Arginine transport in human erythroid cells: discrimination of CAT1 and 4F2hc/y+LAT2 roles

Since arginine metabolites, such as nitric oxide and polyamines, influence the expression of genes involved in erythroid differentiation, the transport of the cationic amino acid may play an important role in erythroid cells. However, available data only concern the presence in these cells of CAT1 transporter (system y⁺), while no information exists on the role of the heterodimeric transporters of system y⁺L (4F2hc/y⁺LAT1 and 4F2hc/y⁺LAT2) which operates transmembrane arginine fluxes cis-inhibited by neutral amino acids in the presence of sodium. Using erythroleukemia K562 cells and normal erythroid precursors, we demonstrate here that arginine transport in human erythroid cells is due to the additive contributions of a leucine-sensitive and leucine-insensitive component. In both cell types, leucine inhibition of arginine influx is much less evident in the absence of sodium, a hallmark of system y⁺L. In K562 cells, N-ethylmaleimide, a known inhibitor of CAT transporters (system y⁺), suppresses only a fraction of arginine influx corresponding to leucine-insensitive uptake. Moreover, in Xenopus oocytes coexpressing 4F2hc and y⁺LAT2, leucine exerts a marked inhibition of arginine transport, partially dependent on sodium, while no inhibition is seen in oocytes expressing CAT1. Lastly, silencing of SLC7A6, the gene for y⁺LAT2, lowers arginine transport and doubles the intracellular content of the cationic amino acid in K562 cells. We conclude that arginine transport in human erythroid cells is due to both system y⁺ (CAT1 transporter) and system y⁺L (4F2hc/y⁺LAT2 isoform), which mainly contribute, respectively, to the influx and to the efflux of the cationic amino acid.     

Kidney amino acid transport

Near complete reabsorption of filtered amino acids is a main specialized transport function of the kidney proximal tubule. This evolutionary conserved task is carried out by a subset of luminal and basolateral transporters that together form the transcellular amino acid transport machinery similar to that of small intestine. A number of other amino acid transporters expressed in the basolateral membrane of proximal kidney tubule cells subserve either specialized metabolic functions, such as the production of ammonium, or are part of the cellular housekeeping equipment. A new finding is that the luminal Na⁺-dependent neutral amino acid transporters of the SLC6 family require an associated protein for their surface expression as shown for the Hartnup transporter B⁰AT1 (SLC6A19) and suggested for the l-proline transporter SIT1 (IMINO^B, SLC6A20) and for B⁰AT3 (XT2, SLC6A18). This accessory subunit called collectrin (TMEM27) is homologous to the transmembrane anchor region of the renin–angiotensin system enzyme ACE2 that we have shown to function in small intestine as associated subunit of the luminal SLC6 transporters B⁰AT1 and SIT1. Some mutations of B⁰AT1 differentially interact with these accessory subunits, providing an explanation for differential intestinal phenotypes among Hartnup patients. The basolateral efflux of numerous amino acids from kidney tubular cells is mediated by heteromeric amino acid transporters that function as obligatory exchangers. Thus, other transporters within the same membrane need to mediate the net efflux of exchange substrates, controlling thereby the net basolateral amino transport and thus the intracellular amino acid concentration.     

The molecular basis of neutral aminoacidurias

Recent success in the molecular cloning and identification of apical neutral amino acid transporters has shed a new light on inherited neutral amino acidurias, such as Hartnup disorder and Iminoglycinuria. Hartnup disorder is caused by mutations in the neutral amino acid transporter B(0) AT1 (SLC6A19). The transporter is found in kidney and intestine, where it is involved in the resorption of all neutral amino acids. The molecular defect underlying Iminoglycinuria has not yet been identified. However, two transporters, the proton amino acid transporter PAT1 (SLC36A1) and the IMINO transporter (SLC6A20) appear to play key roles in the resorption of glycine and proline. A model is presented, involving all three transporters that can explain the phenotypic variability of iminoglycinuria.     

Lysine transport in lactating rat mammary tissue: evidence for an interaction between cationic and neutral amino acids

The transport of lysine by the lactating rat mammary gland has been examined to determine whether there is an interaction between cationic and neutral amino acids. Lysine uptake was time dependent and unaffected by replacing Na⁺ with choline. In the presence of Na⁺, lysine influx was inhibited by cationic amino acids (arginine, homoarginine, ornithine and lysine) and by a range of neutral amino acids (methionine, glutamine, leucine, phenylalanine, alanine, asparagine, α-aminoisobutyric acid (AIB), 2-aminobicyclo [2,2,1] heptane-2-carboxylic acid (BCH), proline and tryptophan). Leucine and glutamine also inhibited lysine influx in the absence of Na⁺ but phenylalanine and proline did not. Lysine efflux from mammary tissue was trans-accelerated by various cationic amino acids (lysine, arginine, homoarginine and ornithine). In addition, leucine and glutamine were capable of trans-stimulating lysine efflux in the presence and absence of Na⁺. It appears that cationic and neutral amino acids stimulated lysine efflux at a single locus.     

The SLC38 family of sodium–amino acid co-transporters

Transporters of the SLC38 family are found in all cell types of the body. They mediate Na⁺-dependent net uptake and efflux of small neutral amino acids. As a result they are particularly expressed in cells that grow actively, or in cells that carry out significant amino acid metabolism, such as liver, kidney and brain. SLC38 transporters occur in membranes that face intercellular space or blood vessels, but do not occur in the apical membrane of absorptive epithelia. In the placenta, they play a significant role in the transfer of amino acids to the foetus. Members of the SLC38 family are highly regulated in response to amino acid depletion, hypertonicity and hormonal stimuli. SLC38 transporters play an important role in amino acid signalling and have been proposed to act as transceptors independent of their transport function. The structure of SLC38 transporters is characterised by the 5?+?5 inverted repeat fold, which is observed in a wide variety of transport proteins.     

Effects of glutamate and aspartate on growth performance,serum amino acids,and amino acid transporters in piglets

This study mainly investigated the effects of different dietary levels of glutamate (Glu) and aspartate (Asp) on growth performance, blood amino acids, and amino acid transporters in piglets. Forty-two healthy piglets were randomly divided into six groups (n?=?7): a control group in which piglets were fed 2.9% Glu and 1.5% Asp and other groups in which piglets received 1.3% or 1.7% Asp and 2.6%, 3.2%, or 3.5% Glu for 21 days. Growth performance, serum amino acid profiles from the mesenteric vein, portal vein, and anterior vena cava, and amino acid transporters in the liver were determined. The results showed that lower doses of Asp promoted growth and enhanced the amino acids, while high doses of Asp and Glu reduced growth and the amino acid pool in piglets (P?P?P?相似文献   

In vitro evidence of involvement of the epithelial y transporter in β-defensin production on the ocular surface

To analyse the hypothesis as to whether there is a functional relationship between human cationic amino acid transporters (hCATs, y⁺ transporter, the main transporter of l-arginine and l-lysine) and human β-defensin (important components of immune function) production on the ocular surface, arginase and nitrate monoxide synthase (NOS), enzymes that compete for l-arginine, were inhibited by norNOHA (N(omega)-hydroxy-nor-l-arginine) and/or L-NAME (NG-nitro-l-arginine methyl ester) in cultured human corneal epithelial cells. In addition, the transport activity of hCAT proteins was inhibited or activated through α-tocopherol or PMA (phorbol myristate acetate), respectively. Concentrations of the human inducible β-defensins (hBD) 2 and 3 were determined by ELISA experiments. The basic expression of hBD3 in non-stimulated HCE cells significantly exceeded that of hBD2. Both β-defensins also differed as to how readily their excretion could be stimulated. HBD2 excretion rate was 3.5 time more by L-NAME, whereas norNOHA had no effect. In contrast, hBD3 excretion was increased by norNOHA by a factor of 1.5 but L-NAME alone had no effect. The excretion of both β-defensins was increased 3- and 6-fold by combined administration of L-NAME, norNOHA and interleukin (IL)-1β. Administration of α-tocopherol increased hBD2 excretion twofold. No effect was observed for hBD3. With PMA, on the other hand, a reduction in secretion for both β-defensins was observed. These in vitro findings provide evidence of a functional association between CAT proteins and β-defensins 2 and 3 opening up a new field of research with pharmacological perspectives for treatment of inflammatory diseases such as keratitis or dry eye disease.     

Recycling of aromatic amino acids via TAT1 allows efflux of neutral amino acids via LAT2-4F2hc exchanger

The rate of amino acid efflux from individual cells needs to be adapted to cellular demands and plays a central role for the control of extracellular amino acid homeostasis. A particular example of such an outward amino acid transport is the basolateral efflux from transporting epithelial cells located in the small intestine and kidney proximal tubule. Because LAT2-4F2hc (Slc7a8–Slc3a2), the best known basolateral neutral amino acid transporter of these epithelial cells, functions as an obligatory exchanger, we tested whether TAT1 (Slc16a10), the aromatic amino-acid facilitated diffusion transporter, might allow amino acid efflux via this exchanger by recycling its influx substrates. In this study, we show by immunofluorescence that TAT1 and LAT2 indeed colocalize in the early kidney proximal tubule. Using the Xenopus laevis oocytes expression system, we show that l-glutamine is released from oocytes into an amino-acid-free medium only when both transporters are coexpressed. High-performance liquid chromatography analysis reveals that several other neutral amino acids are released as well. The transport function of both TAT1 and LAT2-4F2hc is necessary for this efflux, as coexpression of functionally inactive but surface-expressed mutants is ineffective. Based on negative results of coimmunoprecipitation and crosslinking experiments, the physical interaction of these transporters does not appear to be required. Furthermore, replacement of TAT1 or LAT2-4F2hc by the facilitated diffusion transporter LAT4 or the obligatory exchanger LAT1, respectively, supports similar functional cooperation. Taken together, the results suggest that the aromatic amino acid diffusion pathway TAT1 can control neutral amino acid efflux via neighboring exchanger LAT2-4F2hc, by recycling its aromatic influx substrates.     

NBCe1 as a model carrier for understanding the structure–function properties of Na+-coupled SLC4 transporters in health and disease

SLC4 transporters are membrane proteins that in general mediate the coupled transport of bicarbonate (carbonate) and share amino acid sequence homology. These proteins differ as to whether they also transport Na⁺ and/or Cl^?, in addition to their charge transport stoichiometry, membrane targeting, substrate affinities, developmental expression, regulatory motifs, and protein–protein interactions. These differences account in part for the fact that functionally, SLC4 transporters have various physiological roles in mammals including transepithelial bicarbonate transport, intracellular pH regulation, transport of Na⁺ and/or Cl^?, and possibly water. Bicarbonate transport is not unique to the SLC4 family since the structurally unrelated SLC26 family has at least three proteins that mediate anion exchange. The present review focuses on the first of the sodium-dependent SLC4 transporters that was identified whose structure has been most extensively studied: the electrogenic Na⁺-base cotransporter NBCe1. Mutations in NBCe1 cause proximal renal tubular acidosis (pRTA) with neurologic and ophthalmologic extrarenal manifestations. Recent studies have characterized the important structure–function properties of the transporter and how they are perturbed as a result of mutations that cause pRTA. It has become increasingly apparent that the structure of NBCe1 differs in several key features from the SLC4 Cl^?–HCO₃ ^? exchanger AE1 whose structural properties have been well-studied. In this review, the structure–function properties and regulation of NBCe1 will be highlighted, and its role in health and disease will be reviewed in detail.     

Activation of cationic amino acid transport through system y+ correlates with expression of the T-cell early antigen gene in human lymphocytes

Lysine influx (2 M) into activated human B and T lymphocytes through transport systems y⁺ and y⁺L was measured. In activated T cells very substantial activation of system y⁺ was detected; system y⁺L was also activated in these cells but with a slower time course and to a smaller extent. No stimulation of either system was found in activated B cells. The time course of activation of system y⁺ precisely matched the expression of the T cell early antigen gene described by MacLeod et al, 1990. The functional significance of these observations with respect to L-arginine transport and nitric oxide synthesis in activated T lymphocytes is discussed.     

精氨酸转运功能异常相关基因SLC7A2的研究

精氨酸是细胞和有机体生存所必需的,其转运主要依赖于Na+非依赖性、pH不敏感的y+型转运体.其中,SLC7A2基因编码的阳离子氨基酸转运体2(CAT2)是介导精氨酸转运的重要转运体之一.本文就SLC7A2基因及CAT2的功能及此转运体功能异常与疾病的关系作一综述.     

The SLC22 drug transporter family

The SLC22 family comprises organic cation transporters (OCTs), zwitterion/cation transporters (OCTNs), and organic anion transporters (OATs). These transporters contain 12 predicted -helical transmembrane domains (TMDs) and one large extracellular loop between TMDs 1 and 2. Transporters of the SLC22 family function in different ways: (1) as uniporters that mediate facilitated diffusion in either direction (OCTs), (2) as anion exchangers (OAT1, OAT3 and URAT1), and (3) as Na⁺/l-carnitine cotransporter (OCTN2). They participate in the absorption and/or excretion of drugs, xenobiotics, and endogenous compounds in intestine, liver and/or kidney, and perform homeostatic functions in brain and heart. The endogenous substrates include monoamine neurotransmitters, choline, l-carnitine, -ketoglutarate, cAMP, cGMP, prostaglandins, and urate. Defect mutations of transporters of the SLC22 family may cause specific diseases such as "primary systemic carnitine deficiency" or "idiopathic renal hypouricemia" or change drug absorption or excretion.     

Sodium-coupled neutral amino acid (System N/A) transporters of the SLC38 gene family

The sodium-coupled neutral amino acid transporters (SNAT) of the SLC38 gene family resemble the classically-described System A and System N transport activities in terms of their functional properties and patterns of regulation. Transport of small, aliphatic amino acids by System A subtypes (SNAT1, SNAT2, and SNAT4) is rheogenic and pH sensitive. The System N subtypes SNAT3 and SNAT5 also countertransport H(+), which may be key to their operation in reverse, and have narrower substrate profiles than do the System A subtypes. Glutamine emerges as a favored substrate throughout the family, except for SNAT4. The SLC38 transporters undoubtedly play many physiological roles including the transfer of glutamine from astrocyte to neuron in the CNS, ammonia detoxification and gluconeogenesis in the liver, and the renal response to acidosis. Probing their regulation has revealed additional roles, and recent work has considered SLC38 transporters as therapeutic targets in neoplasia.     

The SLC36 family: proton-coupled transporters for the absorption of selected amino acids from extracellular and intracellular proteolysis

Whilst Na(+) has replaced H(+) as a major transport driving force at the plasma membrane of animal cells, the evolutionarily older H(+)-driven systems persist on endomembranes and at the plasma membrane of specialized cells. The first member of the SLC36 family, present in both intracellular and plasma membranes, was identified independently as a lysosomal amino acid transporter (LYAAT1) responsible for the export of lysosomal proteolysis products into the cytosol and as a proton/amino acid transporter (PAT1) responsible for the absorption of amino acids in the gut. In addition to LYAAT1/PAT1, the family comprises another characterized member, PAT2, and two orphan transporters. Both PAT1 and PAT2 mediate 1:1 symport of protons and small neutral amino acids such as glycine, alanine, and proline. Their mRNAs are broadly and differentially expressed in mammalian tissues. The PAT1 protein localizes to lysosomes in brain neurons, but is also found in the apical membrane of intestinal epithelial cells with a role in the absorption of amino acids from luminal protein digestion. In both cases, protons supplied by the lysosomal H(+)-ATPase or by the acidic microclimate of the brush border membrane drive transport of the amino acids into the cytosol. The subcellular localization and physiological role of PAT2 have still to be determined. SLC36 transporters are related distantly to other proton-coupled amino acid transporters, such as the vesicular neurotransmitter transporter VIAAT/VGAT (SLC32) and system N transporters (SLC38 family).     

An amino acid transporter involved in gastric acid secretion

Gastric acid secretion is regulated by a variety of stimuli, in particular histamine and acetyl choline. In addition, dietary factors such as the acute intake of a protein-rich diet and the subsequent increase in serum amino acids can stimulate gastric acid secretion only through partially characterized pathways. Recently, we described in mouse stomach parietal cells the expression of the system L heteromeric amino acid transporter comprised of the LAT2-4F2hc dimer. Here we address the potential role of the system L amino acid transporter in gastric acid secretion by parietal cells in freshly isolated rat gastric glands. RT-PCR, western blotting and immunohistochemistry confirmed the expression of 4F2-LAT2 amino acid transporters in rat parietal cells. In addition, mRNA was detected for the B⁰AT1, ASCT2, and ATB(0+) amino acid transporters. Intracellular pH measurements in parietal cells showed histamine-induced and omeprazole-sensitive H⁺-extrusion which was enhanced by about 50% in the presence of glutamine or cysteine (1 mM), two substrates of system L amino acid transporters. BCH, a non-metabolizable substrate and a competitive inhibitor of system L amino acid transport, abolished the stimulation of acid secretion by glutamine or cysteine suggesting that this stimulation required the uptake of amino acids by system L. In the absence of histamine glutamine also stimulated H⁺-extrusion, whereas glutamate did not. Also, phenylalanine was effective in stimulating H⁺/K⁺-ATPase activity. Glutamine did not increase intracellular Ca²⁺ levels indicating that it did not act via the recently described amino acid modulated Ca²⁺-sensing receptor. These data suggest a novel role for heterodimeric amino acid transporters and may elucidate a pathway by which protein-rich diets stimulate gastric acid secretion.P. Kirchhoff and M.H. Dave contributed equally to this study and therefore share first authorship     

Exclusion of linkage between schizophrenia and the gene encoding a neutral amino acid glutamate/aspartate transporter,SLC1A5

An abnormality in glutamatergic function has been hypothesized as being of etiological importance in schizophrenia. Twenty-three multiplex English and Icelandic schizophrenia families were genotyped with a polymorphic dinucleotide repeat sequence in the 3′-untranslated region of the glutamate/aspartate transporter gene called SLC1A5. Using the lod and a model-free method of linkage analysis (MFLINK), no evidence of linkage between SLC1A5 and schizophrenia was found. Our results do not support the hypothesis that SLC1A5 gene mutations or allelic variants provide a major gene contribution to the etiology of schizophrenia. However, because of the likelihood of heterogeneity of linkage in schizophrenia, there is a case for testing other pedigrees for linkage to the SLC1A5 locus. The SLC1A5 locus is one of a complex family of genes encoding neutral amino acid transporter proteins and the genetic relation between these other loci and schizophrenia has not yet been established. Am. J. Med. Genet. 74:50–52, 1997. © 1997 Wiley-Liss, Inc.     

Induced arginine transport via cationic amino acid transporter‐1 is necessary for human T‐cell proliferation

Availability of the semiessential amino acid arginine is fundamental for the efficient function of human T lymphocytes. Tumor‐associated arginine deprivation, mainly induced by myeloid‐derived suppressor cells, is a central mechanism of tumor immune escape from T‐cell‐mediated antitumor immune responses. We thus assumed that transmembranous transport of arginine must be crucial for T‐cell function and studied which transporters are responsible for arginine influx into primary human T lymphocytes. Here, we show that activation via CD3 and CD28 induces arginine transport into primary human T cells. Both naïve and memory CD4⁺ T cells as well as CD8⁺ T cells specifically upregulated the human cationic amino acid transporter‐1 (hCAT‐1), with an enhanced and persistent expression under arginine starvation. When hCAT‐1 induction was suppressed via siRNA transfection, arginine uptake, and cellular proliferation were impaired. In summary, our results demonstrate that hCAT‐1 is a key component of efficient T‐cell activation and a novel potential target structure to modulate adaptive immune responses in tumor immunity or inflammation.     

The plant plasma membrane proton pump ATPase: a highly regulated P-type ATPase with multiple physiological roles

Around 40 P-type ATPases have been identified in Arabidopsis and rice, for which the genomes are known. None seems to exchange sodium and potassium, as does the animal Na⁺/K⁺-ATPase. Instead, plants, together with fungi, possess a proton pumping ATPase (H⁺-ATPase), which couples ATP hydrolysis to proton transport out of the cell, and so establishes an electrochemical gradient across the plasma membrane, which is dissipated by secondary transporters using protons in symport or antiport, as sodium is used in animal cells. Additional functions, such as stomata opening, cell growth, and intracellular pH homeostasis, have been proposed. Crystallographic data and homology modeling suggest that the H⁺-ATPase has a broadly similar structure to the other P-type ATPases but has an extended C-terminal region, which is involved in enzyme regulation. Phosphorylation of the penultimate residue, a Thr, and the subsequent binding of regulatory 14–3–3 proteins result in the formation of a dodecamer (six H⁺-ATPase and six 14–3–3 molecules) and enzyme activation. This type of regulation is unique to the P-type ATPase family. However, the recent identification of additional phosphorylated residues suggests further regulatory features.     

Effects of hexosamines and omega-3/omega-6 fatty acids on pH regulation by interleukin 1-treated isolated bovine articular chondrocytes

Previous work has shown that interleukin 1 (IL-1) increases the activity of acid extruders in articular chondrocytes, while the H⁺-adenosine triphosphatase (ATPase) inhibitor bafilomycin can prevent aggrecanase-mediated cartilage degradation. The H⁺ transport induced by IL-1 may therefore be required for proteinase activity. In the present study, the effects of hexosamines and fish oils on H⁺-ATPase activity have been characterised for isolated bovine articular chondrocytes. Cells isolated in the presence of IL-1 were acidified, and the fraction of acid extrusion mediated by Na⁺–H⁺ exchange and an H⁺-ATPase were determined using specific inhibitors. Exposure to IL-1 significantly enhanced both components of acid extrusion. Co-incubation with glucosamine or mannosamine attenuated the H⁺-ATPase fraction of efflux. The addition of glucosamine at 9 h after exposure to IL-1—when H⁺-ATPase activation is already apparent—was also able to abolish H⁺-ATPase activity, implying that hexosamines do not exert effects at the level of protein synthesis. Co-incubation with the glucose transport inhibitor phloretin elicited similar effects to the hexosamines, suggesting that modulation of adenosine triphosphate levels may underlie their effects on H⁺-ATPase function. The omega-3 fish oil linolenic acid but not the omega-6 fish oil linoleic acid reduced H⁺-ATPase activity to levels seen in IL-1-untreated cells, although total efflux remained elevated, as a result of an enhanced H⁺ leak. These observations support a model whereby IL-1 stimulates an H⁺-ATPase-dependent system, possibly involved in aggrecanase activation, which appears to be one of the target mechanisms interrupted by dietary supplements reported to have symptom-modifying effects on osteoarthritis.     

The SLC2 family of facilitated hexose and polyol transporters

The SLC2 family of glucose and polyol transporters comprises 13 members, the glucose transporters (GLUT) 1–12 and the H⁺-myo-inositol cotransporter (HMIT). These proteins all contain 12 transmembrane domains with both the amino and carboxy-terminal ends located on the cytoplasmic side of the plasma membrane and a N-linked oligosaccharide side-chain located either on the first or fifth extracellular loop. Based on sequence comparison, the GLUT isoforms can be grouped into three classes: class I comprises GLUT1–4; class II, GLUT6, 8, 10, and 12 and class III, GLUT5, 7, 9, 11 and HMIT. Despite their sequence similarity and the presence of class-specific signature sequences, these transporters carry various hexoses and HMIT is a H⁺/myo-inositol co-transporter. Furthermore, the substrate transported by some isoforms has not yet been identified. Tissue- and cell-specific expression of the well-characterized GLUT isoforms underlies their specific role in the control of whole-body glucose homeostasis. Numerous studies with transgenic or knockout mice indeed support an important role for these transporters in the control of glucose utilization, glucose storage and glucose sensing. Much remains to be learned about the transport functions of the recently discovered isoforms (GLUT6–13 and HMIT) and their physiological role in the metabolism of glucose, myo-inositol and perhaps other substrates.An erratum to this article can be found at