we located IFN-gamma protein in the cytoplasm of the neurons in culture as well as in situ during peri- and postnatal development. Transcripts for classic IFN-gamma and for its receptor were determined in probes of cytoplasm sampled from individual cultured neurons, which had been identified by patch clamp electrophysiology. In addition, the cultured neurons expressed both chains of the IFN-gamma receptor. Locally produced IFN-gamma acts back on its cellular source. Phosphorylation and nuclear translocation of the IFN-inducible transcriptional factor STAT1 as well as IFN-gamma-dependent expression of major histocompatibility complex class I molecules on the neuronal membrane were noted in untreated cultures. However, both processes were substantially blocked in the presence of antibodies neutralizing IFN-gamma. Our findings indicate a role of IFN-gamma in autocrine regulation of sensory neurons.

Transcriptional control of neuropeptide gene expression in sensory neurons, using the preprotachykinin-A gene as a model.

Control of neuropeptide gene expression in sensory neurons is determined in part by a variety of tissue-specific, developmental, and stimulus-induced transcription factors that interact with the promoters of these genes. We have analysed the regulation of the rat preprotachykinin-A (rPPT) gene, which is expressed in a subset of dorsal root ganglia neurons. A region of the promoter encompassing approximately 1300 base pairs spanning the transcriptional start site has been analysed in detail both by functional analysis of promoter activity in clonal cell lines and dorsal root ganglia neurons grown in culture and by in vitro characterisation of transcription factor interaction with this region. Interestingly our analysis indicates an important role in rPPT gene expression for the E box transcription factor family. This class of transcription factor has been demonstrated to be a major determinant of calcitonin gene related peptide (CGRP) expression, which is also expressed in dorsal root ganglia neurons often under similar conditions as rPPT. In addition, multiple regulatory domains have been identified in the rPPT promoter, which act as activators in a variety of cell types. These elements are silenced in the context of the rPPT promoter in many non-neuronal cells. Therefore, tissue-specific expression of reporter genes directed by the rPPT promoter in transient transfection is determined in part by a variety of silencer elements, which act to repress the function of several domains that act as constitutive enhancers of expression in a wide range of cells. Removal or modulation of silencer elements in the rPPT promoter allows activity in a wider variety of cell types. We postulate that control of rPPT gene expression is the results of dynamic interplay of both positive and negative regulatory elements, a phenomenon observed in several other neuronal-specific genes, including that encoding CGRP.

Deficits in sciatic nerve neuropeptide content coincide with a reduction in target tissue nerve growth factor messenger RNA in streptozotocin-diabetic rats: Effects of insulin treatment

The role of sub-optimal neurotrophic support in the aetiology of the sensory neuron dysfunction associated with diabetic neuropathy was investigated. The status of sciatic nerve neuropeptide content was related to target tissue nerve growth factor messenger RNA levels in streptozotocin-diabetic rats. The levels of substance P and calcitonin gene-related peptide in diabetic sciatic nerve were significantly lowered by approximately 50% and 28%, respectively, compared with aged matched controls and insulin-treated diabetic rats ( P < 0.01) for both peptides and both comparisons). Measurements of nerve growth factor messenger RNA levels in sensory neuron target tissues, namely foot-skin and soleus muscle, revealed deficits of approximately 50% in diabetic rats, with insulin treatment reversing the decrease in foot-skin but not in soleus muscle. The results show a possible correlation between deficient neuropeptide gene expression in sensory neurons and reduced nerve growth factor messenger RNA levels in target tissue.