TCDB is operated by the Saier Lab Bioinformatics Group

2.A.57 The Equilibrative Nucleoside Transporter (ENT) Family

Several members of the ENT family (Pfam CLN3) have been functionally characterized (Engel et al., 2004Griffiths et al., 1997b; Mäser et al., 1999; Sundaram et al., 1998; Vasudevan et al., 1998). The hENT1 is of human placental origin, is 456 amino acyl residues long and possesses 11 TMSs. It has an N-terminal mitochondrial targetting sequence and is expressed in the mitochondria and other organelles of many human tissues. Homologues have been sequenced from yeast, protozoa, plants, nematodes and mammals.  Most characterized plant (and probably lower eukaryotic) ENTs act in a concentrative manner, defying their name (Girke et al. 2015). C. elegans possesses at least five such homologues. Among these are the two smaller nucleolar ''''delayed early response'''' gene products, HNP36, sequenced from humans and mice (Williams and Lanahan, 1995). The hENT1 and rENT1 proteins appear to exhibit broad specificity for purine and pyrimidine nucleosides and cytotoxic nucleoside analogues used in cancer and viral chemotherapy. Some are sensitive and others are insensitive to inhibition by nitrobenzyl thioinosine. hENT2 has higher affinity for adenosine, inosine and hypoxanthine than hENT1 but lower affinity for other nucleosides. Both human and rat isoforms of hENT1 are cell surface and organellar localized being found in mitochondria, nuclear envelopes and lysosomes. One, PMAT (TC #2.A.57.1.5), transports monoamines, probably by an H+ symport mechanism.

Nucleoside transporters have been identified in Trypanosoma brucei and Leishmania donovani. They transport adenosine and probably other nucleosides and nucleobases as well as several drugs. When reconstituted in yeast, one (called TbAT1) catalyzes adenosine uptake and confers susceptibility to melaminophenyl arsenicals. Tyrpanocide drug-resistant tyrpanosomes have a mutated TbAT1 gene. These protozoan proteins are 460-500 residues long and exhibit 10 putative TMSs. The three Leishmania donovani paralogues (NT1.1, NT1.2 and NT2) are all electrogenic proton symporters (Stein et al., 2003).

The 7 known human nucleosides transporters (hNTs) exhibit varying permeant selectivities and are found into 2 protein families: the solute carrier (SLC) 29 (SLC29A1, SLC29A2, SLC29A3, SLC29A4) and SLC28 (SLC28A1, SLC28A2, SLC28A3) proteins, otherwise known, respectively, as the human equilibrative NTs (hENTs, hENT1, hENT2, hENT3, hENT4) and human concentrative NTs (hCNTs, hCNT1, hCNT2, hCNT3) (Elwi et al., 2006). The well characterized hENTs (hENT1 and hENT2) are bidirectional facilitative diffusion transporters in plasma membranes; hENT3 and hENT4 are much less well known, although hENT3, found in lysosomal membranes, transports nucleosides and is pH dependent.  hENT4-PMAT is a H+/adenosine cotransporter as well as a monoamine-organic cation transporter. The 3 hCNTs are unidirectional secondary active Na+/nucleoside cotransporters. In renal epithelial cells, hCNT1, hCNT2, and hCNT3, at apical membranes, and hENT1 and hENT2 at basolateral membranes, apparently work in concert to mediate reabsorption of nucleosides from lumen to blood, driven by Na+ gradients. Secretion of some physiological nucleosides, therapeutic nucleoside analog drugs, and nucleotide metabolites of therapeutic nucleoside and nucleobase drugs likely occurs through various xenobiotic transporters in renal epithelia, including organic cation transporters, organic anion transporters, multidrug resistance related proteins, and multidrug resistance proteins. Mounting evidence suggests that hENT1 may have a presence at both apical and basolateral membranes of renal epithelia, and thus may participate in both selective secretory and reabsorptive fluxes of nucleosides (Elwi et al., 2006).

Juvenile neuronal ceroid lipofuscinosis (JNCL) is a fatal childhood-onset neurodegenerative disorder caused by mutations in ceroid lipofuscinosis neuronal-3 (CLN3), a transmembrane protein of unresolved function. There may be blood-brain barrier (BBB) defects in JNCL. Cln3 is expressed in mouse brain endothelium. Tecedor et al. 2013 showed that CLN3 is necessary for normal trafficking of the microdomain-associated proteins caveolin-1, syntaxin-6, and multidrug resistance protein 1 (MDR1) in brain endothelial cells. CLN3-null cells have reduced caveolae, and impaired caveolae- and MDR1-related functions including endocytosis, drug efflux, and cell volume regulation. They also detected an abnormal blood-brain barrier response to osmotic stress in vivo and proposed that CLN3 facilitates golgi-to-plasma membrane transport of microdomain-associated proteins. 

The best-characterized members of the human Ent family, hENT1 and hENT2, possess similar broad permeant selectivities for purine and pyrimidine nucleosides, but hENT2 also efficiently transports nucleobases. hENT3 has a similar broad permeant selectivity for nucleosides and nucleobases and appears to function in intracellular membranes, including lysosomes. hENT4 is uniquely selective for adenosine, and also transports a variety of organic cations. hENT3 and hENT4 are pH sensitive and optimally active under acidic conditions. ENTs, including those in parasitic protozoa, function in nucleoside and nucleobase uptake for salvage pathways of nucleotide synthesis and, in humans, are also responsible for the cellular uptake of nucleoside analogues used in the treatment of cancers and viral diseases. By regulating the concentration of adenosine available to cell surface receptors, mammalian ENTs additionally influence physiological processes ranging from cardiovascular activity to neurotransmission (Young et al. 2008).

The generalized transport reaction catalyzed by well characterized ENT family members is:

Nucleoside (out) → Nucleoside (in).

 

This family belongs to the: MFS Superfamily.

References associated with 2.A.57 family:

and Arendt CS. (2013). Crithidia fasciculata adenosine transporter 1 (CfAT1), a novel high-affinity equilibrative nucleoside transporter specific for adenosine. Mol Biochem Parasitol. 191(2):75-9. 24120444
Baldwin, S.A., S.Y. Yao, R.J. Hyde, A.M. Ng, S. Foppolo, K. Barnes, M.W. Ritzel, C.E. Cass, and J.D. Young. (2005). Functional characterization of novel human and mouse equilibrative nucleoside transporters (hENT3 and mENT3) located in intracellular membranes. J. Biol. Chem. 280: 15880-15887. 15701636
Burchmore, R.J.S., L.J.M. Wallace, D. Candlish, M.I. Al-Salabi, P.R. Beal, M.P. Barrett, S.A. Baldwin, and H.P. de Koning. (2003). Cloning, heterologous expression, and in situ characterization of the first high affinity nucleobase transporter from a protozoan. J. Biol. Chem. 278: 23502-23507. 12707261
Carter N.S., M.E. Drew, M. Sanchez, G. Vasudevan, S.M. Landfear, and B. Ullman. (2000). Cloning of a novel inosine-guanosine transporter gene from Leishmania donovani by functional rescue of a transport-deficient mutant. J. Biol. Chem. 275: 20935-20941. 10783393
Carter, N.S., C. Ben Mamoun, W. Liu, E.O. Silva, S.M. Landfear, D.E. Goldberg, and B. Ullman. (2000). Isolation and functional characterization of the PfNT1 nucleoside transporter gene from Plasmodium falciparum. J. Biol. Chem. 275: 10683-10691. 10744765
Crawford, C.R., D.H. Patel, C. Naeve, and J.A. Belt. (1998). Cloning of the human equilibrative, nitrobenzylmercaptopurine riboside (NBMPR)-insensitive nucleoside transporter ei by functional expression in a transport-deficient cell line. J. Biol. Chem. 273: 5288-5293. 9478986
Downie, M.J., K. El Bissati, A.M. Bobenchik, L. Nic Lochlainn, A. Amerik, R. Zufferey, K. Kirk, and C. Ben Mamoun. (2010). PfNT2, a permease of the equilibrative nucleoside transporter family in the endoplasmic reticulum of Plasmodium falciparum. J. Biol. Chem. 285: 20827-20833. 20439460
Downie, M.J., K.J. Saliba, S.M. Howitt, S. Bröer, and K. Kirk. (2006). Transport of nucleosides across the Plasmodium falciparum parasite plasma membrane has characteristics of PfENT1. Mol. Microbiol. 60: 738-748. 16629674
Elwi, A.N., V.L. Damaraju, S.A. Baldwin, J.D. Young, M.B. Sawyer, and C.E. Cass. (2006). Renal nucleoside transporters: physiological and clinical implications. Biochem Cell Biol 84: 844-58. 17215872
Engel, K., M. Zhou, and J. Wang. (2004). Identification and characterization of a novel monoamine transporter in the human brain. J. Biol. Chem. 279: 50042-50049. 15448143
Farooq, M., R.M. Moustafa, A. Fujimoto, H. Fujikawa, O. Abbas, A.G. Kibbi, M. Kurban, and Y. Shimomura. (2012). Identification of Two Novel Mutations in SLC29A3 Encoding an Equilibrative Nucleoside Transporter (hENT3) in Two Distinct Syrian Families with H Syndrome: Expression Studies of SLC29A3 (hENT3) in Human Skin. Dermatology 224: 277-284. 22653152
Girke C., Arutyunova E., Syed M., Traub M., Mohlmann T. and Lemieux MJ. (2015). High yield expression and purification of equilibrative nucleoside transporter 7 (ENT7) from Arabidopsis thaliana. Biochim Biophys Acta. 1850(9):1921-9. 26080001
Griffiths, M., N. Beaumont, S.Y.M. Yao, M. Sundaram, C.E. Boumah, A. Davies, F.Y.P. Kwong, I. Coe, C.E. Cass, J.D. Young, and S.A. Baldwin. (1997). Cloning of a human nucleoside transporter implicated in the cellular uptake of adenosine and chemotherapeutic drugs. Nature Med. 3: 89-93. 8986748
Griffiths, M., S.Y.M. Yao, F. Abidi, S.E.V. Phillips, C.E. Cass, J.D. Young, and S.A. Baldwin. (1997b). Molecular cloning and characterization of a nitrobenzylthioinosine-insensitive (ei) equilibrative nucleoside transporter from human placenta. Biochem. J. 328: 739-743. 9396714
Ho HT., Xia L. and Wang J. (2012). Residue Ile89 in human plasma membrane monoamine transporter influences its organic cation transport activity and sensitivity to inhibition by dilazep. Biochem Pharmacol. 84(3):383-90. 22562044
Hsu, C.L., W. Lin, D. Seshasayee, Y.H. Chen, X. Ding, Z. Lin, E. Suto, Z. Huang, W.P. Lee, H. Park, M. Xu, M. Sun, L. Rangell, J.L. Lutman, S. Ulufatu, E. Stefanich, C. Chalouni, M. Sagolla, L. Diehl, P. Fielder, B. Dean, M. Balazs, and F. Martin. (2012). Equilibrative nucleoside transporter 3 deficiency perturbs lysosome function and macrophage homeostasis. Science 335: 89-92. 22174130
Klein, D.M., K.K. Evans, R.N. Hardwick, W.H. Dantzler, S.H. Wright, and N.J. Cherrington. (2013). Basolateral uptake of nucleosides by Sertoli cells is mediated primarily by equilibrative nucleoside transporter 1. J Pharmacol Exp Ther 346: 121-129. 23639800
Kobayashi, M., E. Yamato, K. Tanabe, F. Tashiro, S. Miyazaki, and J. Miyazaki. (2016). Functional Analysis of Novel Candidate Regulators of Insulin Secretion in the MIN6 Mouse Pancreatic β Cell Line. PLoS One 11: e0151927. 26986842
Lee, E.W., Y. Lai, H. Zhang, and J.D. Unadkat. (2006). Identification of the mitochondrial targeting signal of the human equilibrative nucleoside transporter 1 (hENT1): implications for interspecies differences in mitochondrial toxicity of fialuridine. J. Biol. Chem. 281: 16700-16706. 16595656
Lepist, E.I., V.L. Damaraju, J. Zhang, W.P. Gati, S.Y. Yao, K.M. Smith, E. Karpinski, J.D. Young, K.H. Leung, and C.E. Cass. (2013). Transport of A1 adenosine receptor agonist tecadenoson by human and mouse nucleoside transporters: evidence for blood-brain barrier transport by murine equilibrative nucleoside transporter 1 mENT1. Drug Metab Dispos 41: 916-922. 23388705
Li, X., J. Zhang, Z. Zhang, and C. Zhou. (2010). [Relationship between single nucleotide polymorphism of the equilibrative nucleoside transporter ENT3 and susceptibility to lung cancer]. Zhongguo Fei Ai Za Zhi 13: 458-463. 20677642
Liu, J.W., N. Si, L.Q. Wang, T. Shen, X.J. Zeng, X. Zhang, and D.L. Ma. (2015). Identification of a novel mutation in solute carrier family 29, member 3 in a Chinese patient with H syndrome. Chin Med J (Engl) 128: 1336-1339. 25963354
Mäser, P., C. Sütterlin, A. Kralli, and R. Kaminsky. (1999). A nucleoside transporter from Tyrpanosoma brucei involved in drug resistance. Science 285: 242-244. 10398598
Nugent, T., S.E. Mole, and D.T. Jones. (2008). The transmembrane topology of Batten disease protein CLN3 determined by consensus computational prediction constrained by experimental data. FEBS Lett. 582: 1019-1024. 18314010
Orlandi, A., M.A. Calegari, M. Martini, A. Cocomazzi, C. Bagalà, G. Indellicati, V. Zurlo, M. Basso, A. Cassano, L.M. Larocca, and C. Barone. (2016). Gemcitabine versus FOLFIRINOX in patients with advanced pancreatic adenocarcinoma hENT1-positive: everything was not too bad back when everything seemed worse. Clin Transl Oncol. [Epub: Ahead of Print] 26742940
Ortiz, D., M.A. Sanchez, S. Pierce, T. Herrmann, N. Kimblin, H.G. Archie Bouwer, and S.M. Landfear. (2007). Molecular genetic analysis of purine nucleobase transport in Leishmania major. Mol Microbiol. 64: 1228-1243. 17542917
Paproski, R.J., F. Visser, J. Zhang, T. Tackaberry, V. Damaraju, S.A. Baldwin, J.D. Young, and C.E. Cass. (2008). Mutation of Trp(29) of human equilibrative nucleoside transporter 1 alters affinity for coronary vasodilator drugs and nucleoside selectivity. Biochem. J. 414: 291-300. 18462193
Rager, N., C. Ben Mamoun, N.S. Carter, D.E. Goldberg, and B. Ullman. (2001). Localization of the Plasmodium falciparum PfNT1 nucleoside transporter to the parasite plasma membrane. J. Biol. Chem. 276: 41095-41099. 11682491
Riegelhaupt, P.M., I.J. Frame, and M.H. Akabas. (2010). Transmembrane segment 11 appears to line the purine permeation pathway of the Plasmodium falciparum equilibrative nucleoside transporter 1 (PfENT1). J. Biol. Chem. 285: 17001-17010. 20335165
Sanchez, M.A., R. Tryon, J. Green, I. Boor, and S.M. Landfear. (2002). Six related nucleoside/nucleobase transporters from Trypanosoma brucei exhibit distinct biochemical functions. J. Biol. Chem. 277: 21499-21504. 11937511
Stein, A., G. Vaseduvan, N.S. Carter, B. Ullman, S.M. Landfear, and M.P. Kavanaugh. (2003). Equilibrative nucleoside transporter family members from Leishmania donovani are electrogenic proton symporters. J. Biol. Chem. 278: 35127-35134. 12835315
Sundaram, M., S.Y. Yao, A.M. Ng, M. Griffiths, C.E. Cass, S.A. Baldwin, and J.D. Young. (1998). Chimeric constructs between human and rat equilibrative nucleoside transporters (hENT1 and rENT1) reveal hENT1 structural domains interacting with coronary vasoactive drugs. J. Biol. Chem. 273: 21519-21525. 9705281
Tecedor, L., C.S. Stein, M.L. Schultz, H. Farwanah, K. Sandhoff, and B.L. Davidson. (2013). CLN3 Loss Disturbs Membrane Microdomain Properties and Protein Transport in Brain Endothelial Cells. J. Neurosci. 33: 18065-18079. 24227717
Traub, M., M. Flörchinger, J. Piecuch, H.H. Kunz, A. Weise-Steinmetz, J.W. Deitmer, H. Ekkehard Neuhaus, and T. Möhlmann. (2007). The fluorouridine insensitive 1 (fur1) mutant is defective in equilibrative nucleoside transporter 3 (ENT3), and thus represents an important pyrimidine nucleoside uptake system in Arabidopsis thaliana. Plant J. 49: 855-864. 17253988
Uusi-Rauva, K., K. Luiro, K. Tanhuanpää, O. Kopra, P. Martín-Vasallo, A. Kyttälä, and A. Jalanko. (2008). Novel interactions of CLN3 protein link Batten disease to dysregulation of fodrin-Na+, K+ ATPase complex. Exp Cell Res 314: 2895-2905. 18621045
Valdés, R., J. Elferich, U. Shinde, and S.M. Landfear. (2014). Identification of the Intracellular Gate for a Member of the Equilibrative Nucleoside Transporter (ENT) Family. J. Biol. Chem. 289: 8799-8809. 24497645
Valdés, R., U. Shinde, and S.M. Landfear. (2012). Cysteine Cross-linking Defines the Extracellular Gate for the Leishmania donovani Nucleoside Transporter 1.1 (LdNT1.1). J. Biol. Chem. 287: 44036-44045. 23150661
Valdés, R., W. Liu, B. Ullman, and S.M. Landfear. (2006). Comprehensive examination of charged intramembrane residues in a nucleoside transporter. J. Biol. Chem. 281: 22647-22655. 16769726
Vasudevan, G., N.S. Carter, M.E. Drew, S.M. Beverley, M.A. Sanchez, A. Seyfang, B. Ullman, and S.M. Landfear. (1998). Cloning of Leishmania nucleoside transporter genes by rescue of a transport-deficient mutant. Proc. Natl. Acad. Sci. USA 95: 9873-9878. 9707568
Vickers, M.F., S.Y.M. Yao, S.A. Baldwin, J.D. Young, and C.E. Cass. (2000). Nucleoside transporter proteins of Saccharomyces cerevisiae: demonstration of a transporter (FUI1) with high uridine selectivity in plasma membranes and a transporter (FUN26) with broad nucleoside selectivity in intracellular membranes. J. Biol. Chem. 275: 25931-25938. 10827169
Visser, F., L. Sun, V. Damaraju, T. Tackaberry, Y. Peng, M.J. Robins, S.A. Baldwin, J.D. Young, C.E. Cass. (2007). Residues 334 and 338 in transmembrane segment 8 of human equilibrative nucleoside transporter 1 are important determinants of inhibitor sensitivity, protein folding, and catalytic turnover. J. Biol. Chem. 282: 14148-14157. 17379602
Ward, J.L., A. Sherali, Z. Mo, and C. Tse. (2000). Kinetic and pharmacological properties of cloned human equilibrative nucleoside transporters, ENT1 and ENT2, stably expressed in nucleoside transporter-deficient PK15 cells. J. Biol. Chem. 275: 8375-8381. 10722669
Williams, J.B. and A.A. Lanahan. (1995). A mammalian delayed early response gene encodes HNP36, a novel conserved nucleolar protein. Biochem. Biophys. Res. Commun. 213: 325-333. 7639753
Xia, L., K. Engel, M. Zhou, and J. Wang. (2007). Membrane localization and pH-dependent transport of a newly cloned organic cation transporter (PMAT) in kidney cells. Am. J. Physiol. Renal Physiol 292: F682-690. 17018840
Yao, S.Y.M., A.M.L. Ng, W.R. Muzyka, M. Griffiths, C.E. Cass, S.A. Baldwin, and J.D. Young. (1997). Molecular cloning and functional characterization of nitrobenzylthioinosine (NBMPR)-sensitive (es) and NBMPR-insensitive (ei) equilibrative nucleoside transporter proteins (rENT1 and rENT2) from rat tissues. J. Biol. Chem. 272: 28423-28430. 9353301
Young, J.D., S.Y. Yao, L. Sun, C.E. Cass, and S.A. Baldwin. (2008). Human equilibrative nucleoside transporter (ENT) family of nucleoside and nucleobase transporter proteins. Xenobiotica 38: 995-1021. 18668437