scholarly journals The Ectodomains of rBAT and 4F2h Are Fake or Orphan α-Glucosidases

Molecules ◽  
2021 ◽  
Vol 26 (20) ◽  
pp. 6231
Author(s):  
Joana Fort ◽  
Adrià Nicolàs-Aragó ◽  
Manuel Palacín

It is known that 4F2hc and rBAT are the heavy subunits of the heteromeric amino acid transporters (HATs). These heavy subunits are N-glycosylated proteins, with an N-terminal domain, one transmembrane domain and a bulky extracellular domain (ectodomain) that belongs to the α-amylase family. The heavy subunits are covalently linked to a light subunit from the SLC7 family, which is responsible for the amino acid transport activity, forming a heterodimer. The functions of 4F2hc and rBAT are related mainly to the stability and trafficking of the HATs in the plasma membrane of vertebrates, where they exert the transport activity. Moreover, 4F2hc is a modulator of integrin signaling, has a role in cell fusion and it is overexpressed in some types of cancers. On the other hand, some mutations in rBAT are found to cause the malfunctioning of the b0,+ transport system, leading to cystinuria. The ectodomains of 4F2hc and rBAT share both sequence and structure homology with α-amylase family members. Very recently, cryo-EM has revealed the structure of several HATs, including the ectodomains of rBAT and 4F2hc. Here, we analyze available data on the ectodomains of rBAT and 4Fhc and their relationship with the α-amylase family. The physiological relevance of this relationship remains largely unknown.

2002 ◽  
Vol 364 (3) ◽  
pp. 767-775 ◽  
Author(s):  
Sabine WOLF ◽  
Annette JANZEN ◽  
Nicole VÉKONY ◽  
Ursula MARTINÉ ◽  
Dennis STRAND ◽  
...  

Member 4 of human solute carrier family 7 (SLC7A4) exhibits significant sequence homology with the SLC7 subfamily of human cationic amino acid transporters (hCATs) [Sperandeo, Borsani, Incerti, Zollo, Rossi, Zuffardi, Castaldo, Taglialatela, Andria and Sebastio (1998) Genomics 49, 230–236]. It is therefore often referred to as hCAT-4 even though no convincing transport activity has been shown for this protein. We expressed SLC7A4 in Xenopus laevis oocytes, but could not detect any transport activity for cationic, neutral or anionic amino acids or for the polyamine putrescine. In addition, human glioblastoma cells stably overexpressing a fusion protein between SLC7A4 and the enhanced green fluorescent protein (EGFP) did not exhibit an increased transport activity for l-arginine. The lack of transport activity was not due to a lack of SLC7A4 protein expression in the plasma membrane, as in both cell types SLC7A4-EGFP exhibited a similar subcellular localization and level of protein expression as functional hCAT-EGFP proteins. The expression of SLC7A4 can be induced in NT2 teratocarcinoma cells by treatment with retinoic acid. However, also for this endogenously expressed SLC7A4, we could not detect any transport activity for l-arginine. Our data demonstrate that the expression of SLC7A4 in the plasma membrane is not sufficient to induce an amino acid transport activity in X. laevis oocytes or human cells. Therefore, SLC7A4 is either not an amino acid transporter or it needs additional (protein) factor(s) to be functional.


Toxins ◽  
2021 ◽  
Vol 13 (5) ◽  
pp. 329
Author(s):  
Andrew Holmes ◽  
Jessie Sadlon ◽  
Keith Weaver

A majority of toxins produced by type I toxin–antitoxin (TA-1) systems are small membrane-localized proteins that were initially proposed to kill cells by forming non-specific pores in the cytoplasmic membrane. The examination of the effects of numerous TA-1 systems indicates that this is not the mechanism of action of many of these proteins. Enterococcus faecalis produces two toxins of the Fst/Ldr family, one encoded on pheromone-responsive conjugative plasmids (FstpAD1) and the other on the chromosome, FstEF0409. Previous results demonstrated that overexpression of the toxins produced a differential transcriptomic response in E. faecalis cells. In this report, we identify the specific amino acid differences between the two toxins responsible for the differential response of a gene highly induced by FstpAD1 but not FstEF0409. In addition, we demonstrate that a transporter protein that is genetically linked to the chromosomal version of the TA-1 system functions to limit the toxicity of the protein.


2021 ◽  
Vol 85 (3) ◽  
pp. 587-599
Author(s):  
Akane Sato ◽  
Takumi Kimura ◽  
Kana Hondo ◽  
Miyuki Kawano-Kawada ◽  
Takayuki Sekito

ABSTRACT In Saccharomyces cerevisiae, Avt4 exports neutral and basic amino acids from vacuoles. Previous studies have suggested that the GATA transcription factors, Gln3 and Gat1, which are key regulators that adapt cells in response to changes in amino acid status, are involved in the AVT4 transcription. Here, we show that mutations in the putative GATA-binding sites of the AVT4 promoter reduced AVT4 expression. Consistently, a chromatin immunoprecipitation (ChIP) assay revealed that Gat1-Myc13 binds to the AVT4 promoter. Previous microarray results were confirmed that gln3∆gat1∆ cells showed a decrease in expression of AVT1 and AVT7, which also encode vacuolar amino acid transporters. Additionally, ChIP analysis revealed that the AVT6 encoding vacuolar acidic amino acid exporter represents a new direct target of the GATA transcription factor. The broad effect of the GATA transcription factors on the expression of AVT transporters suggests that vacuolar amino acid transport is integrated into cellular amino acid homeostasis.


1976 ◽  
Vol 154 (1) ◽  
pp. 43-48 ◽  
Author(s):  
J D Young ◽  
J C Ellory ◽  
E M Tucker

1. Uptake rates for 23 amino acids were measured for both normal (high-GSH) and GSH-deficient (low-GSH) erythrocytes from Finnish Landrace sheep. 2. Compared with high-GSH cells, low-GSH cells had a markedly diminished permeability to D-alanine, L-alanine, α-amino-n-butyrate, valine, cysteine, serine, threonine, asparagine, lysine and ornithine. Smaller differences were observed for glycine and proline, whereas uptake of the other amino acids was not significantly different in the two cell types.


1995 ◽  
Vol 268 (6) ◽  
pp. C1321-C1331 ◽  
Author(s):  
A. J. Moe

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.


1988 ◽  
Vol 255 (3) ◽  
pp. 963-969 ◽  
Author(s):  
A R Quesada ◽  
J D McGivan

A rapid method for the functional reconstruction of amino acid transport from liver plasma-membrane vesicles using the neutral detergent decanoyl-N-glucamide (‘MEGA-10’) is described. The method is a modification of that previously employed in this laboratory for reconstitution of amino acid transport systems from kidney brush-border membranes [Lynch & McGivan (1987) Biochem. J. 244, 503-508]. The transport activities termed ‘System A’, ‘System N’, and ‘System L’ are all reconstituted. The reconstitution procedure is rapid and efficient and is suitable as an assay for transport activity in studies involving membrane fractionation. By using this reconstitution procedure, System A transport activity was partially purified by lectin-affinity chromatography.


1996 ◽  
Vol 199 (4) ◽  
pp. 923-931 ◽  
Author(s):  
P M Taylor ◽  
S Kaur ◽  
B Mackenzie ◽  
G J Peter

We have measured rates of uptake of arginine, glutamine, glutamate, serine, phenylalanine and glycine in Xenopus laevis oocytes cultured for periods of up to 24h in saline in the presence or absence of a mixture of 20 amino acids at concentrations approximating those in Xenopus plasma. Amino acid supplementation increased the total intracellular amino acid concentration from 8.2 to 18.4 nmol per oocyte. Specific Na(+)-dependent amino acid transporters (systems B0,+, Xag-) exhibit 'adaptive regulation' (up-regulation during amino acid deprivation and down-regulation during amino acid supplementation). Na(+)-independent transporters of glutamate, glutamine and glycine (including system asc) display an opposite modulation in activity, which may help to combat amino-acid-induced oxidative stress by increasing the supply of glutathione precursors. Single amino acids at physiological plasma concentrations (0.47 mmol l-1 L-alanine, 0.08 mmol l-1 L-glutamate) mimicked at least some effects of the amino acid mixture. The mechanisms of transport modulation do not appear to include trans-amino acid or membrane potential effects and, in the case of Na(+)-independent transport, are independent of protein or mRNA synthesis. Furthermore, activation of protein kinase C by phorbol 12-myristate 13-acetate did not significantly affect endogenous glutamine and glutamate transport. The Xenopus oocyte appears to possess endogenous signalling mechanisms for selectively modulating the activity of amino acid transport proteins expressed in its surface membranes, a factor for consideration when using oocytes as an expression system for structure-function studies of cloned amino acid transporters.


1998 ◽  
Vol 275 (5) ◽  
pp. G1045-G1055 ◽  
Author(s):  
An-Qiang Sun ◽  
Meenakshisundaram Ananthanarayanan ◽  
Carol J. Soroka ◽  
Sundararajah Thevananther ◽  
Benjamin L. Shneider ◽  
...  

The rat ileal apical Na+-dependent bile acid transporter (ASBT) and the liver Na+-taurocholate cotransporting polypeptide (Ntcp) are members of a new family of anion transporters. These transport proteins share limited sequence homology and almost identical predicted secondary structures but are localized to the apical surface of ileal enterocytes and the sinusoidal surface of hepatocytes, respectively. Stably transfected Madin-Darby canine kidney (MDCK) cells appropriately localized wild-type ASBT and Ntcp apically and basolaterally as assessed by functional activity and immunocytochemical localization studies. Truncated and chimeric transporters were used to determine the functional importance of the cytoplasmic tail in bile acid transport activity and membrane localization. Two cDNAs were created encoding a truncated transporter in which the 56-amino-acid COOH-terminal tail of Ntcp was removed or substituted with an eight-amino-acid epitope FLAG. For both mutants there was some loss of fidelity in basolateral sorting in that ∼75% of each protein was delivered to the basolateral surface compared with ∼90% of the wild-type Ntcp protein. In contrast, deletion of the cytoplasmic tail of ASBT led to complete loss of transport activity and sorting to the apical membrane. An Ntcp chimera in which the 56-amino-acid COOH-terminal tail of Ntcp was replaced with the 40-amino-acid cytoplasmic tail of ASBT was largely redirected (82.4 ± 3.9%) to the apical domain of stably transfected MDCK cells, based on polarity of bile acid transport activity and localization by confocal immunofluorescence microscopy. These results indicate that a predominant signal for sorting of the Ntcp protein to the basolateral domain is located in a region outside of the cytoplasmic tail. These studies have further shown that a novel apical sorting signal is localized to the cytoplasmic tail of ASBT and that it is transferable and capable of redirecting a protein normally sorted to the basolateral surface to the apical domain of MDCK cells.


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