RESUMEN
A subpopulation of deeply quiescent, so-called dormant hematopoietic stem cells (dHSCs) resides at the top of the hematopoietic hierarchy and serves as a reserve pool for HSCs. The state of dormancy protects the HSC pool from exhaustion throughout life; however, excessive dormancy may prevent an efficient response to hematological stresses. Despite the significance of dHSCs, the mechanisms maintaining their dormancy remain elusive. Here, we identify CD38 as a novel and broadly applicable surface marker for the enrichment of murine dHSCs. We demonstrate that cyclic adenosine diphosphate ribose (cADPR), the product of CD38 cyclase activity, regulates the expression of the transcription factor c-Fos by increasing the release of Ca2+ from the endoplasmic reticulum (ER). Subsequently, we uncover that c-Fos induces the expression of the cell cycle inhibitor p57Kip2 to drive HSC dormancy. Moreover, we found that CD38 ecto-enzymatic activity at the neighboring CD38-positive cells can promote human HSC quiescence. Together, CD38/cADPR/Ca2+/c-Fos/p57Kip2 axis maintains HSC dormancy. Pharmacological manipulations of this pathway can provide new strategies to improve the success of stem cell transplantation and blood regeneration after injury or disease.
Asunto(s)
ADP-Ribosil Ciclasa 1 , ADP-Ribosa Cíclica , Animales , Humanos , Ratones , Calcio/metabolismo , ADP-Ribosa Cíclica/metabolismo , Células Madre Hematopoyéticas , ADP-Ribosil Ciclasa 1/metabolismo , Inhibidor p57 de las Quinasas Dependientes de la Ciclina/metabolismoRESUMEN
Sterile alpha and HEAT/armadillo motif-containing protein (SARM1) was recently described as a NAD+-consuming enzyme and has previously been shown to regulate immune responses in macrophages. Neuronal SARM1 is known to contribute to axon degeneration due to its NADase activity. However, how SARM1 affects macrophage metabolism has not been explored. Here, we show that macrophages from Sarm1-/- mice display elevated NAD+ concentrations and lower cyclic ADP-ribose, a known product of SARM1-dependent NAD+ catabolism. Further, SARM1-deficient macrophages showed an increase in the reserve capacity of oxidative phosphorylation and glycolysis compared to WT cells. Stimulation of macrophages to a proinflammatory state by lipopolysaccharide (LPS) revealed that SARM1 restricts the ability of macrophages to upregulate glycolysis and limits the expression of the proinflammatory gene interleukin (Il) 1b, but boosts expression of anti-inflammatory Il10. In contrast, we show macrophages lacking SARM1 induced to an anti-inflammatory state by IL-4 stimulation display increased oxidative phosphorylation and glycolysis, and reduced expression of the anti-inflammatory gene, Fizz1. Overall, these data show that SARM1 fine-tunes immune gene transcription in macrophages via consumption of NAD+ and altered macrophage metabolism.
Asunto(s)
Proteínas del Dominio Armadillo , Proteínas del Citoesqueleto , Neuronas , Animales , Ratones , Proteínas del Dominio Armadillo/genética , Proteínas del Dominio Armadillo/metabolismo , Axones/metabolismo , ADP-Ribosa Cíclica/metabolismo , Proteínas del Citoesqueleto/genética , Proteínas del Citoesqueleto/metabolismo , NAD/metabolismo , Neuronas/metabolismoRESUMEN
Adipocytes play a key role in energy storage and homeostasis. Although the role of transcription factors in adipocyte differentiation is known, the effect of endogenous metabolites of low molecular weight remains unclear. Here, we analyzed time-dependent changes in the levels of these metabolites throughout adipocyte differentiation, using metabolome analysis, and demonstrated that there is a positive correlation between cyclic adenosine diphosphate ribose (cADPR) and Pparγ mRNA expression used as a marker of differentiation. We also found that the treatment of C3H10T1/2 adipocytes with cADPR increased the mRNA expression of those marker genes and the accumulation of triglycerides. Furthermore, inhibition of ryanodine receptors (RyR), which are activated by cADPR, caused a significant reduction in mRNA expression levels of the marker genes and triglyceride accumulation in adipocytes. Our findings show that cADPR accelerates adipocytic differentiation via RyR pathway.
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Adipocitos , ADP-Ribosa Cíclica , Ratones , Animales , ADP-Ribosa Cíclica/metabolismo , Adipocitos/metabolismo , Factores de Transcripción/metabolismo , PPAR gamma/metabolismo , Metaboloma , ARN Mensajero/genética , Diferenciación Celular , Adenosina Difosfato Ribosa/metabolismo , Adenosina Difosfato Ribosa/farmacología , Adipogénesis/genética , Células 3T3-L1RESUMEN
Asthma pathobiology includes oxidative stress that modifies cell membranes and extracellular phospholipids. Oxidized phosphatidylcholines (OxPCs) in lung lavage from allergen-challenged human participants correlate with airway hyperresponsiveness and induce bronchial narrowing in murine thin-cut lung slices. OxPCs activate many signaling pathways, but mechanisms for these responses are unclear. We hypothesize that OxPCs stimulate intracellular free Ca2+ flux to trigger airway smooth muscle contraction. Intracellular Ca2+ flux was assessed in Fura-2-loaded, cultured human airway smooth muscle cells. Oxidized 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine (OxPAPC) induced an approximately threefold increase in 20 kD myosin light chain phosphorylation. This correlated with a rapid peak in intracellular cytoplasmic Ca2+ concentration ([Ca2+]i) (143 nM) and a sustained plateau that included slow oscillations in [Ca2+]i. Sustained [Ca2+]i elevation was ablated in Ca2+-free buffer and by TRPA1 inhibition. Conversely, OxPAPC-induced peak [Ca2+]i was unaffected in Ca2+-free buffer, by TRPA1 inhibition, or by inositol 1,4,5-triphosphate receptor inhibition. Peak [Ca2+]i was ablated by pharmacologic inhibition of ryanodine receptor (RyR) Ca2+ release from the sarcoplasmic reticulum. Inhibiting the upstream RyR activator cyclic adenosine diphosphate ribose with 8-bromo-cyclic adenosine diphosphate ribose was sufficient to abolish OxPAPC-induced cytoplasmic Ca2+ flux. OxPAPC induced â¼15% bronchial narrowing in thin-cut lung slices that could be prevented by pharmacologic inhibition of either TRPA1 or RyR, which similarly inhibited OxPC-induced myosin light chain phosphorylation in cultured human airway smooth muscle cells. In summary, OxPC mediates airway narrowing by triggering TRPA1 and RyR-mediated mobilization of intracellular and extracellular Ca2+ in airway smooth muscle. These data suggest that OxPC in the airways of allergen-challenged subjects and subjects with asthma may contribute to airway hyperresponsiveness.
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Asma , Hipersensibilidad Respiratoria , Humanos , Animales , Ratones , Canal Liberador de Calcio Receptor de Rianodina/metabolismo , Miocitos del Músculo Liso/metabolismo , Cadenas Ligeras de Miosina/metabolismo , ADP-Ribosa Cíclica/metabolismo , Asma/metabolismo , Contracción Muscular/fisiología , Hipersensibilidad Respiratoria/metabolismo , Fosfatidilcolinas/metabolismo , Alérgenos/metabolismo , Calcio/metabolismo , Canal Catiónico TRPA1/metabolismoRESUMEN
We previously identified glyceraldehyde 3-phosphate dehydrogenase (GAPDH) as one of the cyclic adenosine diphosphoribose (cADPR)'s binding proteins and found that GAPDH participates in cADPR-mediated Ca2+ release from endoplasmic reticulum via ryanodine receptors (RyRs). Here, we aimed to chemically synthesise and pharmacologically characterise novel cADPR analogues. Based on the simulated cADPR-GAPDH complex structure, we performed the structure-based drug screening, identified several small chemicals with high docking scores to cADPR's binding pocket in GAPDH and showed that two of these compounds, C244 and C346, are potential cADPR antagonists. We further synthesised several analogues of C346 and found that its analogue, G42, also mobilised Ca2+ release from lysosomes. G42 alkalised lysosomal pH and inhibited autophagosome-lysosome fusion. Moreover, G42 markedly inhibited Zika virus (ZIKV, a flavivirus) or murine hepatitis virus (MHV, a ß-coronavirus) infections of host cells. These results suggest that G42 inhibits virus infection, likely by triggering lysosomal Ca2+ mobilisation and inhibiting autophagy.
Asunto(s)
Infección por el Virus Zika , Virus Zika , Animales , Ratones , Humanos , Calcio/metabolismo , ADP-Ribosa Cíclica/metabolismo , Virus Zika/metabolismo , Canal Liberador de Calcio Receptor de Rianodina/genética , Canal Liberador de Calcio Receptor de Rianodina/metabolismo , Autofagia , Lisosomas/metabolismo , Adenosina Difosfato Ribosa/metabolismoRESUMEN
BST-1 (bone marrow stromal cell antigen-1) is thought to be a key molecule involved in regulating the functional activity of cells in various tissues and organs. BST-1 can catalyze the hydrolysis of nicotinamide adenine dinucleotide (NAD+) to produce cyclic ADP ribose (cADPR), which activates the activity of intracellular Ca2+ signaling. Currently, the role of BST-1 regulation of Ca2+ signaling pathway in pathological myocardial hypertrophy is unclear. We found elevated expression of BST-1 in cardiac hypertrophy tissues of spontaneously hypertensive rats in our vivo study, subsequently; the mechanism of BST-1 action on myocardial hypertrophy was explored in vitro experiment. We used aldosterone (ALD) to induce H9C2 cellular hypertrophy. cADPR levels and intracellular Ca2+ concentrations declined and calcium-regulated neurophosphatase (CaN) activity and protein expression were decreased after BST-1 knockdown. And then activated T-cell nuclear factor (NFATc3) entry nucleus was inhibited. All of the above resulted in that H9C2 cells size was reduced by rhodamine-phalloidin staining. Thus, BST-1 may exacerbate cardiac hypertrophy by activating the Ca2+/CaN/NFATc3 pathway.
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Calcio , ADP-Ribosa Cíclica , Ratas , Animales , Calcio/metabolismo , ADP-Ribosil Ciclasa 1/metabolismo , ADP-Ribosa Cíclica/metabolismo , Aldosterona , Factores de Transcripción NFATC , Cardiomegalia/inducido químicamenteRESUMEN
Social behavior is essential for health, survival, and reproduction of animals; however, the role of astrocytes in social behavior remains largely unknown. The transmembrane protein CD38, which acts both as a receptor and ADP-ribosyl cyclase to produce cyclic ADP-ribose (cADPR) regulates social behaviors by promoting oxytocin release from hypothalamic neurons. CD38 is also abundantly expressed in astrocytes in the postnatal brain and is important for astroglial development. Here, we demonstrate that the astroglial-expressed CD38 plays an important role in social behavior during development. Selective deletion of CD38 in postnatal astrocytes, but not in adult astrocytes, impairs social memory without any other behavioral abnormalities. Morphological analysis shows that depletion of astroglial CD38 in the postnatal brain interferes with synapse formation in the medial prefrontal cortex (mPFC) and hippocampus. Moreover, astroglial CD38 expression promotes synaptogenesis of excitatory neurons by increasing the level of extracellular SPARCL1 (also known as Hevin), a synaptogenic protein. The release of SPARCL1 from astrocytes is regulated by CD38/cADPR/calcium signaling. These data demonstrate a novel developmental role of astrocytes in neural circuit formation and regulation of social behavior in adults.
Asunto(s)
Antígenos CD , ADP-Ribosa Cíclica , Animales , ADP-Ribosil Ciclasa 1/genética , Antígenos CD/metabolismo , ADP-Ribosa Cíclica/metabolismo , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/metabolismo , Astrocitos/metabolismo , Sinapsis/metabolismoRESUMEN
Nicotinamide adenine dinucleotide (NAD+) has emerged as a key component in prokaryotic and eukaryotic immune systems. The recent discovery that Toll/interleukin-1 receptor (TIR) proteins function as NAD+ hydrolases (NADase) links NAD+-derived small molecules with immune signaling. We investigated pathogen manipulation of host NAD+ metabolism as a virulence strategy. Using the pangenome of the model bacterial pathogen Pseudomonas syringae, we conducted a structure-based similarity search from 35,000 orthogroups for type III effectors (T3Es) with potential NADase activity. Thirteen T3Es, including five newly identified candidates, were identified that possess domain(s) characteristic of seven NAD+-hydrolyzing enzyme families. Most Pseudomonas syringae strains that depend on the type III secretion system to cause disease, encode at least one NAD+-manipulating T3E, and many have several. We experimentally confirmed the type III-dependent secretion of a novel T3E, named HopBY, which shows structural similarity to both TIR and adenosine diphosphate ribose (ADPR) cyclase. Homologs of HopBY were predicted to be type VI effectors in diverse bacterial species, indicating potential recruitment of this activity by microbial proteins secreted during various interspecies interactions. HopBY efficiently hydrolyzes NAD+ and specifically produces 2'cADPR, which can also be produced by TIR immune receptors of plants and by other bacteria. Intriguingly, this effector promoted bacterial virulence, indicating that 2'cADPR may not be the signaling molecule that directly initiates immunity. This study highlights a host-pathogen battleground centered around NAD+ metabolism and provides insight into the NAD+-derived molecules involved in plant immunity.
Asunto(s)
ADP-Ribosa Cíclica , NAD , Virulencia , NAD/metabolismo , ADP-Ribosa Cíclica/metabolismo , Bacterias/metabolismo , Plantas/metabolismo , Pseudomonas syringae/metabolismo , NAD+ Nucleosidasa/genética , NAD+ Nucleosidasa/metabolismo , Proteínas Bacterianas/genética , Proteínas Bacterianas/metabolismo , Enfermedades de las Plantas/microbiologíaRESUMEN
ADP-ribosyl cyclases (ADPRCs) catalyze the synthesis of the Ca2+-active second messengers Cyclic ADP-ribose (cADPR) and ADP-ribose (ADPR) from NAD+ as well as nicotinic acid adenine dinucleotide phosphate (NAADP+) from NADP+. The best characterized ADPRC in mammals is CD38, a single-pass transmembrane protein with two opposite membrane orientations. The first identified form, type II CD38, is a glycosylated ectoenzyme, while type III CD38 has its active site in the cytosol. The ectoenzymatic nature of type II CD38 raised long ago the question of a topological paradox concerning the access of the intracellular NAD+ substrate to the extracellular active site and of extracellular cADPR product to its intracellular receptors, ryanodine (RyR) channels. Two different transporters, equilibrative connexin 43 (Cx43) hemichannels for NAD+ and concentrative nucleoside transporters (CNTs) for cADPR, proved to mediate cell-autonomous trafficking of both nucleotides. Here, we discussed how type II CD38, Cx43 and CNTs also play a role in mediating several paracrine processes where an ADPRC+ cell supplies a neighboring CNT-and RyR-expressing cell with cADPR. Recently, type II CD38 was shown to start an ectoenzymatic sequence of reactions from NAD+/ADPR to the strong immunosuppressant adenosine; this paracrine effect represents a major mechanism of acquired resistance of several tumors to immune checkpoint therapy.
Asunto(s)
Fenómenos Biológicos , ADP-Ribosa Cíclica , ADP-Ribosil Ciclasa/metabolismo , ADP-Ribosil Ciclasa 1/metabolismo , Animales , Antígenos CD/metabolismo , Conexina 43/metabolismo , ADP-Ribosa Cíclica/metabolismo , Mamíferos/metabolismo , Glicoproteínas de Membrana/metabolismo , NAD/metabolismoRESUMEN
Sleep apnea syndrome (SAS) is characterized by recurrent episodes of oxygen desaturation and reoxygenation (intermittent hypoxia, IH), and it is a risk factor for cardiovascular disease (CVD) and insulin resistance/type 2 diabetes. However, the mechanisms linking IH stress and CVD remain elusive. We exposed rat H9c2 and mouse P19.CL6 cardiomyocytes to experimental IH or normoxia for 24 h to analyze the mRNA expression of the components of Cd38-cyclic ADP-ribose (cADPR) signaling. We found that the mRNA levels of cluster of differentiation 38 (Cd38), type 2 ryanodine receptor (Ryr2), and FK506-binding protein 12.6 (Fkbp12.6) in H9c2 and P19.CL6 cardiomyocytes were significantly decreased by IH, whereas the promoter activities of these genes were not decreased. By contrast, the expression of phosphatase and tensin homolog deleted from chromosome 10 (Pten) was upregulated in IH-treated cells. The small interfering RNA for Pten (siPten) and a non-specific control RNA were introduced into the H9c2 cells. The IH-induced downregulation of Cd38, Ryr2, and Fkbp12.6 was abolished by the introduction of the siPten, but not by the control RNA. These results indicate that IH stress upregulated the Pten in cardiomyocytes, resulting in the decreased mRNA levels of Cd38, Ryr2, and Fkbp12.6, leading to the inhibition of cardiomyocyte functions in SAS patients.
Asunto(s)
Enfermedades Cardiovasculares , Diabetes Mellitus Tipo 2 , ADP-Ribosil Ciclasa/genética , ADP-Ribosil Ciclasa 1 , Animales , Señalización del Calcio , Enfermedades Cardiovasculares/metabolismo , ADP-Ribosa Cíclica/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Regulación hacia Abajo , Hipoxia/metabolismo , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/metabolismo , Ratones , Miocitos Cardíacos/metabolismo , Fosfohidrolasa PTEN/genética , Fosfohidrolasa PTEN/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Ratas , Canal Liberador de Calcio Receptor de Rianodina/genética , Canal Liberador de Calcio Receptor de Rianodina/metabolismo , Regulación hacia ArribaRESUMEN
Calcium (Ca2+) is a ubiquitous and fundamental signaling component that is utilized by cells to regulate a diverse range of cellular functions, such as insulin secretion from pancreatic ß-cells of the islets of Langerhans. Cyclic ADP-ribose (cADPR), synthesized from NAD+ by ADP-ribosyl cyclase family proteins, such as the mammalian cluster of differentiation 38 (CD38), is important for intracellular Ca2+ mobilization for cell functioning. cADPR induces Ca2+ release from endoplasmic reticulum via the ryanodine receptor intracellular Ca2+ channel complex, in which the FK506-binding protein 12.6 works as a cADPR-binding regulatory protein. Recently, involvements of the CD38-cADPR signal system in several human diseases and animal models have been reported. This review describes the biochemical and molecular biological basis of the CD38-cADPR signal system and the diseases caused by its abnormalities.
Asunto(s)
Antígenos CD , ADP-Ribosa Cíclica , ADP-Ribosil Ciclasa 1/metabolismo , Animales , Antígenos CD/metabolismo , Antígenos de Diferenciación/metabolismo , Calcio/metabolismo , Señalización del Calcio , ADP-Ribosa Cíclica/metabolismo , Mamíferos/metabolismo , Glicoproteínas de Membrana/metabolismoRESUMEN
cADPR is a second messenger that releases Ca2+ from intracellular stores via the ryanodine receptor. Over more than 15 years, it has been controversially discussed whether cADPR also contributes to the activation of the nucleotide-gated cation channel TRPM2. While some groups have observed activation of TRPM2 by cADPR alone or in synergy with ADPR, sometimes only at 37 °C, others have argued that this is due to the contamination of cADPR by ADPR. The identification of a novel nucleotide-binding site in the N-terminus of TRPM2 that binds ADPR in a horseshoe-like conformation resembling cADPR as well as the cADPR antagonist 8-Br-cADPR, and another report that demonstrates activation of TRPM2 by binding of cADPR to the NUDT9H domain raised the question again and led us to revisit the topic. Here we show that (i) the N-terminal MHR1/2 domain and the C-terminal NUDT9H domain are required for activation of human TRPM2 by ADPR and 2'-deoxy-ADPR (2dADPR), (ii) that pure cADPR does not activate TRPM2 under a variety of conditions that have previously been shown to result in channel activation, (iii) the cADPR antagonist 8-Br-cADPR also inhibits activation of TRPM2 by ADPR, and (iv) cADPR does not bind to the MHR1/2 domain of TRPM2 while ADPR does.
Asunto(s)
ADP-Ribosa Cíclica , Canales Catiónicos TRPM , Sitios de Unión , Calcio/metabolismo , Señalización del Calcio , ADP-Ribosa Cíclica/metabolismo , Humanos , Canales Catiónicos TRPM/metabolismoRESUMEN
Since the discovery of the pyridine nucleotide metabolites Ca2+ mobilizing messengers cyclic ADP-ribose (cADPR) and nicotinic acid adenine dinucleotide phosphate (NAADP), they have been demonstrated to function as Ca2+ signaling messengers in a wide range of cell types. In this review, I will briefly summarize the roles of cADPR and NAADP in the physiological process of stimulus-secretion coupling in pancreatic ß cells. I am also going to outline the current breadth of knowledge regarding intracellular Ca2+ stores and Ca2+ channels targeted by cADPR and NAADP, as well as the biogenesis of these Ca2+ signaling messengers. I focused on receptor-mediated Ca2+ signaling in mediating the effects of GLP-1 and insulin in pancreatic ß cells. A better grasp in the roles of these signaling messengers will assist in our understanding of Ca2+ signaling as well as pathophysiology.
Asunto(s)
ADP-Ribosa Cíclica , Células Secretoras de Insulina , Calcio/metabolismo , Señalización del Calcio/fisiología , ADP-Ribosa Cíclica/metabolismo , Células Secretoras de Insulina/metabolismo , NADP/análogos & derivados , NADP/metabolismoRESUMEN
Adenosine diphosphate ribose cyclase (ADPRC) exists widely in eukaryotes and lower metazoans cells. It can degrade nicotinamide adenine dinucleotide (NAD) into cyclic ADP ribose (cADPR) and nicotinamide, and subsequently hydrolyses cADPR to ADP ribose (ADPR). In this paper, we have summarized the relative subcellular localization of ADPRC and enzymes with ADPRC activity in organisms, related enzyme family members of ADPRC are also described. In addition, we discussed the main biological functions of ADPRC, the regulation of Ca2+ signal, the regulation of insulin and glucagon secretion, oxytocin secretion, and the effects of renal and pulmonary vasomotor tension. Finally, we expounded the relationship between ADPRC and human health and disease occurrence. It provides a theoretical basis for the targeted treatment of ADPRC as a pharmacological tool for related diseases, and has important significance in clinical diagnosis and disease intervention.
Asunto(s)
ADP-Ribosil Ciclasa/metabolismo , Señalización del Calcio , ADP-Ribosil Ciclasa/análisis , ADP-Ribosil Ciclasa 1 , ADP-Ribosa Cíclica/metabolismo , Humanos , NAD/metabolismo , Fenómenos FisiológicosRESUMEN
CD38 is a single-pass transmembrane enzyme catalyzing the synthesis of two nucleotide second messengers, cyclic ADP-ribose (cADPR) from NAD and nicotinic acid adenine dinucleotide phosphate (NAADP) from NADP. The former mediates the mobilization of the endoplasmic Ca2+-stores in response to a wide range of stimuli, while NAADP targets the endo-lysosomal stores. CD38 not only possesses multiple enzymatic activities, it also exists in two opposite membrane orientations. Type III CD38 has the catalytic domain facing the cytosol and is responsible for producing cellular cADPR. The type II CD38 has an opposite orientation and is serving as a surface receptor mediating extracellular functions such as cell adhesion and lymphocyte activation. Its ecto-NADase activity also contributes to the recycling of external NAD released by apoptosis. Endocytosis can deliver surface type II CD38 to endo-lysosomes, which acidic environment favors the production of NAADP. This article reviews the rationale and evidence that have led to CD38 as a paradigm for membrane topology defining distinct functions of proteins. Also described is the recent discovery of a hitherto unknown cADPR-synthesizing enzyme, SARM1, ushering in a new frontier in cADPR-mediated Ca2+-signaling.
Asunto(s)
Señalización del Calcio , Calcio , ADP-Ribosil Ciclasa 1/metabolismo , Calcio/metabolismo , ADP-Ribosa Cíclica/metabolismo , Lisosomas/metabolismo , NADP/metabolismoRESUMEN
Cancer patients frequently develop chemotherapy-induced peripheral neuropathy (CIPN), a painful and long-lasting disorder with profound somatosensory deficits. There are no effective therapies to prevent or treat this disorder. Pathologically, CIPN is characterized by a "dying-back" axonopathy that begins at intra-epidermal nerve terminals of sensory neurons and progresses in a retrograde fashion. Calcium dysregulation constitutes a critical event in CIPN, but it is not known how chemotherapies such as paclitaxel alter intra-axonal calcium and cause degeneration. Here, we demonstrate that paclitaxel triggers Sarm1-dependent cADPR production in distal axons, promoting intra-axonal calcium flux from both intracellular and extracellular calcium stores. Genetic or pharmacologic antagonists of cADPR signaling prevent paclitaxel-induced axon degeneration and allodynia symptoms, without mitigating the anti-neoplastic efficacy of paclitaxel. Our data demonstrate that cADPR is a calcium-modulating factor that promotes paclitaxel-induced axon degeneration and suggest that targeting cADPR signaling provides a potential therapeutic approach for treating paclitaxel-induced peripheral neuropathy (PIPN).
Asunto(s)
Proteínas del Dominio Armadillo/metabolismo , Axones/metabolismo , Calcio/metabolismo , ADP-Ribosa Cíclica/metabolismo , Proteínas del Citoesqueleto/metabolismo , Degeneración Nerviosa/patología , Paclitaxel/efectos adversos , Enfermedades del Sistema Nervioso Periférico/inducido químicamente , Enfermedades del Sistema Nervioso Periférico/metabolismo , Animales , Canales de Calcio/metabolismo , ADP-Ribosa Cíclica/antagonistas & inhibidores , Femenino , Células HEK293 , Humanos , Ratones Endogámicos C57BL , Ratas Sprague-DawleyRESUMEN
The Toll/interleukin-1 receptor (TIR) domain is a canonical component of animal and plant immune systems1,2. In plants, intracellular pathogen sensing by immune receptors triggers their TIR domains to generate a molecule that is a variant of cyclic ADP-ribose3,4. This molecule is hypothesized to mediate plant cell death through a pathway that has yet to be resolved5. TIR domains have also been shown to be involved in a bacterial anti-phage defence system called Thoeris6, but the mechanism of Thoeris defence remained unknown. Here we show that phage infection triggers Thoeris TIR-domain proteins to produce an isomer of cyclic ADP-ribose. This molecular signal activates a second protein, ThsA, which then depletes the cell of the essential molecule nicotinamide adenine dinucleotide (NAD) and leads to abortive infection and cell death. We also show that, similar to eukaryotic innate immune systems, bacterial TIR-domain proteins determine the immunological specificity to the invading pathogen. Our results describe an antiviral signalling pathway in bacteria, and suggest that the generation of intracellular signalling molecules is an ancient immunological function of TIR domains that is conserved in both plant and bacterial immunity.
Asunto(s)
Bacillus/inmunología , Bacillus/virología , Proteínas Bacterianas/química , Proteínas Bacterianas/inmunología , Bacteriófagos/inmunología , Receptores de Interleucina-1/química , Transducción de Señal/inmunología , Receptores Toll-Like/química , ADP-Ribosa Cíclica/análogos & derivados , ADP-Ribosa Cíclica/metabolismo , Evolución Molecular , Modelos Moleculares , NAD/metabolismo , Dominios Proteicos , Especificidad por Sustrato/inmunologíaRESUMEN
Although a monoclonal antibody targeting the multifunctional ectoenzyme CD38 is an FDA-approved drug, few small molecule inhibitors exist for this enzyme that catalyzes inter alia the formation and metabolism of the N1-ribosylated, Ca2+-mobilizing, second messenger cyclic adenosine 5'-diphosphoribose (cADPR). N1-Inosine 5'-monophosphate (N1-IMP) is a fragment directly related to cADPR. 8-Substituted-N1-IMP derivatives, prepared by degradation of cyclic parent compounds, inhibit CD38-mediated cADPR hydrolysis more efficiently than related cyclic analogues, making them attractive for inhibitor development. We report a total synthesis of the N1-IMP scaffold from adenine and a small initial compound series that facilitated early delineation of structure-activity parameters, with analogues evaluated for inhibition of CD38-mediated hydrolysis of cADPR. The 5'-phosphate group proved essential for useful activity, but substitution of this group by a sulfonamide bioisostere was not fruitful. 8-NH2-N1-IMP is the most potent inhibitor (IC50 = 7.6 µM) and importantly HPLC studies showed this ligand to be cleaved at high CD38 concentrations, confirming its access to the CD38 catalytic machinery and demonstrating the potential of our fragment approach.
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ADP-Ribosil Ciclasa 1/antagonistas & inhibidores , ADP-Ribosa Cíclica/metabolismo , Inosina/metabolismo , Bibliotecas de Moléculas Pequeñas/química , ADP-Ribosil Ciclasa 1/metabolismo , Adenosina Difosfato Ribosa/metabolismo , Calcio/metabolismo , Catálisis/efectos de los fármacos , Humanos , Hidrólisis/efectos de los fármacos , Inosina Monofosfato/química , Bibliotecas de Moléculas Pequeñas/farmacología , Relación Estructura-ActividadRESUMEN
Sleep apnea syndrome is characterized by recurrent episodes of oxygen desaturation and reoxygenation (intermittent hypoxia [IH]), and it is a known risk factor for hypertension. The upregulation of the renin-angiotensin system has been reported in IH, and the correlation between renin and CD38 has been noted. We exposed human HEK293 and mouse As4.1 renal cells to experimental IH or normoxia for 24 h and then measured the mRNA levels using a real-time reverse transcription polymerase chain reaction. The mRNA levels of Renin (Ren) and Cd38 were significantly increased by IH, indicating that they could be involved in the CD38-cyclic ADP-ribose signaling pathway. We next investigated the promotor activities of both genes, which were not increased by IH. Yet, a target mRNA search of the microRNA (miRNA) revealed both mRNAs to have a potential target sequence for miR-203. The miR-203 level of the IH-treated cells was significantly decreased when compared with the normoxia-treated cells. The IH-induced upregulation of the genes was abolished by the introduction of the miR-203 mimic, but not the miR-203 mimic NC negative control. These results indicate that IH stress downregulates the miR-203 in renin-producing cells, thereby resulting in increased mRNA levels of Ren and Cd38, which leads to hypertension.
Asunto(s)
ADP-Ribosil Ciclasa 1/genética , Regulación hacia Abajo/genética , Hipoxia/genética , MicroARNs/genética , Renina/genética , Regulación hacia Arriba/genética , ADP-Ribosil Ciclasa 1/metabolismo , Animales , ADP-Ribosa Cíclica/análogos & derivados , ADP-Ribosa Cíclica/metabolismo , Células HEK293 , Humanos , Ratones , MicroARNs/metabolismo , Regiones Promotoras Genéticas , ARN Mensajero/genética , ARN Interferente Pequeño/metabolismo , Renina/metabolismo , Canal Liberador de Calcio Receptor de Rianodina/metabolismoRESUMEN
It has been recently reported that CD38 expressed on tumor cells of multiple murine and human origins could be upregulated in response to PD-L1 antibody therapy, which led to dysfunction of tumor-infiltrating CD8+ T immune cells due to increasing the production of adenosine. However, the role of tumor expressed-CD38 on neoplastic formation and progression remains elusive. In the present study, we aimed to delineate the molecular and biochemical function of the tumor-associated CD38 in lung adenocarcinoma progression. Our clinical data showed that the upregulation of tumor-originated CD38 was correlated with poor survival of lung cancer patients. Using multiple in vitro assays we found that the enzymatic activity of tumor expressed-CD38 facilitated lung cancer cell migration, proliferation, colony formation, and tumor development. Consistently, our in vivo results showed that inhibition of the enzymatic activity or antagonizing the enzymatic product of CD38 resulted in the similar inhibition of tumor proliferation and metastasis as CD38 gene knock-out or mutation. At biochemical level, we further identified that cADPR, the mainly hydrolytic product of CD38, was responsible for inducing the opening of TRPM2 iron channel leading to the influx of intracellular Ca2+ and then led to increasing levels of NRF2 while decreasing expression of KEAP1 in lung cancer cells. These findings suggested that malignant lung cancer cells were capable of using cADPR catalyzed by CD38 to facilitate tumor progression, and blocking the enzymatic activity of CD38 could be represented as an important strategy for preventing tumor progression.