Equilibrative nucleoside transporter 1 inhibition rescues energy dysfunction and pathology in a model of tauopathy.
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ABSTRACT:
SUBMITTER: Chang CP
PROVIDER: S-EPMC8220833 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Background5-Lipoxygenase (5-LO) is a protein widely distributed in the central nervous system where it modulates amyloidosis and memory impairments in transgenic mouse models of Alzheimer's disease. However, no data are available as to whether 5-LO is elevated in human tauopathy or if it directly influences tau pathology in a relevant model of the disease.MethodsWe assayed 5-LO levels in brain samples from patients with tauopathy and transgenic tau mice, and we evaluated the effect of 5-LO pharmacologic inhibition on the phenotype of these mice.ResultsThe 5-LO protein is upregulated in human tauopathy and transgenic tau mice brains. Pharmacologic blockade of 5-LO in tau mice resulted in significant memory improvement, rescue of synaptic integrity and dysfunction, and reduction of tau pathology via a cdk5-dependent mechanism.ConclusionsThese results establish a key role of 5-LO in the development of the tau pathology phenotype and demonstrate it to be a novel viable therapeutic target for the pharmacologic treatment of human tauopathy.
| S-EPMC4529386 | biostudies-literature
Project description:Mitochondrial and metabolic dysfunction are often implicated in neurological disease, but effective mechanism-based therapies remain elusive. We performed a genome-scale forward genetic screen in a Drosophila model of tauopathy, a class of neurodegenerative disorders characterized by the accumulation of the protein tau, and identified manipulation of the B-vitamin biotin as a potential therapeutic approach in tauopathy. We show that tau transgenic flies have an innate biotin deficiency due to tau-mediated relaxation of chromatin and consequent aberrant expression of multiple biotin-related genes, disrupting both carboxylase and mitochondrial function. Biotin depletion alone causes mitochondrial pathology and neurodegeneration in both flies and human neurons, implicating mitochondrial dysfunction as a mechanism in biotin deficiency. Finally, carboxylase biotin levels are reduced in mammalian tauopathies, including brains of human Alzheimer's disease patients. These results provide insight into pathogenic mechanisms of human biotin deficiency, the resulting effects on neuronal health, and a potential therapeutic pathway in the treatment of tau-mediated neurotoxicity.
| S-EPMC7777082 | biostudies-literature
Project description:Equilibrative nucleoside transporters (ENTs), which facilitate cross-membrane transport of nucleosides and nucleoside-derived drugs, play an important role in the salvage pathways of nucleotide synthesis, cancer chemotherapy, and treatment for virus infections. Functional characterization of ENTs at the molecular level remains technically challenging and hence scant. In this study, we report successful purification and biochemical characterization of human equilibrative nucleoside transporter 1 (hENT1) in vitro. The HEK293F-derived, recombinant hENT1 is homogenous and functionally active in proteoliposome-based counter flow assays. hENT1 transports the substrate adenosine with a Km of 215 ± 34 µmol/L and a Vmax of 578 ± 23.4 nmol mg-1 min-1. Adenosine uptake by hENT1 is competitively inhibited by nitrobenzylmercaptopurine ribonucleoside (NBMPR), nucleosides, deoxynucleosides, and nucleoside-derived anti-cancer and anti-viral drugs. Binding of hENT1 to adenosine, deoxyadenosine, and adenine by isothermal titration calorimetry is in general agreement with results of the competitive inhibition assays. These results validate hENT1 as a bona fide target for potential drug target and serve as a useful basis for future biophysical and structural studies.
| S-EPMC5359181 | biostudies-literature
Project description:Equilibrative nucleoside transporter 2 (ENT2) is a bidirectional transporter embedded in the biological membrane and is ubiquitously found in most tissue and cell types. ENT2 mediates the uptake of purine and pyrimidine nucleosides and nucleobase besides transporting a variety of nucleoside-derived drugs, mostly in anticancer therapy. Since high expression of ENT2 has been correlated with advanced stages of different types of cancers, consequently, this has gained significant interest in the role of ENT2 as a potential therapeutic target. Furthermore, ENT2 plays critical roles in signaling pathway and cell cycle progression. Therefore, elucidating the physiological roles of ENT2 and its properties may contribute to a better understanding of ENT2 roles beyond their transportation mechanism. This review is aimed at highlighting the main roles of ENT2 and at providing a brief update on the recent research.
| S-EPMC7732376 | biostudies-literature
Project description:Alzheimer's disease (AD) is a devastating neurodegenerative disease causing irreversible cognitive decline in the elderly. There is no disease-modifying therapy for this condition and the mechanisms underpinning neuronal dysfunction and neurodegeneration are unclear. Compromised cytoskeletal integrity within neurons is reported in AD. This is believed to result from loss-of-function of the microtubule-associated protein tau, which becomes hyper-phosphorylated and deposits into neurofibrillary tangles in AD. We have developed a Drosophila model of tauopathy in which abnormal human tau mediates neuronal dysfunction characterised by microtubule destabilisation, axonal transport disruption, synaptic defects and behavioural impairments. Here we show that a microtubule-stabilising drug, NAPVSIPQ (NAP), prevents as well as reverses these phenotypes even after they have become established. Moreover, it does not alter abnormal tau levels indicating that it by-passes toxic tau altogether. Thus, microtubule stabilisation is a disease-modifying therapeutic strategy protecting against tau-mediated neuronal dysfunction, which holds great promise for tauopathies like AD.
| S-EPMC3690421 | biostudies-literature
Project description:The human equilibrative nucleoside transporters hENT1 and hENT2 (each with 456 residues) are 40% identical in amino acid sequence and contain 11 putative transmembrane helices. Both transport purine and pyrimidine nucleosides and are distinguished functionally by a difference in sensitivity to inhibition by nanomolar concentrations of nitrobenzylmercaptopurine ribonucleoside (NBMPR), hENT1 being NBMPR-sensitive. Previously, we used heterologous expression in Xenopus oocytes to demonstrate that recombinant hENT2 and its rat ortholog rENT2 also transport purine and pyrimidine bases, h/rENT2 representing the first identified mammalian nucleobase transporter proteins (Yao, S. Y., Ng, A. M., Vickers, M. F., Sundaram, M., Cass, C. E., Baldwin, S. A., and Young, J. D. (2002) J. Biol. Chem. 277, 24938-24948). The same study also revealed lower, but significant, transport of hypoxanthine by h/rENT1. In the present investigation, we have used the enhanced Xenopus oocyte expression vector pGEMHE to demonstrate that hENT1 additionally transports thymine and adenine and, to a lesser extent, uracil and guanine. Fluxes of hypoxanthine, thymine, and adenine by hENT1 were saturable and inhibited by NBMPR. Ratios of V(max) (pmol/oocyte · min(-1)):K(m) (mm), a measure of transport efficiency, were 86, 177, and 120 for hypoxantine, thymine, and adenine, respectively, compared with 265 for uridine. Hypoxanthine influx was competitively inhibited by uridine, indicating common or overlapping nucleobase and nucleoside permeant binding pockets, and the anticancer nucleobase drugs 5-fluorouracil and 6-mercaptopurine were also transported. Nucleobase transport activity was absent from an engineered cysteine-less version hENT1 (hENT1C-) in which all 10 endogenous cysteine residues were mutated to serine. Site-directed mutagenesis identified Cys-414 in transmembrane helix 10 of hENT1 as the residue conferring nucleobase transport activity to the wild-type transporter.
| S-EPMC3173155 | biostudies-literature
Project description:Augustine (AUG) is a blood group system comprising four antigens: AUG1, AUG2 (Ata), and AUG4 are of very high frequency; AUG3 is of very low frequency. These antigens are located on ENT1, an equilibrative nucleoside transporter encoded by SLC19A1. AUG antibodies are of clinical relevance in blood transfusion and pregnancy: anti-AUG2 have caused haemolytic transfusion reactions; the only anti-AUG3 was associated with severe haemolytic disease of the fetus and newborn. ENT1 is present in almost all human tissues. It facilitates the transfer of purine and pyrimidine nucleosides and is responsible for the majority of adenosine transport across plasma membranes. Adenosine transport appears to be an important factor in the regulation of bone metabolism. The AUGnull phenotype (AUG:-1,-2,-3,-4) has been found in three siblings, who are homozygous for an inactivating splice-site mutation in SLC29A1. Although ENT1 is very likely to be absent from all cells in these three individuals, they were apparently healthy with normal lifestyles. However, they suffered frequent attacks of pseudogout, a form of arthritis, in various joints with multiple calcifications around their hand joints. Ectopic calcification in the hips, pubic symphysis, and lumbar discs was present in the propositus. The three AUGnull individuals had misshapen red cells with deregulated protein phosphorylation, but no anaemia or shortening of red cell lifespan. Defective in vitro erythropoiesis in the absence of ENT1 was confirmed by shRNA-mediated knockdown of ENT1 during in vitro erythropoiesis of CD34+ progenitor cells from individuals with normal ENT1. Nucleoside transporters, such as ENT1, are vital in the uptake of synthetic nucleoside analogue drugs, used in cancer and viral chemotherapy. It is feasible that the efficacy of these drugs would be compromised in patients with the extremely rare AUGnull phenotype.
| S-EPMC8832251 | biostudies-literature
Project description:The plant-derived cannabinoids delta9-tetrahydrocannabinol (THC) and cannabidiol (CBD) both have immunosuppressive effects; although some effects of THC are mediated by the CB2 receptor, CB2 binds CBD weakly. In examining the effects of THC and CBD on microglial proliferation, we found that these compounds potently inhibit [3H]thymidine incorporation into a murine microglial cell line with no effect on cell cycle. Treatment with THC and CBD decreased [3H]thymidine uptake into microglia, with IC50 values that match inhibition of [3H]thymidine incorporation into DNA. CBD and, less potently, THC decreased uptake of [3H]adenosine to a similar extent as [3H]thymidine in both murine microglia and RAW264.7 macrophages. Binding studies confirm that CBD binds to the equilibrative nucleoside transporter 1 with a Ki < 250 nM. Because adenosine agonists have antiinflammatory effects, and because uptake of adenosine is a primary mechanism of terminating adenosine signaling, we tested the hypothesis that CBD is immunosuppressive because it enhances endogenous adenosine signaling. In vivo treatment with a low dose of CBD decreases TNFalpha production in lipopolysaccharide-treated mice; this effect is reversed with an A2A adenosine receptor antagonist and abolished in A2A receptor knockout mice. These studies demonstrate that CBD has the ability to enhance adenosine signaling through inhibition of uptake and provide a non-cannabinoid receptor mechanism by which CBD can decrease inflammation.
| S-EPMC1472541 | biostudies-other
Project description:Immunosuppression by adenosine is an important cancer immune checkpoint. Extracellular adenosine signals through specific receptors and can be transported across the cell membrane through nucleoside transporters. While adenosine receptors are well-known to regulate tumor immunity, the impact of adenosine transporters remains unexplored. In this study, we investigated the effect on tumor immunity of equilibrative nucleoside transporter-1 (ENT1), the major regulator of extracellular adenosine concentrations. Using selective inhibitors and gene targeting approaches, we demonstrated that blocking or deleting host ENT1 significantly enhanced CD8+ T cell-dependent anti-tumor responses. Tumors inoculated into ENT1-deficient mice showed increased infiltration of effector CD8+ T cells with an enhanced cytotoxic transcriptomic profile and significant upregulation of granzyme B, IFN-g, IL-2, TNF-a and CXCL10. ENT1-deficiency was further associated with decreased tumor-infiltrating T regulatory cells and CD206high macrophages, and suppressed CCL17 production. ENT1-deficiency notably potentiated the therapeutic activity of PD-1 blockade. T cells upregulated ENT1 upon activation, and blocking ENT1 enhanced their function when co-cultured with cognate antigen/HLA-matched melanoma cells. Mechanistically, ENT1-mediated adenosine uptake inhibited the activity of phosphoribosylpyrophosphate synthetase (PRPS) in activated T cells, thereby suppressing production of uridine 5′-monophosphate (UMP) and its derivatives required for DNA and RNA synthesis. In summary, our study identified ENT1-mediated adenosine uptake as an important and previously unappreciated mechanism of adenosine-mediated immunosuppression via pyrimidine starvation that can be targeted to enhance antitumor T cell responses.
2024-10-13 | GSE279144 | GEO
Project description:Equilibrative nucleoside transporters (ENTs) are polytopic membrane transporters responsible for the translocation of nucleosides, nucleobases-to a lesser extent-and nucleoside analog therapeutics across cellular membranes. ENTs function in a diffusion controlled bidirectional manner and are thought to utilize an alternating access transport mechanism. However, a detailed understanding of ENT function at the molecular level has remained elusive. ScENT1 (formerly known as Function Unknown Now 26 or FUN26) is the only known ENT ortholog endogenously expressed in S. cerevisiae, and a proteoliposome assay system was used to study homogenously overexpressed and purified ScENT1 (wildtype relative to L390A and F249I mutants). L390 and F249 are highly conserved residues and were found to alter transporter function. L390A produced a reduction of mean transport activity while F249I increased mean substrate translocation relative to wildtype protein. However, both mutations resulted in transport of UTP-a novel gain of function for any ENT. These residues were then mapped onto an ab initio model of FUN26 which suggests they function in substrate translocation (L390) or cytoplasmic gating (F249). Furthermore, wildtype, L390A, and F249I were found to be sensitive to the presence of alcohols. Ethanol attenuated ScENT1-mediated transport of uridine by ~50%. These findings further demonstrate functional similarities between ScENT1 and human ENT isoforms and support identification of FUN26 as ScENT1, the first ENT isoform in S. cerevisiae.
| S-EPMC6017673 | biostudies-literature