Mechanisms activating latent functions of PIP aquaporin water channels via the interaction between PIP1 and PIP2 proteins

2020 ◽  
Author(s):  
Mineo Shibasaka ◽  
Tomoaki Horie ◽  
Maki Katsuhara
Keyword(s):  
Plasma Membrane ◽  
Water Transport ◽  
Molecular Mechanisms ◽  
Water Channel ◽  
Amino Acid Replacement ◽  
Middle Part ◽  
Single Amino Acid ◽  
Xenopus Laevis Oocytes ◽  
Transport Activity ◽  
Membrane Type

Abstract Plant plasma-membrane type PIP aquaporins are classified into two groups, PIP1s and PIP2s. In this study, we focused on HvPIP1; 2, a PIP1 in barley (Hordeum vulgare), to dissect the molecular mechanisms that evoke HvPIP1-mediated water transport. No HvPIP1; 2 protein was localized to the plasma membrane when expressed alone in Xenopus laevis oocytes. In contrast, a chimeric HvPIP1; 2 protein (HvPIP1; 2_24NC), in which the N- and C-terminal regions were replaced with the corresponding regions from HvPIP2; 4, was found to localize to the plasma membrane of oocytes. However, HvPIP1; 2_24NC showed no water transport activity in swelling assays. These results suggested that the terminal regions of PIP2 proteins direct PIP proteins to the plasma-membrane, but the re-localization of PIP1 proteins was not sufficient to PIP1s functionality as water channel in a membrane. A single amino acid replacement of threonine by methionine in HvPIP2; 4 (HvPIP2; 4T229M) abolished water transport activity. Co-expression of HvPIP1; 2_24NC either with HvPIP2; 4_12NC or HvPIP2; 4TM_12NC, in which the N- and C-terminal regions were replaced with the corresponding regions of HvPIP1; 2, increased the water transport activity in oocytes. These data provided evidence that the HvPIP1; 2 molecule has own water transport activity and an interaction with the middle part of the HvPIP2; 4 protein (except for the N- and C- termini) is required for HvPIP1; 2 functionality as water channel. This molecular mechanism could be applied to other PIP1s and PIP2s in addition to the known mechanism that the terminal regions of some PIP2s lead some PIP1s to the plasma membrane.

Role of membrane trafficking in plasma membrane solute transport

1994 ◽  
Vol 267 (1) ◽  
pp. C1-C24 ◽  
Author(s):  
N. A. Bradbury ◽  
R. J. Bridges
Keyword(s):  
Plasma Membrane ◽  
Solute Transport ◽  
Membrane Trafficking ◽  
Molecular Mechanisms ◽  
Glucose Transporter ◽  
Water Channel ◽  
Transport Proteins ◽  
Transmembrane Conductance Regulator ◽  
Transmembrane Conductance ◽  
Regulated Trafficking

Cells can rapidly and reversibly alter solute transport rates by changing the kinetics of transport proteins resident within the plasma membrane. Most notably, this can be brought about by reversible phosphorylation of the transporter. An additional mechanism for acute regulation of plasma membrane transport rates is by the regulated exocytic insertion of transport proteins from intracellular vesicles into the plasma membrane and their subsequent regulated endocytic retrieval. Over the past few years, the number of transporters undergoing this regulated trafficking has increased dramatically, such that what was once an interesting translocation of a few transporters has now become a widespread modality for regulating plasma membrane solute permeabilities. The aim of this article is to review the models proposed for the regulated trafficking of transport proteins and what lines of evidence should be obtained to document regulated exocytic insertion and endocytic retrieval of transport proteins. We highlight four transporters, the insulin-responsive glucose transporter, the antidiuretic hormone-responsive water channel, the urinary bladder H(+)-ATPase, and the cystic fibrosis transmembrane conductance regulator Cl- channel, and discuss the various approaches taken to document their regulated trafficking. Finally, we discuss areas of uncertainty that remain to be investigated concerning the molecular mechanisms involved in regulating the trafficking of proteins.

scholarly journals Expression of solute carrier 7A4 (SLC7A4) in the plasma membrane is not sufficient to mediate amino acid transport activity

2002 ◽  
Vol 364 (3) ◽  
pp. 767-775 ◽  
Author(s):  
Sabine WOLF ◽  
Annette JANZEN ◽  
Nicole VÉKONY ◽  
Ursula MARTINÉ ◽  
Dennis STRAND ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Amino Acid ◽  
Protein Expression ◽  
Amino Acid Transport ◽  
Fluorescent Protein ◽  
Xenopus Laevis Oocytes ◽  
Amino Acid Transporters ◽  
Acid Transport ◽  
Transport Activity ◽  
Solute Carrier

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.

scholarly journals The Cruciality of Single Amino Acid Replacement for the Spectral Tuning of Biliverdin-Binding Cyanobacteriochromes

2020 ◽  
Vol 21 (17) ◽  
pp. 6278
Author(s):  
Keiji Fushimi ◽  
Hiroki Hoshino ◽  
Naeko Shinozaki-Narikawa ◽  
Yuto Kuwasaki ◽  
Keita Miyake ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Molecular Mechanisms ◽  
Near Infrared ◽  
Structural Characteristics ◽  
Blue Shift ◽  
Amino Acid Replacement ◽  
Red Shift ◽  
Single Amino Acid ◽  
Structural Basis ◽  
Small Unit

Cyanobacteriochromes (CBCRs), which are known as linear tetrapyrrole-binding photoreceptors, to date can only be detected from cyanobacteria. They can perceive light only in a small unit, which is categorized into various lineages in correlation with their spectral and structural characteristics. Recently, we have succeeded in identifying specific molecules, which can incorporate mammalian intrinsic biliverdin (BV), from the expanded red/green (XRG) CBCR lineage and in converting BV-rejective molecules into BV-acceptable ones with the elucidation of the structural basis. Among the BV-acceptable molecules, AM1_1870g3_BV4 shows a spectral red-shift in comparison with other molecules, while NpF2164g5_BV4 does not show photoconversion but stably shows a near-infrared (NIR) fluorescence. In this study, we found that AM1_1870g3_BV4 had a specific Tyr residue near the d-ring of the chromophore, while others had a highly conserved Leu residue. The replacement of this Tyr residue with Leu in AM1_1870g3_BV4 resulted in a blue-shift of absorption peak. In contrast, reverse replacement in NpF2164g5_BV4 resulted in a red-shift of absorption and fluorescence peaks, which applies to fluorescence bio-imaging in mammalian cells. Notably, the same Tyr/Leu-dependent color-tuning is also observed for the CBCRs belonging to the other lineage, which indicates common molecular mechanisms.

scholarly journals Syntaxin specificity of aquaporins in the inner medullary collecting duct

2009 ◽  
Vol 297 (2) ◽  
pp. F292-F300 ◽  
Author(s):  
Abinash C. Mistry ◽  
Rickta Mallick ◽  
Janet D. Klein ◽  
Thomas Weimbs ◽  
Jeff M. Sands ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Collecting Duct ◽  
Water Channel ◽  
Snare Complex ◽  
Apical Plasma Membrane ◽  
Transport Activity ◽  
Inner Medullary Collecting Duct ◽  
Syntaxin 4 ◽  
Medullary Collecting Duct ◽  
Syntaxin 3

Proper targeting of the aquaporin-2 (AQP2) water channel to the collecting duct apical plasma membrane is critical for the urine concentrating mechanism and body water homeostasis. However, the trafficking mechanisms that recruit AQP2 to the plasma membrane are still unclear. Snapin is emerging as an important mediator in the initial interaction of trafficked proteins with target soluble N-ethylmaleimide-sensitive factor attachment protein (SNAP) receptor (t-SNARE) proteins, and this interaction is functionally important for AQP2 regulation. We show that in AQP2-Madin-Darby canine kidney cells subjected to adenoviral-mediated expression of both snapin and syntaxins, the association of AQP2 with both syntaxin-3 and syntaxin-4 is highly enhanced by the presence of snapin. In pull-down studies, snapin detected AQP2, syntaxin-3, syntaxin-4, and SNAP23 from the inner medullary collecting duct. AQP2 transport activity, as probed by AQP2's urea permeability, was greatly enhanced in oocytes that were coinjected with cRNAs of SNARE components (snapin+syntaxin-3+SNAP23) over those injected with AQP2 cRNA alone. It was not enhanced when syntaxin-3 was replaced by syntaxin-4 (snapin+syntaxin-4+SNAP23). On the other hand, the latter combination significantly enhanced the transport activity of the related AQP3 water channel while the presence of syntaxin-3 did not. This AQP-syntaxin interaction agrees with the polarity of these proteins' expression in the inner medullary collecting duct epithelium. Thus our findings suggest a selectivity of interactions between different aquaporin and syntaxin isoforms, and thus in the regulation of AQP2 and AQP3 activities in the plasma membrane. Snapin plays an important role as a linker between the water channel and the t-SNARE complex, leading to the fusion event, and the pairing with specific t-SNAREs is essential for the specificity of membrane recognition and fusion.

scholarly journals Aquaporin-1 Facilitates Transmesothelial Water Permeability: In Vitro and Ex Vivo Evidence and Possible Implications in Peritoneal Dialysis

2021 ◽  
Vol 22 (22) ◽  
pp. 12535
Author(s):  
Francesca Piccapane ◽  
Andrea Gerbino ◽  
Monica Carmosino ◽  
Serena Milano ◽  
Arduino Arduini ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Peritoneal Dialysis ◽  
Tight Junctions ◽  
Water Transport ◽  
Water Flux ◽  
Water Channel ◽  
Bathing Solution ◽  
Ussing Chambers ◽  
Aquaporin 1 ◽  
Osmotic Water

We previously showed that mesothelial cells in human peritoneum express the water channel aquaporin 1 (AQP1) at the plasma membrane, suggesting that, although in a non-physiological context, it may facilitate osmotic water exchange during peritoneal dialysis (PD). According to the three-pore model that predicts the transport of water during PD, the endothelium of peritoneal capillaries is the major limiting barrier to water transport across peritoneum, assuming the functional role of the mesothelium, as a semipermeable barrier, to be negligible. We hypothesized that an intact mesothelial layer is poorly permeable to water unless AQP1 is expressed at the plasma membrane. To demonstrate that, we characterized an immortalized cell line of human mesothelium (HMC) and measured the osmotically-driven transmesothelial water flux in the absence or in the presence of AQP1. The presence of tight junctions between HMC was investigated by immunofluorescence. Bioelectrical parameters of HMC monolayers were studied by Ussing Chambers and transepithelial water transport was investigated by an electrophysiological approach based on measurements of TEA+ dilution in the apical bathing solution, through TEA+-sensitive microelectrodes. HMCs express Zo-1 and occludin at the tight junctions and a transepithelial vectorial Na+ transport. Real-time transmesothelial water flux, in response to an increase of osmolarity in the apical solution, indicated that, in the presence of AQP1, the rate of TEA+ dilution was up to four-fold higher than in its absence. Of note, we confirmed our data in isolated mouse mesentery patches, where we measured an AQP1-dependent transmesothelial osmotic water transport. These results suggest that the mesothelium may represent an additional selective barrier regulating water transport in PD through functional expression of the water channel AQP1.

scholarly journals The inhibition of monocarboxylate transporter 2 (MCT2) by AR-C155858 is modulated by the associated ancillary protein

2010 ◽  
Vol 431 (2) ◽  
pp. 217-225 ◽  
Author(s):  
Matthew J. Ovens ◽  
Christine Manoharan ◽  
Marieangela C. Wilson ◽  
Clarey M. Murray ◽  
Andrew P. Halestrap
Keyword(s):  
Plasma Membrane ◽  
Mammalian Cells ◽  
Monocarboxylate Transporter ◽  
Xenopus Laevis Oocytes ◽  
Transport Activity ◽  
Membrane Expression ◽  
Binding Partner ◽  
Reverse Transcription Pcr ◽  
C Terminus ◽  
Plasma Membrane Expression

In mammalian cells, MCTs (monocarboxylate transporters) require association with an ancillary protein to enable plasma membrane expression of the active transporter. Basigin is the preferred binding partner for MCT1, MCT3 and MCT4, and embigin for MCT2. In rat and rabbit erythrocytes, MCT1 is associated with embigin and basigin respectively, but its sensitivity to inhibition by AR-C155858 was found to be identical. Using RT (reverse transcription)–PCR, we have shown that Xenopus laevis oocytes contain endogenous basigin, but not embigin. Co-expression of exogenous embigin was without effect on either the expression of MCT1 or its inhibition by AR-C155858. In contrast, expression of active MCT2 at the plasma membrane of oocytes was significantly enhanced by co-expression of exogenous embigin. This additional transport activity was insensitive to inhibition by AR-C155858 unlike that by MCT2 expressed with endogenous basigin that was potently inhibited by AR-C155858. Chimaeras and C-terminal truncations of MCT1 and MCT2 were also expressed in oocytes in the presence and absence of exogenous embigin. L-Lactate Km values for these constructs were determined and revealed that the TM (transmembrane) domains of an MCT, most probably TM7–TM12, but not the C-terminus, are the major determinants of L-lactate affinity, whereas the associated ancillary protein has little or no effect. Inhibitor titrations of lactate transport by these constructs indicated that embigin modulates MCT2 sensitivity to AR-C155858 through interactions with both the intracellular C-terminus and TMs 3 and 6 of MCT2. The C-terminus of MCT2 was found to be essential for its expression with endogenous basigin.

Aquaporins in the plasma membrane of leaf callus protoplasts of Actinidia deliciosa var. deliciosa cv. Hayward

2000 ◽  
Vol 27 (1) ◽  
pp. 71
Author(s):  
Quan-Sheng Qiu ◽  
Ze-Zhou Wang ◽  
Nang Zhang ◽  
Qi-Gui Cai ◽  
Rong-Xi Jiang
Keyword(s):  
Plasma Membrane ◽  
Water Transport ◽  
Imaging System ◽  
Inhibitory Effect ◽  
Actinidia Deliciosa ◽  
Transport Activity ◽  
Water Permeability Coefficient ◽  
Osmotic Water ◽  
A Cell ◽  
Protoplast Volume

The water transport activity of Actinidia deliciosa protoplasts was determined using a cell imaging system. Results showed that the protoplast volume increased swiftly when placed in a hypoton-ic medium, and also increased with an increase in medium osmotic gradients. The osmotic water permeability coefficient (Pf) values were 0.118 × 10–3, 0.121 × 10–3, and 0.133 × 10–3 cm s–1 when the osmotic gradients were 75, 100, and 125 mosmol, respectively. The water transport activity of protoplas-ts could be inhibited by HgCl2 and stimulated by amphotericin B. Moreover, ZnCl2 and ZnSO4 had a significant inhibitory effect on the water transport activity of the protoplasts. Our results indicate that the Actinidia deliciosa protoplasts had properties typical of aquaporins, suggesting that aquaporins were present at the plasma membrane.

scholarly journals Lovastatin-induced cholesterol depletion affects both apical sorting and endocytosis of aquaporin-2 in renal cells

2010 ◽  
Vol 298 (2) ◽  
pp. F266-F278 ◽  
Author(s):  
G. Procino ◽  
C. Barbieri ◽  
M. Carmosino ◽  
F. Rizzo ◽  
G. Valenti ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Molecular Mechanisms ◽  
Collecting Duct ◽  
Rat Kidney ◽  
Water Channel ◽  
Membrane Rafts ◽  
Apical Plasma Membrane ◽  
Cholesterol Depletion ◽  
Aquaporin 2 ◽  
Apical Sorting

Vasopressin causes the redistribution of the water channel aquaporin-2 (AQP2) from cytoplasmic storage vesicles to the apical plasma membrane of collecting duct principal cells, leading to urine concentration. The molecular mechanisms regulating the selective apical sorting of AQP2 are only partially uncovered. In this work, we investigate whether AQP2 sorting/trafficking is regulated by its association with membrane rafts. In both MCD4 cells and rat kidney, AQP2 preferentially associated with Lubrol WX-insoluble membranes regardless of its presence in the storage compartment or at the apical membrane. Block-and-release experiments indicate that 1) AQP2 associates with detergent-resistant membranes early in the biosynthetic pathway; 2) strong cholesterol depletion delays the exit of AQP2 from the trans-Golgi network. Interestingly, mild cholesterol depletion promoted a dramatic accumulation of AQP2 at the apical plasma membrane in MCD4 cells in the absence of forskolin stimulation. An internalization assay showed that AQP2 endocytosis was clearly reduced under this experimental condition. Taken together, these data suggest that association with membrane rafts may regulate both AQP2 apical sorting and endocytosis.

cDNA cloning of a functional water channel from toad urinary bladder epithelium

1996 ◽  
Vol 271 (5) ◽  
pp. C1699-C1704 ◽  
Author(s):  
T. Ma ◽  
B. Yang ◽  
A. S. Verkman
Keyword(s):  
Amino Acid ◽  
Urinary Bladder ◽  
Water Channel ◽  
Single Amino Acid ◽  
Xenopus Laevis Oocytes ◽  
Toad Urinary Bladder ◽  
Bladder Epithelium ◽  
Base Pairs ◽  
Reading Frame ◽  
Acid Protein

A cDNA was cloned from the epithelium of toad (Bufo marinas) urinary bladder, based on homology to the mammalian aquaporins (AQP). The cDNA [947 base pairs (bp), identified as AQP-t1] encoded a 272-amino acid protein with 76% identity to mammalian aquaporin-1 (AQP-1) and 88% identity to frog water channel FA-CHIP. AQP-t1 cDNA was nearly identical to a fragment of a nonfunctional cDNA cloned recently from toad bladder ["AQP-TB"; J. Siner, A. Paredes, C. Hosselet, T. Hammond, K. Strange, and H.W. Harris, Am. J. Physiol. 270 (Cell Physiol. 39): C372-C381, 1996], except for reading frame shifts at bp 253, 264, and 682, two single amino acid deletions, a different 3'-coding sequence downstream from bp 786, and a different 5' sequence upstream from bp 9. Water permeability (Pf) in Xenopus laevis oocytes expressing AQP-t1 cRNA was strongly increased from (0.83 +/- 0.06) x 10(-3) cm/s (water-injected control) to (17 +/- 4) x 10(-3) cm/s, with 80% inhibition by 0.3 mM HgCl2; glycerol and urea permeabilities were not increased. Northern blot analysis showed a single AQP-t1 mRNA of 2.8 kb in eye > lung > urinary bladder > skin > stomach approximately heart, brain, and intestine. AQP-t1 mRNA expression was not changed by a 3-day dehydration of toads or an 8-h stimulation of Pf in isolated bladders by forskolin. These results indicate that the epithelium of toad urinary bladder expresses a functional homologue of AQP-1 and FA-CHIP that is probably not vasopressin regulated.

N-glycosylation controls functional activity of Oatp1, an organic anion transporter

2003 ◽  
Vol 285 (2) ◽  
pp. G371-G381 ◽  
Author(s):  
Thomas K. Lee ◽  
Albert S. Koh ◽  
Zhifeng Cui ◽  
Robert H. Pierce ◽  
Nazzareno Ballatori
Keyword(s):  
Plasma Membrane ◽  
Functional Activity ◽  
Plasma Membranes ◽  
Organic Anion ◽  
Xenopus Laevis Oocytes ◽  
Transport Activity ◽  
Anion Transporter ◽  
Glycosylation Sites ◽  
Quadruple Mutant ◽  
Taurocholate Transport

Rat Oatp1 (Slc21a1) is an organic anion-transporting polypeptide believed to be an anion exchanger. To characterize its mechanism of transport, Oatp1 was expressed in Saccharomyces cerevisiae under control of the GAL1 promoter. Protein was present at high levels in isolated S. cerevisiae secretory vesicles but had minimal posttranslational modifications and failed to exhibit taurocholate transport activity. Apparent molecular mass ( M) of Oatp1 in yeast was similar to that of unmodified protein, ∼62 kDa, whereas in liver plasma membranes Oatp1 has an M of ∼85 kDa. To assess whether underglycosylation of Oatp1 in yeast suppressed functional activity, Oatp1 was expressed in Xenopus laevis oocytes with and without tunicamycin, a glycosylation inhibitor. With tunicamycin, M of Oatp1 decreased from ∼72 to ∼62 kDa and transport activity was nearly abolished. Mutations to four predicted N-glycosylation sites on Oatp1 (Asn to Asp at positions 62, 124, 135, and 492) revealed a cumulative effect on function of Oatp1, leading to total loss of taurocholate transport activity when all glycosylation sites were removed. M of the quadruple mutant was ∼ 62 kDa, confirming that these asparagine residues are sites of glycosylation in Oatp1. Relatively little of the quadruple mutant was able to reach the plasma membrane, and most remained in unidentified intracellular compartments. In contrast, two of the triple mutants tested (N62/124/135D and N124/135/492D) were present in the plasma membrane fraction yet exhibited minimal transport activity. These results demonstrate that both membrane targeting and functional activity of Oatp1 are controlled by the extent of N-glycosylation.

Sign in / Sign up

Export Citation Format

Share Document