Localization of the high-affinity glutamate transporter EAAC1 in rat kidney

1997 ◽  
Vol 273 (6) ◽  
pp. F1023-F1029 ◽  
Author(s):  
Chairat Shayakul ◽  
Yoshikatsu Kanai ◽  
Wen-Sen Lee ◽  
Dennis Brown ◽  
Jeffrey D. Rothstein ◽  
...  
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Rat Kidney ◽  
Cell Volume Regulation ◽  
Glutamate Transporter ◽  
Transport Systems ◽  
Kidney Cortex ◽  
Acid Base Balance ◽  
Western Blots ◽  
High Affinity

Most amino acids filtered by the glomerulus are reabsorbed in the kidney via specialized transport systems. Recently, the cDNA encoding a high-affinity glutamate transporter, EAAC1, has been isolated and shown to be expressed at high levels in the kidney. To determine the potential role of EAAC1 in renal acidic amino acid reabsorption, the distribution of EAAC1 mRNA and protein in rat kidney was examined. In situ hybridization revealed that EAAC1 mRNA is expressed predominantly in S2 and S3 segments of the proximal tubules and at low levels in the inner stripe of outer medulla and inner medulla. Polyclonal antibodies raised against the carboxy terminus of EAAC1 recognized a single band of ∼70 kDa on Western blots of membrane protein from kidney cortex and medulla. Immunofluorescence microscopy revealed intense signals in the luminal membrane of S2 and S3 segments and weaker signals in S1 segments, descending thin limbs of long-loop nephrons, medullary thick ascending limbs, and distal convoluted tubules. These results are consistent with EAAC1 encoding the previously described apical high-affinity glutamate transporter in the kidney that mediates reabsorption of acidic amino acids in tubules beyond early proximal tubule S1 segments. Potential additional roles of EAAC1 in acid/base balance, cell volume regulation, and amino acid metabolism are discussed.

Expression of mRNA (D2) encoding a protein involved in amino acid transport in S3 proximal tubule

1992 ◽  
Vol 263 (6) ◽  
pp. F1087-F1092 ◽  
Author(s):  
Y. Kanai ◽  
M. G. Stelzner ◽  
W. S. Lee ◽  
R. G. Wells ◽  
D. Brown ◽  
...  
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Proximal Tubule ◽  
Amino Acid Transport ◽  
Glucose Transporter ◽  
Rat Kidney ◽  
Expression Cloning ◽  
Acid Transport ◽  
Specific Expression ◽  
High Affinity

A rat kidney- and intestine-specific cDNA (D2) that induces high-affinity, Na(+)-independent uptake of cystine and dibasic and neutral amino acids into cRNA-injected Xenopus oocytes was recently isolated by expression cloning in our laboratory (R. G. Wells and M. A. Hediger. Proc. Natl. Acad. Sci. USA 89: 5596-5600, 1992). At present it is not known whether the D2-encoded protein functions as a transporter or as a transporter activator. To gain more insight into the role of D2 in renal amino acid transport, we studied the site of its expression in the kidney. This was determined by Northern blot analysis and by using a combination of in situ hybridization and immunocytochemistry with antibodies that recognize specific proximal tubule segments. D2 antisense RNA hybridized to the same tubular segments that were strongly positive for anti-ecto-adenosinetriphosphatase but negative for carbonic anhydrase type IV and the facilitated glucose transporter GLUT2. We conclude that D2 mRNA is strongly expressed in the rat kidney proximal tubule S3 segment, although there is weak hybridization to the S1 and S2 segments. The signal is absent in all other parts of the kidney. The S3 specific expression of D2 mRNA coincides with the site of high-affinity transport of cystine and other amino acids, consistent with the proposed involvement of D2 in these processes.

Neutral amino acid transport systems of microvillous membrane of human placenta

1988 ◽  
Vol 254 (6) ◽  
pp. C773-C780 ◽  
Author(s):  
L. W. Johnson ◽  
C. H. Smith
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Amino Acid Transport ◽  
Plasma Membranes ◽  
Competitive Inhibition ◽  
Transport Systems ◽  
Acid Uptake ◽  
Acid Transport ◽  
Independent System ◽  
High Affinity

Placental transport produces concentrations of amino acids in fetal blood greater than those of maternal blood. Competitive inhibition studies of zwitterionic amino acid transport in isolated vesicles from the microvillous (maternal facing) plasma membranes of syncytiotrophoblast defined three transport systems: 1) a sodium-dependent system that supports methylaminoisobutyric acid (MeAIB) transport and has the characteristics of an A system; 2) a sodium-independent system with a high affinity for leucine and other amino acids with branched or aromatic side chains; and 3) a sodium-independent system with a preference for alanine as a substrate. The two sodium-independent systems could be further discriminated by marked specificity for trans stimulation with alanine or with leucine. System ASC, known to be present in whole placenta, and the neutral brush-border or imino systems of other polarized epithelia were apparently absent. Kinetic characteristics of the A system make it the probable primary driving force for concentrative transfer of its substrate amino acids to the fetus. Characteristics of the high-affinity leucine system demonstrated that it is saturated by normal serum leucine concentrations. Regulation of either system has the potential to alter placental amino acid uptake and transfer to the fetus.

AMINO ACID EXCRETION PATTERNS IN DEVELOPING RATS

1967 ◽  
Vol 45 (5) ◽  
pp. 867-872 ◽  
Author(s):  
William A. Webber
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Rat Kidney ◽  
Kidney Cortex ◽  
Acid Excretion ◽  
Human Infants ◽  
Rat Kidney Cortex ◽  
Progressive Development ◽  
Cortex Slices ◽  
Amino Acid Concentrations

Amino acid excretion patterns were studied in rats 2 to 12 weeks old. In general there was a decline in amino acid excretion over this period which paralleled that reported in human infants by other workers. The decrease was most marked for certain amino acids (glycine, histidine, and arginine). These changes in excretion are not explicable in terms of changes in plasma amino acid concentrations, nor is it likely that they result from differences in filtered load. They may reflect a progressive development of transport mechanisms for some amino acids over the period studied, in which case similar changes in the concentrating ability of rat kidney cortex slices would be predicted. Other possible explanations which are less readily tested include changes in permeability of the tubular cell membranes and differences in the glomerular filtering capacity relative to the amount of tubular tissue which has developed.

A comparison of the amino acid concentrating ability of the kidney cortex of newborn and mature rats

1968 ◽  
Vol 46 (2) ◽  
pp. 165-169 ◽  
Author(s):  
W. A. Webber ◽  
J. A. Cairns
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Cell Layer ◽  
Rat Kidney ◽  
The Other ◽  
Kidney Cortex ◽  
Acid Transport ◽  
Newborn Rat ◽  
Rat Kidney Cortex ◽  
A Cell

It has frequently been demonstrated that there are multiple mechanisms for amino acid transport and that these function to maintain a favorable intracellular level of amino acids within cells. In some instances they also make possible the transport of amino acids from one face of a cell layer to the other. In general, developing tissues have a higher concentrating ability than mature tissues. In the kidney, however, it has been observed that the ability to reabsorb amino acids may be less effective in developing than in mature organisms. Studies were carried out to determine whether the newborn rat kidney cortex differed from mature cortex in its ability to concentrate a representative group of amino acids. In general, the patterns observed for the concentrative uptake of glycine, L-leucine, α-aminoisobutyric acid, L-aspartic acid, and L-lysine were the same. In all cases uptake was initially more rapid in the mature tissue, but the concentration ratios ultimately reached were higher in the newborn tissues. It is concluded that, as in other developing tissues, newborn rat kidney cortex has a high concentrating ability and might therefore be expected to reabsorb amino acids at least as effectively as mature cortex. However the observation that uptake is relatively slow initially suggests that although the ability to establish a gradient at equilibrium is high the capacity of the system is relatively low and this may account for the apparent low capacity of the immature kidney to reabsorb amino acids.

Effect on kidney S35O4 uptake of compounds related to SO4 transport and metabolism

1964 ◽  
Vol 207 (1) ◽  
pp. 84-88 ◽  
Author(s):  
Ingrith J. Deyrup
Keyword(s):  
Amino Acids ◽  
Rat Kidney ◽  
Point Of View ◽  
Transport Systems ◽  
Kidney Cortex ◽  
Cellular Transport ◽  
Rat Kidney Cortex ◽  
Aryl Sulfatase

Experiments have been carried out to test the effects on S35O4 accumulation by rat kidney cortex slices in vitro of 1) compounds known to affect renal SO4 reabsorption (thiosulfate, amino acids); 2) compounds secreted by the kidney, or known to affect specific cellular transport systems (including tetraethylammonium ions, guanidine, creatinine, carinamide, probenecid, phloretin, diethylstilbestrol, ethylenediaminetetraacetate sodium); and 3) compounds related to SO4 metabolism (aryl sulfatase substrates). Under the conditions of the experiment, net S35O4 uptake was depressed by thiosulfate, certain amino acids, carinamide, phloretin, diethylstilbestrol, and aryl sulfatase substrates. It was enhanced by ethylenediaminetetraacetate sodium. Other compounds were without effect. These results are discussed from the point of view of the possible relationship between SO4 accumulation in vitro and transport in vivo.

scholarly journals A new family of proteins (rBAT and 4F2hc) involved in cationic and zwitterionic amino acid transport: a tale of two proteins in search of a transport function.

1994 ◽  
Vol 196 (1) ◽  
pp. 123-137 ◽  
Author(s):  
M Palacín
Keyword(s):  
Amino Acids ◽  
Small Intestine ◽  
Amino Acid ◽  
Amino Acid Transport ◽  
Transport Systems ◽  
Amino Acid Transporters ◽  
Cdna Clones ◽  
Missense Mutations ◽  
Acid Transport ◽  
High Affinity

The currently identified cDNA clones of mammalian amino acid transporters can be grouped into five different families. One family is composed of the proteins rBAT and the heavy chain (hc) of the cell surface antigen 4F2. RNAs encoding these two proteins induce a system b(o,+)-like (rBAT) and a system y+L-like (4F2hc) activity in Xenopus oocytes. Surprisingly, rBAT and 4F2hc do not seem to be pore-forming proteins. This finding supports the hypothesis that rBAT and 4F2hc are subunits or modulators of the corresponding amino acid transport systems. Expression of rBAT in oocytes induces high-affinity transport of cystine, which is shared with transport of cationic and zwitterionic amino acids. The rBAT gene is expressed mainly in kidney and small intestine. The rBAT protein is localized to the microvilli of proximal straight tubules of the kidney and mucosa from the small intestine. This finding is consistent with the involvement of rBAT in a high-affinity resorption system for cystine in the proximal straight tubule of the nephron. All of these characteristics suggest that rBAT is a good candidate for a cystinuria gene. Cystinuria is an inheritable defect in high-affinity transport of cystine, shared with cationic amino acids, through epithelial cells of the renal tubule and intestinal tract. Very recently, point missense mutations have been found in the rBAT gene of cystinuria patients. The most frequent rBAT mutation, M467T (threonine substitution of methionine at residue 467) nearly abolished the amino acid transport activity elicited by rBAT in oocytes. This result offers convincing evidence that rBAT is a cystinuria gene. Biochemical, cytological and genetic approaches are now needed to delineate the mechanism of action of rBAT and 4F2hc in the transport of amino acids.

Amino Acid Transport in a Water-mould: The Possible Regulatory Roles of Calcium and N6-(Δ2-isopentenyl)adenine

1975 ◽  
Vol 53 (9) ◽  
pp. 975-988 ◽  
Author(s):  
Danny P. Singh ◽  
Hérb. B. LéJohn
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Amino Acid Transport ◽  
Osmotic Shock ◽  
Inhibitory Effect ◽  
Transport Systems ◽  
Acid Transport ◽  
Isopentenyl Adenine ◽  
Energy Dependent ◽  
Water Mould

Transport of amino acids in the water-mould Achlya is an energy-dependent process. Based on competition kinetics and studies involving the influence of pH and temperature on the initial transport rates, it was concluded that the 20 amino acids (L-isomers) commonly found in proteins were transported by more than one, possibly nine, uptake systems. This is similar to the pattern elucidated for some bacteria but unlike those uncovered for all fungi studied to date. The nine different transport systems elucidated are: (i) methionine, (ii) cysteine, (iii) proline, (iv) serine–threonine, (v) aspartic and glutamic acids, (vi) glutamine and asparagine, (vii) glycine and alanine, (viii) histidine, lysine, and arginine, and (ix) phenylalanine–tyrosine–tryptophan and leucine–isoleucine–valine as two overlapping groups. Transport of all of these amino acids was inhibited by azide, cyanide, and its derivatives and 2,4-dinitrophenol. These agents normally interfere with metabolism at the level of the electron transport chain and oxidative phosphorylation. Osmotic shock treatment of the cells released, into the shock fluid, a glycopeptide that binds calcium as well as tryptophan but no other amino acid. The shocked cells are incapable of concentrating amino acids, but remain viable and reacquire this capacity when the glycopeptide is resynthesized.Calcium played more than a secondary role in the transport of the amino acids. When bound to the membrane-localized glycopeptide, it permits concentrative transport to take place. However, excess calcium can inhibit transport which can be overcome by chelating with citrate. Calculations show that the concentration of free citrate is most important. At low citrate concentrations (less than 1 mM) in the absence of exogenously supplied calcium, enhancement of amino acid transport occurs. At high concentrations (greater than 5 mM), citrate inhibits but this effect can be reversed by titrating with calcium. Evidently, the glycopeptide acts as a calcium sink to regulate the concentration of calcium made available to the cell for its membrane activities.N6-(Δ2-isopentenyl) adenine (a plant growth 'hormone') and analogues mimic the inhibitory effect of citrate and bind to the glycopeptide as well. Replot data for citrate and N6-(Δ2-isopentyl) adenine inhibition indicate that both agents have no more than one binding constant. These results implicate calcium, glycopeptide, and energy-dependent transport of solutes in some, as yet undefinable, way.

Placental amino acid transport

1995 ◽  
Vol 268 (6) ◽  
pp. C1321-C1331 ◽  
Author(s):  
A. J. Moe
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Human Placenta ◽  
Amino Acid Transport ◽  
Single Layer ◽  
Maternal Blood ◽  
Transport Systems ◽  
Amino Acid Transporters ◽  
Acid Transport ◽  
Principal Mechanism

Normal fetal growth and development depend on a continuous supply of amino acids from the mother to the fetus. The placenta is responsible for the transfer of amino acids between the two circulations. The human placenta is hemomonochorial, meaning that the maternal and fetal circulations are separated by a single layer of polarized epithelium called the syncytiotrophoblast, which is in direct contact with maternal blood. Transport proteins located in the microvillous and basal membranes of the syncytiotrophoblast are the principal mechanism for transfer from maternal blood to fetal blood. Knowledge of the function and regulation of syncytiotrophoblast amino acid transporters is of great importance in understanding the mechanism of placental transport and potentially improving fetal and newborn outcomes. The development of methods for the isolation of microvillous and basal membrane vesicles from human placenta over the past two decades has contributed greatly to this understanding. Now a primary cultured trophoblast model is available to study amino acid transport and regulation as the cells differentiate. The types of amino acid transporters and their distribution between the syncytiotrophoblast microvillous and basal membranes are somewhat unique compared with other polarized epithelia. These differences may reflect the unusual circumstance of this epithelium that is exposed to blood on both sides. The current state of knowledge as to the types of transport systems present in syncytiotrophoblast, their regulation, and the effects of maternal consumption of drugs on transport are discussed.

Sign in / Sign up

Export Citation Format

Share Document