scholarly journals Allosteric Regulation of Transport Activity by Heterotrimerization of Arabidopsis Ammonium Transporter Complexes in Vivo

2013 ◽  
Vol 25 (3) ◽  
pp. 974-984 ◽  
Author(s):  
Lixing Yuan ◽  
Riliang Gu ◽  
Yuanhu Xuan ◽  
Erika Smith-Valle ◽  
Dominique Loqué ◽  
...  
Keyword(s):  
Allosteric Regulation ◽  
Ammonium Transporter ◽  
Transport Activity ◽  
Regulation Of Transport

scholarly journals Structure and lipid-mediated remodelling mechanism of the Na+/H+ exchanger NHA2

2021 ◽  
Author(s):  
Rei Matsuoka ◽  
Roman Fudim ◽  
Sukkyeong Jung ◽  
Chenou Zhang ◽  
Andre Bazzone ◽  
...  
Keyword(s):  
Ion Exchange ◽  
Structural Information ◽  
Salt Bridge ◽  
Structural Similarity ◽  
Ion Binding ◽  
Transport Activity ◽  
Regulation Of Transport ◽  
Fine Tune ◽  
Exchange Activity

Na+/H+ exchangers catalyse an ion-exchange activity that is carried out in most, if not all cells. SLC9B2, also known as NHA2, correlates with the long-sought after sodium/lithium (Na+/Li+) exchanger linked to the pathogenesis of diabetes mellitus and essential hypertension in humans. Despite its functional importance, structural information and the molecular basis of its ion-exchange mechanism have been lacking. Here, we report the cryo EM structures of bison NHA2 in detergent and in nanodiscs at 3.0 and 3.5 Å resolution, respectively. NHA2 shares closest structural similarity to the bacterial electrogenic Na+/H+ antiporter NapA, rather than other mammalian SLC9A members. Nevertheless, SSM-based electrophysiology results with NHA2 show the catalysis of electroneutral rather than electrogenic ion exchange, and the ion binding site is quite distinctive, with a tryptophan arginine-glutamate triad separated from the well established ion-binding aspartates. These triad residues fine-tune ion binding specificity, as demonstrated by a salt-bridge swap mutant that converts NHA2 into a Li+ specific transporter. Strikingly, an additional N terminal helix in NHA2 establishes a unique homodimer with a large ~ 25 Å intracellular gap between protomers. In the presence of phosphatidylinositol lipids, the N-terminal helix rearranges and closes this gap. We confirm that dimerization of NHA2 is required for activity in vivo, and propose that the N- terminal helix has evolved as a lipid-mediated remodelling switch for regulation of transport activity.

Assessment of extracellular glucose distribution and glucose transport activity in conscious rats

1995 ◽  
Vol 268 (4) ◽  
pp. E712-E721 ◽  
Author(s):  
J. H. Youn ◽  
J. K. Kim ◽  
G. M. Steil
Keyword(s):  
Glucose Transport ◽  
Insulin Infusion ◽  
Compartmental Analysis ◽  
Time Profile ◽  
Cellular Transport ◽  
Transport Activity ◽  
Conscious Rats ◽  
Distribution Kinetics ◽  
Extracellular Glucose

The effects of insulin on extracellular glucose distribution and cellular glucose transport activity were studied by simultaneously analyzing the plasma kinetics of L-[1-14C]glucose and 3-O-[3H]methylglucose after an intravenous injection during saline or insulin infusion (euglycemic glucose clamp) in conscious rats (n = 7 for each). The time profiles of plasma L-glucose were almost superimposable in the two protocols, and compartmental analysis showed that neither distribution volumes nor distribution rate constants were affected with insulin (P > 0.05 for all), suggesting that glucose distribution within the extracellular space was not influenced with insulin. In contrast, the time profile of plasma 3-O-methylglucose (3-MG) was markedly altered with insulin; the initial decrease was much faster during insulin infusion than during saline infusion, indicating stimulation of 3-MG transport into intracellular spaces with insulin. The 3-MG data were analyzed using a comprehensive model separately describing extracellular distribution and cellular transport of 3-MG by incorporating information on extracellular distribution kinetics obtained from L-glucose data. The combined L-glucose and 3-MG kinetic analysis precisely estimated insulin's effect in vivo to stimulate glucose transport into and out of intracellular spaces. We conclude that 1) insulin does not affect extracellular glucose distribution kinetics or volumes in conscious rats and 2) insulin's effects on cellular glucose transport in vivo can be assessed by simultaneous analysis of plasma L-glucose and 3-MG kinetics.

scholarly journals CALCIUM-DEPENDENT PROTEIN KINASE 32-mediated phosphorylation is essential for the ammonium transport activity of AMT1;1 in Arabidopsis roots

2020 ◽  
Vol 71 (16) ◽  
pp. 5087-5097
Author(s):  
De-Bin Qin ◽  
Meng-Yuan Liu ◽  
Lixing Yuan ◽  
Yun Zhu ◽  
Xi-Dong Li ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Ammonium Transporter ◽  
Ammonium Uptake ◽  
Ammonium Transport ◽  
Toxicity Screening ◽  
Plant Signaling ◽  
Transport Activity ◽  
Calcium Dependent ◽  
Positive Regulator ◽  
Dependent Protein

Abstract Protein kinase-mediated phosphorylation modulates the absorption of many nutrients in plants. CALCIUM-DEPENDENT PROTEIN KINASES (CPKs) are key players in plant signaling to translate calcium signals into diverse physiological responses. However, the regulatory role of CPKs in ammonium uptake remains largely unknown. Here, using methylammonium (MeA) toxicity screening, CPK32 was identified as a positive regulator of ammonium uptake in roots. CPK32 specifically interacted with AMMONIUM TRANSPORTER 1;1 (AMT1;1) and phosphorylated AMT1;1 at the non-conserved serine residue Ser450 in the C-terminal domain. Functional analysis in Xenopus oocytes showed that co-expression of CPK32 and AMT1;1 significantly enhanced the AMT1;1-mediated inward ammonium currents. In transgenic plants, the phosphomimic variant AMT1;1S450E, but not the non-phosphorylatable variant AMT1;1S450A, fully complemented the MeA insensitivity and restored high-affinity 15NH4+ uptake in both amt1;1 and cpk32 mutants. Moreover, in the CPK32 knockout background, AMT1;1 lost its ammonium transport activity entirely. These results indicate that CPK32 is a crucial positive regulator of ammonium uptake in roots and the ammonium transport activity of AMT1;1 is dependent on CPK32-mediated phosphorylation.

scholarly journals 2,4,6-Tribromophenol Exposure Decreases P-Glycoprotein Transport at the Blood-Brain Barrier

2019 ◽  
Vol 171 (2) ◽  
pp. 463-472 ◽  
Author(s):  
Andrew W Trexler ◽  
Gabriel A Knudsen ◽  
Sascha C T Nicklisch ◽  
Linda S Birnbaum ◽  
Ronald E Cannon
Keyword(s):  
Blood Brain Barrier ◽  
Ex Vivo ◽  
Female Rats ◽  
Brain Barrier ◽  
Transport Activity ◽  
Fish Food ◽  
P Glycoprotein ◽  
Blood Brain ◽  
Male And Female Rats

Abstract 2,4,6-Tribromophenol (TBP, CAS No. 118-79-6) is a brominated chemical used in the production of flame-retardant epoxy resins and as a wood preservative. In marine environments, TBP is incorporated into shellfish and consumed by predatory fish. Food processing and water treatment facilities produce TBP as a byproduct. 2,4,6-Tribromophenol has been detected in human blood and breast milk. Biologically, TBP interferes with estrogen and thyroid hormone signaling, which regulate important transporters of the blood-brain barrier (BBB). The BBB is a selectively permeable barrier characterized by brain microvessels which are composed of endothelial cells mortared by tight-junction proteins. ATP-binding cassette (ABC) efflux transporters on the luminal membrane facilitate the removal of unwanted endobiotics and xenobiotics from the brain. In this study, we examined the in vivo and ex vivo effects of TBP on two important transporters of the BBB: P-glycoprotein (P-gp, ABCB1) and Multidrug Resistance-associated Protein 2 (MRP2, ABCC2), using male and female rats and mice. 2,4,6-Tribromophenol exposure ex vivo resulted in a time- (1–3 h) and dose- (1–100 nM) dependent decrease in P-gp transport activity. MRP2 transport activity was unchanged under identical conditions. Immunofluorescence and western blotting measured decreases in P-gp expression after TBP treatment. ATPase assays indicate that TBP is not a substrate and does not directly interact with P-gp. In vivo dosing with TBP (0.4 µmol/kg) produced decreases in P-gp transport. Co-treatment with selective protein kinase C (PKC) inhibitors prevented the TBP-mediated decreases in P-gp transport activity.

Treat and trick: common regulation and manipulation of sugar transporters during sink establishment by the plant and the pathogen

2020 ◽  
Vol 71 (14) ◽  
pp. 3930-3940
Author(s):  
Benjamin Pommerrenig ◽  
Christina Müdsam ◽  
Dominik Kischka ◽  
H Ekkehard Neuhaus
Keyword(s):  
Intracellular Transport ◽  
Sugar Transport ◽  
Transport Proteins ◽  
Dependent Manner ◽  
Transport Activity ◽  
Sugar Transporters ◽  
Dependent Processes ◽  
Sink Establishment ◽  
Source And Sink

Abstract Sugar transport proteins are crucial for the coordinated allocation of sugars. In this Expert View we summarize recent key findings of the roles and regulation of sugar transporters in inter- and intracellular transport by focusing on applied approaches, demonstrating how sucrose transporter activity may alter source and sink dynamics and their identities. The plant itself alters its sugar transport activity in a developmentally dependent manner to either establish or load endogenous sinks, for example, during tuber formation and filling. Pathogens represent aberrant sinks that trigger the plant to induce the same processes, resulting in loss of carbon assimilates. We explore common mechanisms of intrinsic, developmentally dependent processes and aberrant, pathogen-induced manipulation of sugar transport. Transporter activity may also be targeted by breeding or genetic modification approaches in crop plants to alter source and sink metabolism upon the overexpression or heterologous expression of these proteins. In addition, we highlight recent progress in the use of sugar analogs to study these processes in vivo.

scholarly journals SOS2 Promotes Salt Tolerance in Part by Interacting with the Vacuolar H+-ATPase and Upregulating Its Transport Activity

2007 ◽  
Vol 27 (22) ◽  
pp. 7781-7790 ◽  
Author(s):  
Giorgia Batelli ◽  
Paul E. Verslues ◽  
Fernanda Agius ◽  
Quansheng Qiu ◽  
Hiroaki Fujii ◽  
...  
Keyword(s):  
Salt Stress ◽  
Salt Tolerance ◽  
Atpase Activity ◽  
Ion Transport ◽  
Affinity Purification ◽  
Tandem Affinity Purification ◽  
Mass Spectrometry Analysis ◽  
Transport Activity ◽  
Sos Pathway

ABSTRACT The salt overly sensitive (SOS) pathway is critical for plant salt stress tolerance and has a key role in regulating ion transport under salt stress. To further investigate salt tolerance factors regulated by the SOS pathway, we expressed an N-terminal fusion of the improved tandem affinity purification tag to SOS2 (NTAP-SOS2) in sos2-2 mutant plants. Expression of NTAP-SOS2 rescued the salt tolerance defect of sos2-2 plants, indicating that the fusion protein was functional in vivo. Tandem affinity purification of NTAP-SOS2-containing protein complexes and subsequent liquid chromatography-tandem mass spectrometry analysis indicated that subunits A, B, C, E, and G of the peripheral cytoplasmic domain of the vacuolar H+-ATPase (V-ATPase) were present in a SOS2-containing protein complex. Parallel purification of samples from control and salt-stressed NTAP-SOS2/sos2-2 plants demonstrated that each of these V-ATPase subunits was more abundant in NTAP-SOS2 complexes isolated from salt-stressed plants, suggesting that the interaction may be enhanced by salt stress. Yeast two-hybrid analysis showed that SOS2 interacted directly with V-ATPase regulatory subunits B1 and B2. The importance of the SOS2 interaction with the V-ATPase was shown at the cellular level by reduced H+ transport activity of tonoplast vesicles isolated from sos2-2 cells relative to vesicles from wild-type cells. In addition, seedlings of the det3 mutant, which has reduced V-ATPase activity, were found to be severely salt sensitive. Our results suggest that regulation of V-ATPase activity is an additional key function of SOS2 in coordinating changes in ion transport during salt stress and in promoting salt tolerance.

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