scholarly journals Increased Expression of UMAMIT Amino Acid Transporters Results in Activation of Salicylic Acid Dependent Stress Response

2021 ◽  
Vol 11 ◽  
Author(s):  
Julien Besnard ◽  
Unnati Sonawala ◽  
Bal Maharjan ◽  
Eva Collakova ◽  
Scott A. Finlayson ◽  
...  
Keyword(s):  
Salicylic Acid ◽  
Amino Acid ◽  
Stress Response ◽  
Stress Responses ◽  
Amino Acid Transporter ◽  
Marker Genes ◽  
Amino Acid Transporters ◽  
Loss Of Function ◽  
Over Expression ◽  
Acid Transporter

In addition to their role in the biosynthesis of important molecules such as proteins and specialized metabolites, amino acids are known to function as signaling molecules through various pathways to report nitrogen status and trigger appropriate metabolic and cellular responses. Moreover, changes in amino acid levels through altered amino acid transporter activities trigger plant immune responses. Specifically, loss of function of major amino acid transporter, over-expression of cationic amino acid transporter, or over-expression of the positive regulators of membrane amino acid export all lead to dwarfed phenotypes and upregulated salicylic acid (SA)-induced stress marker genes. However, whether increasing amino acid exporter protein levels lead to similar stress phenotypes has not been investigated so far. Recently, a family of transporters, namely USUALLY MULTIPLE ACIDS MOVE IN AND OUT TRANSPORTERS (UMAMITs), were identified as amino acid exporters. The goal of this study was to investigate the effects of increased amino acid export on plant development, growth, and reproduction to further examine the link between amino acid transport and stress responses. The results presented here show strong evidence that an increased expression of UMAMIT transporters induces stress phenotypes and pathogen resistance, likely due to the establishment of a constitutive stress response via a SA-dependent pathway.

scholarly journals Pichia pastoris and the Recombinant Human Heterodimeric Amino Acid Transporter 4F2hc-LAT1: From Clone Selection to Pure Protein

2021 ◽  
Vol 4 (3) ◽  
pp. 51
Author(s):  
Satish Kantipudi ◽  
Daniel Harder ◽  
Sara Bonetti ◽  
Dimitrios Fotiadis ◽  
Jean-Marc Jeckelmann
Keyword(s):  
Amino Acid ◽  
Pichia Pastoris ◽  
Protein Complexes ◽  
Amino Acid Transporter ◽  
Amino Acid Transporters ◽  
Expression Host ◽  
Amino Acids And Derivatives ◽  
Heterodimeric Complex ◽  
Acid Transporter ◽  
And Function

Heterodimeric amino acid transporters (HATs) are protein complexes composed of two subunits, a heavy and a light subunit belonging to the solute carrier (SLC) families SLC3 and SLC7. HATs transport amino acids and derivatives thereof across the plasma membrane. The human HAT 4F2hc-LAT1 is composed of the type-II membrane N-glycoprotein 4F2hc (SLC3A2) and the L-type amino acid transporter LAT1 (SLC7A5). 4F2hc-LAT1 is medically relevant, and its dysfunction and overexpression are associated with autism and tumor progression. Here, we provide a general applicable protocol on how to screen for the best membrane transport protein-expressing clone in terms of protein amount and function using Pichia pastoris as expression host. Furthermore, we describe an overexpression and purification procedure for the production of the HAT 4F2hc-LAT1. The isolated heterodimeric complex is pure, correctly assembled, stable, binds the substrate L-leucine, and is thus properly folded. Therefore, this Pichia pastoris-derived recombinant human 4F2hc-LAT1 sample can be used for downstream biochemical and biophysical characterizations.

scholarly journals Downregulation of Placental Amino Acid Transporter Expression and mTORC1 Signaling Activity Contributes to Fetal Growth Retardation in Diabetic Rats

2020 ◽  
Vol 21 (5) ◽  
pp. 1849
Author(s):  
Jie Xu ◽  
Jiao Wang ◽  
Yang Cao ◽  
Xiaotong Jia ◽  
Yujia Huang ◽  
...  
Keyword(s):  
Amino Acid ◽  
Rat Model ◽  
Amino Acid Transporter ◽  
Amino Acid Transporters ◽  
Amino Acid Transport System ◽  
Intrauterine Growth ◽  
System L ◽  
Acid Transporter ◽  
Transporter Expression

Alterations in placental transport may contribute to abnormal fetal intrauterine growth in pregnancies complicated by diabetes, but it is not clear whether the placental amino acid transport system is altered in diabetic pregnancies. We therefore studied the changes in the expressions of placental amino acid transporters in a rat model of diabetes induced by streptozotocin, and tested the effects of hyperglycemia on trophoblast amino acid transporter in vitro. Our results showed that the expressions for key isoforms of system L amino acid transporters were significantly reduced in the placentas of streptozotocin-induced diabetic pregnant rats, which was associated with the decreased birthweight in the rats. A decreased placental efficiency and decreased placental mammalian target of rapamycin (mTOR) complex 1 (mTORC1) activity were also found in the rat model. In addition, hyperglycemia in vitro could inhibit amino acid transporter expression and mTORC1 activity in human trophoblast. Inhibition of mTORC1 activity led to reduced amino acid transporter expression in placental trophoblast. We concluded that reduced placental mTORC1 activity during pregnancy resulted in decreased placental amino acid transporter expression and, subsequently, contributed to fetal intrauterine growth restriction in pregnancies complicated with diabetes.

scholarly journals Amino Acid Transport Defects in Human Inherited Metabolic Disorders

2019 ◽  
Vol 21 (1) ◽  
pp. 119 ◽  
Author(s):  
Raquel Yahyaoui ◽  
Javier Pérez-Frías
Keyword(s):  
Amino Acid ◽  
Metabolic Disorders ◽  
Strong Association ◽  
Cell Types ◽  
Amino Acid Transporter ◽  
Molecular Defect ◽  
Amino Acid Transporters ◽  
Inherited Disorders ◽  
Laboratory Findings ◽  
Acid Transporter

Amino acid transporters play very important roles in nutrient uptake, neurotransmitter recycling, protein synthesis, gene expression, cell redox balance, cell signaling, and regulation of cell volume. With regard to transporters that are closely connected to metabolism, amino acid transporter-associated diseases are linked to metabolic disorders, particularly when they involve different organs, cell types, or cell compartments. To date, 65 different human solute carrier (SLC) families and more than 400 transporter genes have been identified, including 11 that are known to include amino acid transporters. This review intends to summarize and update all the conditions in which a strong association has been found between an amino acid transporter and an inherited metabolic disorder. Many of these inherited disorders have been identified in recent years. In this work, the physiological functions of amino acid transporters will be described by the inherited diseases that arise from transporter impairment. The pathogenesis, clinical phenotype, laboratory findings, diagnosis, genetics, and treatment of these disorders are also briefly described. Appropriate clinical and diagnostic characterization of the underlying molecular defect may give patients the opportunity to avail themselves of appropriate therapeutic options in the future.

scholarly journals Transport of butyryl-l-carnitine, a potential prodrug, via the carnitine transporter OCTN2 and the amino acid transporter ATB0,+

2007 ◽  
Vol 293 (5) ◽  
pp. G1046-G1053 ◽  
Author(s):  
Sonne R. Srinivas ◽  
Puttur D. Prasad ◽  
Nagavedi S. Umapathy ◽  
Vadivel Ganapathy ◽  
Prem S. Shekhawat
Keyword(s):  
Amino Acid ◽  
Mammalian Cells ◽  
High Capacity ◽  
Amino Acid Transporter ◽  
Xenopus Laevis Oocytes ◽  
Loss Of Function ◽  
Bacterial Fermentation ◽  
Carnitine Transporter ◽  
Inward Currents ◽  
Acid Transporter

l-Carnitine is absorbed in the intestinal tract via the carnitine transporter OCTN2 and the amino acid transporter ATB0,+. Loss-of-function mutations in OCTN2 may be associated with inflammatory bowel disease (IBD), suggesting a role for carnitine in intestinal/colonic health. In contrast, ATB0,+ is upregulated in bowel inflammation. Butyrate, a bacterial fermentation product, is beneficial for prevention/treatment of ulcerative colitis. Butyryl-l-carnitine (BC), a butyrate ester of carnitine, may have potential for treatment of gut inflammation, since BC would supply both butyrate and carnitine. We examined the transport of BC via ATB0,+ to determine if this transporter could serve as a delivery system for BC. We also examined the transport of BC via OCTN2. Studies were done with cloned ATB0,+ and OCTN2 in heterologous expression systems. BC inhibited ATB0,+-mediated glycine transport in mammalian cells (IC50, 4.6 ± 0.7 mM). In Xenopus laevis oocytes expressing human ATB0,+, BC induced Na+-dependent inward currents under voltage-clamp conditions. The currents were saturable with a K0.5 of 1.4 ± 0.1 mM. Na+ activation kinetics of BC-induced currents suggested involvement of two Na+ per transport cycle. BC also inhibited OCTN2-mediated carnitine uptake (IC50, 1.5 ± 0.3 μM). Transport of BC via OCTN2 is electrogenic, as evidenced from BC-induced inward currents. These currents were Na+ dependent and saturable ( K0.5, 0.40 ± 0.02 μM). We conclude that ATB0,+ is a low-affinity/high-capacity transporter for BC, whereas OCTN2 is a high-affinity/low-capacity transporter. ATB0,+ may mediate intestinal absorption of BC when OCTN2 is defective.

Function and structure of heterodimeric amino acid transporters

2001 ◽  
Vol 281 (4) ◽  
pp. C1077-C1093 ◽  
Author(s):  
Carsten A. Wagner ◽  
Florian Lang ◽  
Stefan Bröer
Keyword(s):  
Amino Acid ◽  
Membrane Protein ◽  
Heavy Chain ◽  
Light Chain ◽  
Disulfide Bridge ◽  
Amino Acid Transporter ◽  
Light Chains ◽  
Amino Acid Transporters ◽  
Amino Acid Transport System ◽  
Acid Transporter

Heterodimeric amino acid transporters are comprised of two subunits, a polytopic membrane protein (light chain) and an associated type II membrane protein (heavy chain). The heavy chain rbAT (related to b0,+ amino acid transporter) associates with the light chain b0,+AT (b0,+ amino acid transporter) to form the amino acid transport system b0,+, whereas the homologous heavy chain 4F2hc interacts with several light chains to form system L (with LAT1 and LAT2), system y+L (with y+LAT1 and y+LAT2), system x[Formula: see text](with xAT), or system asc (with asc1). The association of light chains with the two heavy chains is not unambiguous. rbAT may interact with LAT2 and y+LAT1 and vice versa; 4F2hc may interact with b0,+AT when overexpressed. 4F2hc is necessary for trafficking of the light chain to the plasma membrane, whereas the light chains are thought to determine the transport characteristics of the respective heterodimer. In contrast to 4F2hc, mutations in rbAT suggest that rbAT itself takes part in the transport besides serving for the trafficking of the light chain to the cell surface. Heavy and light subunits are linked together by a disulfide bridge. The disulfide bridge, however, is not necessary for the trafficking of rbAT or 4F2 heterodimers to the membrane or for the functioning of the transporter. However, there is experimental evidence that the disulfide bridge in the 4F2hc/LAT1 heterodimer plays a role in the regulation of a cation channel. These results highlight complex interactions between the different subunits of heterodimeric amino acid transporters and suggest that despite high grades of homology, the interactions between rbAT and 4F2hc and their respective partners may be different.

192 AMINO ACID TRANSPORTER EXPRESSION IN BOVINE OVIDUCT EPITHELIAL CELLS

2008 ◽  
Vol 20 (1) ◽  
pp. 175
Author(s):  
S. L. Whitear ◽  
H. J. Leese
Keyword(s):  
Amino Acid ◽  
Epithelial Cells ◽  
Amino Acid Transporter ◽  
Positive Control ◽  
Amino Acid Transporters ◽  
Oviduct Epithelial Cells ◽  
Acid Transporter ◽  
Oviduct Fluid ◽  
Transporter Expression

Oviduct fluid provides the environment for the gametes and early embryo but little is known about the mechanisms underlying its formation. Components of oviduct fluid have been shown to be present at concentrations different from that in blood, indicative of selective transport by the epithelial cells lining the lumen. For example, amino acid concentrations in oviduct fluid differ from those in extracellular fluid and have also been shown to be important to preimplantation embryos in vitro, enhancing development, especially when added at physiological concentrations. However, little is known about amino acid transport systems in the oviduct, and the aim of this work was to search for mRNA transcripts for amino acid transporters in bovine oviduct epithelial cells. Contra- and ipsi-lateral oviducts were removed from abattoir-derived reproductive tracts at specific stages of the reproductive cycle. Oviducts were trimmed of surrounding tissue and fat and slit longitudinally to expose the luminal surface. Bovine oviduct epithelial cells (bOEC) were scraped from the surface using a sterile glass coverslip and washed by centrifugation. mRNA was isolated using Trizol-chloroform extraction and lithium chloride precipitation methods. PCR was used to detect cDNA encoding the amino acid transporters CAT-1, CAT-4, and LAT1. A negative control (water) and a positive control (human placental cDNA) were included in each experiment and β-actin expression was used as a positive control for cDNA library generation. Products were separated by agarose gel electrophoresis. PCR for β-actin resulted in the presence of a positive band in all samples, showing successful extraction of mRNA and generation of cDNA libraries. mRNA for CAT-1 and LAT1 was detected in bOEC from contra- and ipsi-lateral oviducts and from each cycle stage tested. There was, however, no detectable mRNA for CAT-4 in any of the samples. To our knowledge, this is the first report of amino acid transporter expression in the mammalian oviduct. CAT-1 is a ubiquitous sodium-independent uniporter of cationic amino acids that has been localized to the basolateral membrane of epithelial cells. The presence of mRNA for this amino acid transporter in all samples tested is therefore to be expected. LAT1 is a obligatory exchanger which exports glutamine and cystine and imports large uncharged branched-chain amino acids. This transporter may be partly responsible for the high concentration of glutamate in the basal compartment of in vitro cell cultures reported in our previous work (Whitear and Leese 2007 Biennial Meet. Joint Fertil. Soc., York, UK). CAT-4 shares only 40% sequence homology with CAT-1 and its function is unknown. Its expression appears to be restricted to brain, testis, and placenta, and the absence of mRNA for the oviduct was, perhaps, not surprising. Further experiments will investigate expression levels of other amino acid transporters in bOEC and transporter localization using immunohistochemistry. This work was funded by the BBSRC and ANGLE Technology Ltd.

scholarly journals Skeletal muscle amino acid transporter expression is increased in young and older adults following resistance exercise

2011 ◽  
Vol 111 (1) ◽  
pp. 135-142 ◽  
Author(s):  
Micah J. Drummond ◽  
Christopher S. Fry ◽  
Erin L. Glynn ◽  
Kyle L. Timmerman ◽  
Jared M. Dickinson ◽  
...  
Keyword(s):  
Older Adults ◽  
Amino Acid ◽  
Resistance Exercise ◽  
Amino Acid Transporter ◽  
Initiation Factor ◽  
Amino Acid Transporters ◽  
Α Subunit ◽  
Mtorc1 Signaling ◽  
Acid Transporter ◽  
Transporter Expression

Amino acid transporters and mammalian target of rapamycin complex 1 (mTORC1) signaling are important contributors to muscle protein anabolism. Aging is associated with reduced mTORC1 signaling following resistance exercise, but the role of amino acid transporters is unknown. Young ( n = 13; 28 ± 2 yr) and older ( n = 13; 68 ± 2 yr) subjects performed a bout of resistance exercise. Skeletal muscle biopsies ( vastus lateralis) were obtained at basal and 3, 6, and 24 h postexercise and were analyzed for amino acid transporter mRNA and protein expression and regulators of amino acid transporter transcription utilizing real-time PCR and Western blotting. We found that basal amino acid transporter expression was similar in young and older adults ( P > 0.05). Exercise increased L-type amino acid transporter 1/solute-linked carrier (SLC) 7A5, CD98/SLC3A2, sodium-coupled neutral amino acid transporter 2/SLC38A2, proton-assisted amino acid transporter 1/SLC36A1, and cationic amino acid transporter 1/SLC7A1 mRNA expression in both young and older adults ( P < 0.05). L-type amino acid transporter 1 and CD98 protein increased only in younger adults ( P < 0.05). eukaryotic initiation factor 2 α-subunit (S52) increased similarly in young and older adults postexercise ( P < 0.05). Ribosomal protein S6 (S240/244) and activating transcription factor 4 nuclear protein expression tended to be higher in the young, while nuclear signal transducer and activator of transcription 3 (STAT3) (Y705) was higher in the older subjects postexercise ( P < 0.05). These results suggest that the rapid upregulation of amino acid transporter expression following resistance exercise may be regulated differently between the age groups, but involves a combination of mTORC1, activating transcription factor 4, eukaryotic initiation factor 2 α-subunit, and STAT3. We propose an increase in amino acid transporter expression may contribute to enhanced amino acid sensitivity following exercise in young and older adults. In older adults, the increased nuclear STAT3 phosphorylation may be indicative of an exercise-induced stress response, perhaps to export amino acids from muscle cells.

scholarly journals Cloning, functional expression and dietary regulation of the mouse neutral and basic amino acid transporter (NBAT)

1997 ◽  
Vol 328 (2) ◽  
pp. 657-664 ◽  
Author(s):  
Hiroko SEGAWA ◽  
Ken-ichi MIYAMOTO ◽  
Yoshio OGURA ◽  
Hiromi HAGA ◽  
Kyoko MORITA ◽  
...  
Keyword(s):  
Amino Acid ◽  
Xenopus Oocytes ◽  
Physiological Role ◽  
Basic Amino Acid ◽  
Functional Expression ◽  
Amino Acid Transporter ◽  
Amino Acid Transporters ◽  
Transport Activity ◽  
Acid Transporter ◽  
Mouse Ileum

The Na+-independent dibasic and neutral amino acid transporter NBAT is among the least hydrophobic of mammalian amino acid transporters. The transporter contains one to four transmembrane domains and induces amino acid transport activity via a b0,+-like system when expressed in Xenopus oocytes. However, the physiological role of NBAT remains unclear. Complementary DNA clones encoding mouse NBAT have now been isolated. The expression of mouse NBAT in Xenopus oocytes also induced an obligatory amino acid exchange activity similar to that of the b0,+-like system. The amount of NBAT mRNA in mouse kidney increased during postnatal development, consistent with the increase in renal cystine and dibasic transport activity. Dietary aspartate induced a marked increase in cystine transport via the b0,+ system in mouse ileum. A high-aspartate diet also increased the amount of NBAT mRNA in mouse ileum. In the ileum of mice fed on the aspartate diet, the extent of cystine transport was further increased by preloading brush border membrane vesicles with lysine. Hybrid depletion of NBAT mRNA from ileal polyadenylated RNA revealed that the increase in cystine transport activity induced by the high-aspartate diet, as measured in Xenopus oocytes, was attributable to NBAT. These results demonstrate that mouse NBAT has an important role in cystine transport.

scholarly journals Wnt regulates amino acid transporter Slc7a5 and so constrains the integrated stress response in mouse embryos

EMBO Reports ◽  
2019 ◽  
Vol 21 (1) ◽  
Author(s):  
Nadège Poncet ◽  
Pamela A Halley ◽  
Christopher Lipina ◽  
Marek Gierliński ◽  
Alwyn Dady ◽  
...  
Keyword(s):  
Amino Acid ◽  
Stress Response ◽  
Amino Acid Transporter ◽  
Mouse Embryos ◽  
Integrated Stress Response ◽  
Acid Transporter

scholarly journals IL-6 stimulates system A amino acid transporter activity in trophoblast cells through STAT3 and increased expression of SNAT2

2009 ◽  
Vol 297 (5) ◽  
pp. C1228-C1235 ◽  
Author(s):  
H. N. Jones ◽  
T. Jansson ◽  
T. L. Powell
Keyword(s):  
Amino Acid ◽  
Protein Expression ◽  
Nutrient Transport ◽  
Amino Acid Transporter ◽  
Amino Acid Transporters ◽  
Trophoblast Cells ◽  
System A ◽  
Tnf Α ◽  
Gene And Protein Expression ◽  
Acid Transporter

Changes in placental nutrient transport are closely associated with abnormal fetal growth. However, the molecular mechanisms underlying the regulation of placental amino acid transporters are unknown. We demonstrate that physiological concentrations of the proinflammatory cytokines interleukin (IL)-6 and tumor necrosis factor (TNF)-α stimulate the activity of amino acid transporter system A, but not system L, in cultured human primary trophoblast cells. Both cytokines increased the gene and protein expression of the Na+-coupled neutral amino acid transporter (SNAT)2 isoform and upregulated SNAT1 protein expression. IL-6 increased Tyr705 phosphorylation of signal transducer and activator of transcription 3 (STAT3). In cells transfected with small interfering RNA (siRNA) targeting STAT3, the RNA and protein expression of SNAT2, but not SNAT1, was reduced and the stimulating effect of IL-6 on system A activity was abolished. Despite eliciting similar responses in amino acid transport activity and transporter expression, TNF-α effects on system A activity were not mediated through the JAK/STAT pathway. In conclusion, we have identified a novel regulatory pathway involving increased gene expression of the SNAT2 isoform mediated by a STAT-dependent pathway, which links IL-6 to increased activity of system A, a ubiquitously expressed transporter of neutral amino acids. From these new findings, we propose that upregulation of amino acid transporters by cytokines may contribute to increased placental nutrient transport and fetal overgrowth, which are commonly found in pregnancies complicated by maternal diabetes and obesity.

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