Reciprocal modulation of TrkA and p75NTR affinity states is mediated by direct receptor interactions

Equilibrium binding of ¹²⁵I-nerve growth factor (¹²⁵I-NGF) to cells coexpressing the tyrosine kinase receptor A (TrkA) and common neurotrophin receptor (p75^NTR), cells coexpressing both receptors where p75^NTR is occupied, and cells expressing only p75^NTR, revealed reciprocal modulation of receptor affinity states. Analysis of receptor affinity states in PC12 cells, PC12 cells in the presence of brain-derived neurotrophic factor (BDNF), and PC12^nnr5 cells suggested that liganded and unliganded p75^NTR induce a higher affinity state within TrkA, while TrkA induces a lower affinity state within p75^NTR. These data are consistent with receptor allosterism, and prompted a search for TrkA/p75^NTR complexes in the absence of NGF. Chemical crosslinking studies revealed high molecular weight receptor complexes that specifically bound ¹²⁵I-NGF, and were immunoprecipitated by antibodies to both receptors. The heteroreceptor complex of TrkA and p75^NTR alters conformation and/or dissociates in the presence of NGF, as indicated by the ability of low concentrations of NGF to prevent heteroreceptor crosslinking. These data suggest a new model of receptor interaction, whereby structural changes within a heteroreceptor complex are induced by ligand binding.     

Jekyll-Hyde neurotrophins: the story of proNGF

Neurotrophins promote neuronal survival and differentiation by binding to two classes of cell surface receptors: members of the Trk-family receptor tyrosine kinases and p75(NTR), a member of the tumor necrosis factor receptor superfamily. The discoveries that the unprocessed proneurotrophin peptide is both a preferential high-affinity ligand for p75(NTR) (with minimal affinity for trkA) and a potent inducer of p75(NTR)-mediated neuronal cell death are likely to change our understanding of neurotrophin physiology and our ability to manipulate their signaling pathways.     

3,4-methylenedioxyamphetamine upregulates p75 neurotrophin receptor protein expression in the rat brain

The p75 neurotrophin receptor,which is a member of the tumor necrosis factor receptor superfamily,facilitates apoptosis during development and following central nervous system injury.Previous studies have shown that programmed cell death is likely involved in the neurotoxic effects of 3,4-methylenedioxy-N-methylamphetamine (MDMA),because MDMA induces apoptosis of immortalized neurons through regulation of proteins belonging to the Bcl-2 family.In the present study,intraperitoneal injection of different doses of MDMA (20,50,and 100 mg/kg) induced significant behavioral changes,such as increased excitability,increased activity,and irritability in rats.Moreover,changes exhibited dose-dependent adaptation.Following MDMA injection in rat brain tissue,the number of apoptotic cells dose-dependently increased and p75 neurotrophin receptor expression significantly increased in the prefrontal cortex,cerebellum,and hippocampus.These findings confirmed that MDMA induced neuronal apoptosis,and results suggested that this effect was related by upregulated protein expression of the p75 neurotrophin receptor.     

Characterization of NADE,NRIF and SC-1 gene expression during mouse neurogenesis

The p75 neurotrophin receptor (p75NTR) is a member of the tumor necrosis factor receptor superfamily. p75NTR signaling events have been implicated in both cell cycle arrest and apoptosis depending on which effector molecules are associated with its intracellular domain after ligand binding. Two such effector proteins, p75NTR-associated cell death executor (NADE) and neurotrophin receptor interacting factor (NRIF) promote p75NTR-mediated apoptosis, whereas Schwann cell factor-1 (SC-1) mediates neurotrophin-dependent withdrawal from the cell cycle. An understanding of the expression profiles of these three interacting proteins and p75NTR during embryogenesis is critical for addressing whether these effector proteins might function outside of p75NTR-mediated signaling events. The distribution of NADE, NRIF and SC-1 mRNAs during murine development suggests that the action of these genes is in fact not limited to regions of p75NTR expression. Specifically, a detailed comparison of the spatial and temporal expression domains of NADE, NRIF and SC-1 during brain development revealed regions of co-expression with p75NTR but also illustrates a distinct and discordant spatial and temporal expression. These results yield novel insights into the unique developmental characteristics of the three p75NTR-interacting proteins, thus revealing their diverse signaling potential during embryonic development.     

Neurotrophin dependence domain

The mechanisms underlying neurotrophin dependence, and cellular dependent states in general, are unknown. We show that a 29 amino acid region in the intracellular domain of the common neurotrophin receptor, p75^NTR, is required for the mediation of apoptosis by p75^NTR. Furthermore, contrary to results obtained with Fas, monomeric p75^NTR is required for apoptosis induction, whereas multimerization inhibits the pro-apoptotic effect. Within the 29-residue domain required for apoptosis induction by p75^NTR, a 14-residue region is sufficient as a peptide inducer of apoptosis. This 14-residue peptide requires the positively charged carboxyterminal residues for its effect on cell death, and these same residues are required by the full-length p75^NTR. These studies define a novel type of domain that mediates neurotrophin dependence, and suggest that other cellular dependent states may be mediated by proteins displaying similar domains.     

Phosphorylation of the common neurotrophin receptor p75 by p38β2 kinase affects NF-κB and AP-1 activities

The signaling pathways invoked by ligand binding to the common neurotrophin receptor p75^NTR are incompletely understood. Using the yeast two-hybrid system, we identified the mitogen-activated protein (MAP) kinase p38β2 as a specific interactor with the 5th and 6th alpha helices of the p75^NTR intracytoplasmic region. The consequences of this interaction were studied, using primary cultures of Schwann cells and the 293T cell line. Phosphorylation of p75^NTR by p38β2 was induced in vitro and in vivo by MAP kinase kinases (MKK) 6 activation. This pathway demonstrated feedback in that nerve growth factor (NGF) binding increased p38β2 activity, causing an increase of nuclear factor-κB (NF-κB) activation and a decrease of AP-1 activation. The mechanisms described explain at least in part why NGF binding to p75^NTR increases cell survival in certain circumstances.     

Stimulation of glucocorticoid‐induced tumor necrosis factor receptor family‐related protein ligand (GITRL) induces inflammatory activation of microglia in culture

Glucocorticoid‐induced tumor necrosis factor receptor family‐related protein ligand (GITRL) is a member of the tumor necrosis factor superfamily (TNFSF) and is known to act as a costimulator in the immune system by binding to GITR. GITRL is expressed in endothelial cells, dendritic cells, macrophages, and B cells, but it is not known whether GITRL is expressed in brain microglia cells. Here, we investigated the expression of GITR and GITRL and their potential role in microglia cells. Using BV‐2 mouse microglia cells and mouse primary microglia cultures, we have demonstrated that 1) both GITR and GITRL are expressed in microglia cells; 2) stimulation of GITRL induces inflammatory activation of microglia on the basis of production of nitric oxide (NO) and expression of inducible nitric oxide synthase, cyclooxygenase‐2, CD40, and matrix metalloproteinase‐9; 3) GITRL‐mediated microglial NO production partially depends on p38 MAPK, JNK, and nuclear factor‐κB pathways; and 4) GITRL stimulation also induces microglia cell death. These results indicate that GITR and GITRL are functionally expressed on brain microglia and that the stimulation of GITRL can induce inflammatory activation of microglia. The GITR/GITRL system may play an important role in neuroinflammation. © 2010 Wiley‐Liss, Inc.     

Photic injury promotes cleavage of p75NTR by TACE and nuclear trafficking of the p75 intracellular domain

The p75 neurotrophin receptor (p75NTR) is a member of the tumor necrosis factor receptor superfamily that paradoxically mediates neuronal survival and differentiation or apoptotic cell death. Cleavage of p75NTR by a constitutively active metalloprotease could result in shedding of its extracellular domain (p75ECD) and generation of a pro-apoptotic intracellular domain (p75ICD). In this study, we established that exposure of a transgenic mouse photoreceptor cell line to intense light upregulated the expression of p75NTR and of the disintegrin metalloprotease tumor necrosis factor-converting enzyme (TACE) and resulted in apoptotic cell death. Light damage promoted TACE cleavage of p75NTR resulting in shedding of the soluble p75ECD and nuclear translocation of the p75ICD. Overexpression of TACE and p75NTR-induced p75NTR cleavage and secretion of p75ECD, but not nuclear transport of p75ICD. Light-induced cleavage of p75NTR, nuclear localization of p75ICD, and apoptosis were inhibited by IC-3, a metalloprotease inhibitor. Increased levels of p75NTR and TACE were observed in photoreceptor cells of animals with photic injury. Our findings support a role for TACE in the proteolytic cleavage of p75NTR and light-induced apoptosis.     

The p75 neurotrophin receptor: at the crossroad of neural repair and death

The strong repair and pro-survival functions of neurotrophins at their primary receptors, TrkA, TrkB and TrkC, have made them attractive candidates for treatment of nervous system injury and disease. However, difficulties with the clinical implementation of neurotrophin therapies have prompted the search for treatments that are stable, easier to deliver and allow more precise regulation of neurotrophin actions. Recently, the p75 neurotrophin receptor (p75^NTR) has emerged as a potential target for pharmacological control of neurotrophin activity, supported in part by studies demonstrating 1) regulation of neural plasticity in the mature nervous system, 2) promotion of adult neurogenesis and 3) increased expression in neurons, macrophages, microglia, astrocytes and/or Schwann cells in response to injury and neurodegenerative diseases. Although the receptor has no intrinsic catalytic activity it interacts with and modulates the function of TrkA, TrkB, and TrkC, as well as sortilin and the Nogo receptor. This provides substantial cellular and molecular diversity for regulation of neuron survival, neurogenesis, immune responses and processes that support neural function. Upregulation of the p75^NTR under pathological conditions places the receptor in a key position to control numerous processes necessary for nervous system recovery. Support for this possibility has come from recent studies showing that small, non-peptide p75^NTR ligands can selectively modify pro-survival and repair functions. While a great deal remains to be discovered about the wide ranging functions of the p75^NTR, studies summarized in this review highlight the immense potential for development of novel neuroprotective and neurorestorative therapies.     

Expression of low-affinity neurotrophin receptor p75NTR in the peripheral nervous system of human neuropathies

Expression of low-affinity neurotrophin receptor (p75^NTR) was immunohistochemically examined in the peripheral nerve trunks, dorsal root ganglia, sympathetic nerve ganglia and spinal cords in various human neurological diseases manifesting peripheral neuropathies. p75^NTR was expressed in the nerves with axonal degeneration, and was also prominent in the nerves with newly regenerating axons. In contrast, axonal pathology tended to reduce the expression of p75^NTR in the neuronal perikarya of the dorsal root genglion and sympathetic nerve ganglion neurons. In the ventral and lateral horn cells, the p75^NTR immunoreactivity was not detected in the normal and diseased nerves except for amyloid polyneuropathy. These p75^NTR expressions in the diseased human peripheral nervous tissues would be regulated by an underlying pathology-related process, and could play a role in peripheral nerve repair. Received: 28 April 1997 / Revised: 5 August 1997 / Revised, accepted: 17 November 1997     

Effect of glucocorticoids on neuronal and vascular pathology in a transgenic model of selective Müller cell ablation

Retinal diseases such as macular telangiectasis type 2 (MacTel), age‐related macular degeneration (AMD) and diabetic retinopathy (DR) affect both neurons and blood vessels. Treatments addressing both at the same time might have advantages over more specific approaches, such as vascular endothelial growth factor (VEGF) inhibitors, which are used to treat vascular leak but are suspected to have a neurotoxic effect. Here, we studied the effects of an intravitreal injection of triamcinolone acetonide (TA) in a transgenic model in which patchy Müller cell ablation leads to photoreceptor degeneration, vascular leak, and intraretinal neovascularization. TA was injected 4 days before Müller cell ablation. Changes in photoreceptors, microglia and Müller cells, retinal vasculature, differential expression of p75 neurotrophin receptor (p75^NTR), tumor necrosis factor‐α (TNFα), the precursor and mature forms of neurotrophin 3 (pro‐NT3 and mature NT3) and activation of the p53 and p38 stress‐activated protein kinase (p38/SAPK) signaling pathways were examined. We found that TA prevented photoreceptor degeneration and inhibited activation of microglial and Müller cells. TA attenuated Müller cell loss and inhibited overexpression of p75^NTR, TNFα, pro‐NT, and the activation of p53 and p38/SAPK signaling pathways. TA not only prevented the development of retinal vascular lesions but also inhibited fluorescein leakage from established vascular lesions. TA inhibited overexpression of VEGF in transgenic mice but without affecting its basal level expression in the normal retina. Our data suggest that glucocorticoid treatment may be beneficial for treatment of retinal diseases such as MacTel, AMD, and DR that affect both neurons and the vasculature. GLIA 2014;62:1110–1124     

Characterization of the signaling pathway downstream p75 neurotrophin receptor involved in β-amyloid peptide-dependent cell death

The accumulation of β-amyloid (Aβ) peptide is a key pathogenic event in Alzheimer’s disease. Previous studies have shown that Aβ peptide can damage neurons by activating the p75 neurotrophin receptor (p75^NTR). However, the signaling pathway leading to neuronal cell death is not completely understood. By using a neuroblastoma cell line devoid of neurotrophin receptors and engineered to express either a full-length or a death domain (DD)-truncated form of p75^NTR, we demonstrated that Aβ peptide activates the mitogen-activated protein kinases (MAPKs) p38 and c-Jun N-terminal kinase (JNK). We also found that Aβ peptide induces the translocation of nuclear factor-κB (NF-κB). These events depend on the DD of p75^NTR. β-Amyloid (Aβ) peptide was found not to be toxic when the above interactors were inhibited, indicating that they are required for Aβ-induced neuronal cell death. p75 neurotrophin receptor (p75^NTR)-expressing cells became resistant to Aβ toxicity when transfected with dominant-negative mutants of MAPK kinases 3, 4, or 6 (MKK3, MKK4, or MKK6), the inhibitor of κBα, or when treated with chemical inhibitors of p38 and JNK. Furthermore, p75^NTR-expressing cells became resistant to Aβ peptide upon transfection with a dominant-negative mutant of p53. These results were obtained in the presence of normal p38 and JNK activation, indicating that p53 acts downstream of p38 and JNK. Finally, we demonstrated that NF-κB activation is dependent on p38 and JNK activation. Therefore, our data suggest a signaling pathway in which Aβ peptide binds to p75^NTR and activates p38 and JNK in a DD-dependent manner, followed by NF-κB translocation and p53 activation.     

Knockout of p75NTR does not alter the viability of striatal neurons following a metabolic or excitotoxic injury

Following metabolic or excitotoxic injury to the striatum, there is de novo expression of the low-affinity p75 neurotrophin receptor (p75^NTR). The novel expression of this pan neurotrophin receptor in rodents occurs within the lesion core and surrounding area, creating a division between viable and nonviable tissue. The present series of experiments sought to elucidate whether the p75^NTR expression seen following metabolic and excitotoxic injury alters neuronal viability within the striatum. Toward this end, we compared the extent of striatal lesion created with quinolinic acid (QA) or 3-nitropropionic acid (3-NP) in p75^NTR null and wild-type mice. Using stereological techniques, we found that the lesion volume and neuronal cell counts between p75^NTR null and wild-type mice were similar 1, 2, and 4 weeks post-QA or -3-NP lesion. The results indicate that the expression of p75^NTR within reactive astrocytes in the mouse striatum is not a key factor in protecting neuronal cell death following metabolic and excitotoxic insults.     

The cholinergic system, nerve growth factor and the cytoskeleton

Cholinergic neurons of the basal forebrain provide the major cholinergic innervation to the cortex and hippocampus, and play a key role in memory and attentional processes. Dysfunction of basal forebrain cholinergic neurons (BFCN) is a cardinal feature of Alzheimer's disease (AD) and correlates with cognitive decline. Survival of BFCN neurons depends upon binding of nerve growth factor (NGF), which is synthesized and secreted by cells in the cortex and hippocampus, with high-affinity (TrkA) and low-affinity (p75^NTR) neurotrophin receptors produced within BFCN neurons. NGF released from target cells activates TrkA on axon terminals and triggers activation of PI3K/Akt, MEK/ERK, and PLCγ (phospholipase C) signaling pathways. The signal then travels retrogradely along axon to cell body to promote neuronal survival. However, the nature of the retrograde signal remains mysterious. p75^NTR receptors could mediate a fundamentally different signaling pathway leading to apoptic cell death. Dysfunction of NGF and its receptors has been suggested to underlie the selective degeneration of the BFCN in end stage Alzheimer disease. In this regard, NGF, the founding member of the neurotrophin family, has generated great interest as a potential target for the treatment of AD. This review focuses on NGF-cholinergic dependency, NGF/receptor binding, signal transduction, retrograde transport, regulation of specific cellular endpoints, and the potential involvement of cytoskeleton dysfunction in defected NGF signaling.     

p75NTR regulates brain mononuclear cell function and neuronal structure in Toxoplasma infection-induced neuroinflammation

Neurotrophins mediate neuronal growth, differentiation, and survival via tropomyosin receptor kinase (Trk) or p75 neurotrophin receptor (p75^NTR) signaling. The p75^NTR is not exclusively expressed by neurons but also by certain immune cells, implying a role for neurotrophin signaling in the immune system. In this study, we investigated the effect of p75^NTR on innate immune cell behavior and on neuronal morphology upon chronic Toxoplasma gondii (T. gondii) infection-induced neuroinflammation. Characterization of the immune cells in the periphery and central nervous system (CNS) revealed that innate immune cell subsets in the brain upregulated p75^NTR upon infection in wild-type mice. Although cell recruitment and phagocytic capacity of p75^NTRexonIV knockout (p75^−/−) mice were not impaired, the activation status of resident microglia and recruited myeloid cell subsets was altered. Importantly, recruited mononuclear cells in brains of infected p75^−/− mice upregulated the production of the cytokines interleukin (IL)-10, IL-6 as well as IL-1α. Protein levels of proBDNF, known to negatively influence neuronal morphology by binding p75^NTR, were highly increased upon chronic infection in the brain of wild-type and p75^−/− mice. Moreover, upon infection the activated immune cells contributed to the proBDNF release. Notably, the neuroinflammation-induced changes in spine density were rescued in the p75^−/− mice. In conclusion, these findings indicate that neurotrophin signaling via the p75^NTR affects innate immune cell behavior, thus, influencing the structural plasticity of neurons under inflammatory conditions.     

Nerve growth factor attenuates proliferation of astrocytes via the p75 neurotrophin receptor

The p75 neurotrophin receptor has been implicated in the regulation of multiple cellular functions that differ depending on the cell context. We have observed that p75^NTR is strongly induced on astrocytes as well as neurons in the hippocampal CA3 region after seizures; however, the function of this receptor on these glial cells has not been defined. We have employed a primary culture system to investigate the effects of neurotrophins on astrocytes. Treatment of hippocampal astrocytes with nerve growth factor (NGF) caused a reduction in cell number, but did not elicit an apoptotic response, in contrast to hippocampal neurons. Instead, activation of p75^NTR by NGF attenuated proliferation induced by mitogens such as EGF or serum. These studies demonstrate the cell type specificity of neurotrophin functions in the brain. © 2009 Wiley‐Liss, Inc.     

Nerve growth factor (NGF) induces motoneuron apoptosis in rat embryonic spinal cord in vitro

Recent studies have demonstrated that nerve growth factor (NGF) induces apoptosis of several cell types in the central nervous system through its low-affinity p75 neurotrophin receptor (p75NTR). To test the effect of NGF on embryonic motoneuron survival, we developed an organotypic culture system which allowed the in vitro development of intact embryonic rat spinal cords. In our system, neural tubes were taken and cultured at E13, just before the onset of physiological motoneuron death. After 2 days in vitro (DIV), motoneurons underwent apoptosis over a time-course similar to that in vivo. In this system, the addition of NGF (200 ng/mL) for 2 days enhanced the number of apoptotic motoneurons by 37%. This pro-apoptotic effect was completely reversed by the blocking anti-p75NTR (REX) antibody which inhibits NGF binding to p75NTR. Other neurotrophins, e.g. brain-derived neurotrophic factor (BDNF), neurotrophin 3 (NT3) and neurotrophin 4/5 (NT4/5) did not have any effect, while glial cell-derived neurotrophic factor (GDNF) promoted motoneuron survival. Anti-BDNF blocking antibodies enhanced motoneuron death indicating that endogenous BDNF promotes motoneuron survival in explants. Our results demonstrate, for the first time, that NGF can induce embryonic motoneuron apoptosis through its receptor p75NTR.