Bidirectional transport of glutamine across the cell membrane in rat liver.
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ABSTRACT:
SUBMITTER: Fafournoux P
PROVIDER: S-EPMC1152517 | biostudies-other | 1983 Nov
REPOSITORIES: biostudies-other
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Project description:Transport of glutamine by brush-border vesicles prepared from the renal cortex was studied. The transport system had both Na+-dependent and Na+-independent components. The presence of Na+ in the incubation resulted in an 'overshoot' at 30s at which time the rates of transport were approx. 8 times the values obtained in the absence of Na+. Variation of the glutamine concentration showed that the system obeyed Michaelis-Menten kinetics with Km and Vmax. values for the Na+-dependent system of 0.86 mM and 9.6 nmol/min per mg of protein respectively. Vesicles obtained from chronically acidotic rats showed similar kinetic characteristics. The Km and Vmax. values for the Na+-dependent system were 0.76 mM and 9.6 nmol/min per mg of protein respectively. There was increased uptake of glutamine by vesicles from acidotic rats and this increase was associated with increased activity of gamma-glutamyltransferase in these preparations. Vesicles from acidotic rats, however, showed no increase in glucose transport and no increase in the activity of maltase, another brush-border enzyme.
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Project description:In BtuB, the Escherichia coli TonB-dependent transporter for vitamin B12, substrate binding to the extracellular surface unfolds a conserved energy coupling motif termed the Ton box into the periplasm. This transmembrane signaling event facilitates an interaction between BtuB and the inner-membrane protein TonB. In this study, continuous-wave and pulse electron paramagnetic resonance in a native outer-membrane preparation demonstrate that signaling also occurs from the periplasmic to the extracellular surface in BtuB. The binding of a TonB fragment to the periplasmic interface alters the configuration of the second extracellular loop and partially dissociates a spin-labeled substrate analog. Moreover, mutants in the periplasmic Ton box that are transport-defective alter the binding site for vitamin B12 in BtuB. This work demonstrates that the Ton box and the extracellular substrate binding site are allosterically coupled in BtuB, and that TonB binding may initiate a partial round of transport.
| S-EPMC5103032 | biostudies-literature
Project description:1. Alanine, glutamine and serine were actively accumulated in liver cells isolated from starved rats. 2. This accumulation was inhibited when either Na+ or HCO3- ions were omitted from the incubation medium. In general the degree of dependence on Na+ was quantitatively similar to that on HCO3-. 3. The apparent Km values for the transport of all three amino acids were in the range 3--5mM with Vmax. values in the range 15--25nmol/min per mg of cell protein at 37 degrees C. 4. Alanine and serine transport were mutually competitive; glutamine inhibited the transport of alanine and serine non-competitively. 5. The initial rate of transport of these amino acids was inhibited when the intracellular content of ATP was decreased. 6. Ouabain inhibited the rate of alanine transport without inhibiting the rate of alanine metabolism. 7. It is concluded that a minimum of three transport systems must be postulated to exist in the liver cell plasma membrane to account for the transport of alanine, serine and glutamine. The rate of transport of these amino acids in isolated hepatocytes is unlikely to limit the rate at which they are metabolized.
| S-EPMC1186306 | biostudies-other
Project description:Mitochondrial-inner-membrane vesicles, isolated from Ehrlich ascites carcinoma cells by titration with detergents, accumulated L-glutamine by a very efficient transport system. The vesicles lack any phosphate-activated glutaminase activity, allowing measurement of transport rates without interference by L-glutamine metabolism. The time course of the transport was linear for the first 60 s, reaching a steady state after 120 min. L-Glutamine transport showed co-operativity, with a Hill coefficient of 2.2; the kinetic parameters S0.5 and Vmax had values of 5 mM and 26 nmol/30 s per mg of protein respectively. The pH-dependence curve showed a bell shape, with a pH optimum about 8.0. The uptake of L-glutamine was not affected by the presence of a 50-fold molar excess of D-glutamine, L-cysteine, L-histidine, L-alanine, L-serine and L-leucine, whereas L-glutamate behaved as a poor inhibitor. The structural analogue L-glutamate gamma-hydroxamate (5mM) inhibited the net uptake by 68%; interestingly, other analogues (6-diazo-5-oxo-L-norleucine, acivicin and L-glutamate gamma-hydrazide) were ineffective. The impermeant thiol reagent p-chloromercuriphenylsulphonic acid (0.5mM) completely abolished the mitochondrial L-glutamine uptake; in contrast, other thiol reagents (mersalyl and N-ethylmaleimide) did not significantly affect the transport. These data confirm the existence of a specific transport system with high capacity for L-glutamine in the mitochondrial inner membrane, a step preceding the highly operative glutaminolysis in tumour cells.
| S-EPMC1136972 | biostudies-other
Project description:The human MHC class I-related neonatal Fc receptor, hFcRn, mediates bidirectional transport of IgG across mucosal barriers. Here, we find that at steady state hFcRn distributes predominantly to an apical intracellular compartment and almost exclusively to the basolateral cell surface of polarized epithelial cells. It moves only transiently to the apical membrane. Ligand binding does not redistribute the steady state location of the receptor. Removal of the cytoplasmic tail that contains di-leucine and tryptophan-based endocytosis motifs or incubation at low temperature (18 degrees C) redistributes the receptor apically. The rates of endocytosis of the full-length hFcRn from the apical or basolateral membrane domains, however, are equal. Thus, the strong cell surface polarity displayed by hFcRn results from dominant basolateral sorting by motifs in the cytoplasmic tail that nonetheless allows for a cycle of bidirectional transcytosis.
| S-EPMC379272 | biostudies-literature
Project description:The concept that cancer is a metabolic disease is now well acknowledged: many cancer cell types rely mostly on glucose and some amino acids, especially glutamine for energy supply. These findings were corroborated by overexpression of plasma membrane nutrient transporters, such as the glucose transporters (GLUTs) and some amino acid transporters such as ASCT2, LAT1, and ATB0,+, which became promising targets for pharmacological intervention. On the basis of their sodium-dependent transport modes, ASCT2 and ATB0+ have the capacity to sustain glutamine need of cancer cells; while LAT1, which is sodium independent will have the role of providing cancer cells with some amino acids with plausible signaling roles. According to the metabolic reprogramming of many types of cancer cells, glucose is mainly catabolized by aerobic glycolysis in tumors, while the fate of Glutamine is completed at mitochondrial level where the enzyme Glutaminase converts Glutamine to Glutamate. Glutamine rewiring in cancer cells is heterogeneous. For example, Glutamate is converted to ?-Ketoglutarate giving rise to a truncated form of Krebs cycle. This reprogrammed pathway leads to the production of ATP mainly at substrate level and regeneration of reducing equivalents needed for cells growth, redox balance, and metabolic energy. Few studies on hypothetical mitochondrial transporter for Glutamine are reported and indirect evidences suggested its presence. Pharmacological compounds able to inhibit Glutamine metabolism may represent novel drugs for cancer treatments. Interestingly, well acknowledged targets for drugs are the Glutamine transporters of plasma membrane and the key enzyme Glutaminase.
| S-EPMC5770653 | biostudies-literature
Project description:High boron (B) concentration is toxic to plants that limit plant productivity. Recent studies have shown the involvement of the members of major intrinsic protein (MIP) family in controlling B transport. Here, we have provided experimental evidences showing the bidirectional transport activity of rice OsPIP1;3 and OsPIP2;6. Boron transport ability of OsPIP1;3 and OsPIP2;6 were displayed in yeast HD9 mutant strain (?fps1?acr3?ycf1) as a result of increased B sensitivity, influx and accumulation by OsPIP1;3, and rapid efflux activity by OsPIP2;6. RT-PCR analysis showed strong upregulation of OsPIP1;3 and OsPIP2;6 transcripts in roots by B toxicity. Transgenic Arabidopsis lines overexpressing OsPIP1;3 and OsPIP2;6 exhibited enhanced tolerance to B toxicity. Furthermore, B concentration was significantly increased after 2 and 3 hours of tracer boron ((10)B) treatment. Interestingly, a rapid efflux of (10)B from the roots of the transgenic plants was observed within 1 h of (10)B treatment. Boron tolerance in OsPIP1;3 and OsPIP2;6 lines was inhibited by aquaporin inhibitors, silver nitrate and sodium azide. Our data proved that OsPIP1;3 and OsPIP2;6 are indeed involved in both influx and efflux of boron transport. Manipulation of these PIPs could be highly useful in improving B tolerance in crops grown in high B containing soils.
| S-EPMC4763227 | biostudies-literature
Project description:Glutamine uptake was examined in isolated renal brush-border and basolateral-membrane vesicles from control and acidotic rats. In brush-border vesicles from acidotic animals, there was a significant increase in the initial rate of glutamine uptake compared with that in controls. Lowering the pH of the medium increased the initial rate of glutamine uptake in brush-border vesicles from acidotic, but not from control, rats. In brush-border vesicles from both groups of animals, two saturable transport systems mediated glutamine uptake. There was a 2-fold increase in the Vmax. of the low-affinity high-capacity system in the brush-border vesicles from the acidotic animals compared with that from control animals, with no alteration in the other kinetic parameters. There was no difference in glutamine uptake by the two saturable transport systems in basolateral vesicles from control and acidotic animals. Lowering the incubation-medium pH increased the uptake of glutamine by basolateral vesicles from both control and acidotic rats to a similar extent. The data indicate that during acidosis there are alterations in glutamine transport by both the basolateral and brush-border membrane which could enhance its uptake by the renal-tubule cell for use in ammoniagenesis.
| S-EPMC1153148 | biostudies-other
Project description:1. On perfusion of livers from fed rats with a semi-synthetic medium containing no added amino acids there is a rapid release of glutamine during the first 15min (15.6+/-0.8mumol/h per g wet wt.; mean+/-s.e.m. of 35 experiments), followed by a low linear rate of production (3.6+/-0.3mumol/h per g wet wt.; mean+/-s.e.m. of three experiments). The rapid initial release can be accounted for as wash-out of preexisting intracellular glutamine. 2. Addition of readily permeating substrates, or substrate combinations, giving rise to intracellular glutamate or ammonia, or both, did not appreciably increase the rate of glutamine production over the endogenous rate. The maximum rate obtained was from proline plus alanine; even then the rate represented less than one-fortieth of the capacity of glutamine synthetase measured in vitro. 3. Complete inhibition of respiration in the perfusions [no erythrocytes in the medium; 1mm-cyanide; N(2)+CO(2) (95:5) in the gas phase] or perfusion with glutamine synthetase inhibitors [l-methionine dl-sulphoximine; dl-(+)-allo-delta-hydroxylysine] abolishes the low linear rate of glutamine synthesis, but not the initial rapid release of glutamine. 4. In livers from 48h-starved rats initial release (0-15min) of glutamine was decreased (10.6+/-1.1mumol/h per g wet wt.; mean+/-s.e.m. of 11 experiments) and the subsequent rate of glutamine production was lower than in livers from fed rats. Again proline plus alanine was the only substrate combination giving an increase significantly above the endogenous rate. 5. The rate of glutamine synthesis de novo by the liver is apparently unrelated to the tissue content of glutamate or ammonia. 6. The blood glutamine concentration is increased by 50% within 1h of elimination of the liver from the circulation of rats in vivo. 7. There is an output of glutamine by the brain under normal conditions; the mean arterio-venous difference for six rats was 0.023mumol/ml. 8. The high potential activity of liver glutamine synthetase appears to be inhibited by some unknown mechanism: the function of the liver under normal conditions is probably the disposal of glutamine produced by extrahepatic tissues.
| S-EPMC1177235 | biostudies-other
Project description:The transport of glutamine and alanine into isolated rat colonocytes was studied. The transport of both amino acids appears to be dependent on a Na+ gradient. The apparent Km values for the transport of glutamine and alanine were 2.56 +/- 0.84 and 5.35 +/- 1.20 mM respectively, but with similar Vmax. values. Glutamine and alanine transport were mutually competitive, and the transport of both amino acids was competitively inhibited by 2-methylaminoisobutyrate. In contrast, histidine inhibited the transport of both glutamine and alanine non-competitively. It is concluded that glutamine and alanine are transported into rat colonocytes by a common carrier system similar to System A of other cells. It is suggested that the metabolic function of this transport system in rat colonocytes might be the partial exchange of extracellular glutamine for intracellular alanine.
| S-EPMC1147106 | biostudies-other