Human nongastric H+-K+-ATPase: transport properties of ATP1al1 assembled with different β-subunits

2002 ◽  
Vol 283 (1) ◽  
pp. C305-C314 ◽  
Author(s):  
Gilles Crambert ◽  
Jean-Daniel Horisberger ◽  
Nikolai N. Modyanov ◽  
Käthi Geering
Keyword(s):  
Cell Surface ◽  
Transport Properties ◽  
Functional Properties ◽  
Conformational Changes ◽  
Xenopus Oocytes ◽  
Structural Integrity ◽  
Catalytic Subunit ◽  
Α Subunit ◽  
Distinct Effect ◽  
New Evidence

To investigate whether nongastric H+-K+-ATPases transport Na+ in exchange for K+ and whether different β-isoforms influence their transport properties, we compared the functional properties of the catalytic subunit of human nongastric H+-K+-ATPase, ATP1al1 (AL1), and of the Na+-K+-ATPase α1-subunit (α1) expressed in Xenopus oocytes, with different β-subunits. Our results show that βHK and β1-NK can produce functional AL1/β complexes at the oocyte cell surface that, in contrast to α1/β1 NK and α1/βHK complexes, exhibit a similar apparent K+ affinity. Similar to Na+-K+-ATPase, AL1/β complexes are able to decrease intracellular Na+ concentrations in Na+-loaded oocytes, and their K+ transport depends on intra- and extracellular Na+ concentrations. Finally, controlled trypsinolysis reveals that β-isoforms influence the protease sensitivity of AL1 and α1 and that AL1/β complexes, similar to the Na+-K+-ATPase, can undergo distinct K+-Na+- and ouabain-dependent conformational changes. These results provide new evidence that the human nongastric H+-K+-ATPase interacts with and transports Na+ in exchange for K+ and that β-isoforms have a distinct effect on the overall structural integrity of AL1 but influence its transport properties less than those of the Na+-K+-ATPase α-subunit.

Unusual degradation of α-β complexes inXenopusoocytes by β-subunits ofXenopusgastric H-K-ATPase

1998 ◽  
Vol 275 (1) ◽  
pp. C139-C145 ◽  
Author(s):  
Pei-Xian Chen ◽  
Paul M. Mathews ◽  
Peter J. Good ◽  
Bernard C. Rossier ◽  
Käthi Geering
Keyword(s):  
Endoplasmic Reticulum ◽  
Cell Surface ◽  
Xenopus Oocytes ◽  
Experimental System ◽  
Surface Expression ◽  
Cell Surface Expression ◽  
Β Subunit ◽  
Α Subunit ◽  
P Type ◽  
Α Subunits

The catalytic α-subunit of oligomeric P-type ATPases such as Na-K-ATPase and H-K-ATPase requires association with a β-subunit after synthesis in the endoplasmic reticulum (ER) to become stably expressed and functionally active. In this study, we have expressed the β-subunit of Xenopus gastric H-K-ATPase (βHK) in Xenopus oocytes together with α-subunits of H-K-ATPase (αHK) or Na-K-ATPase (αNK) and have followed the biosynthesis, assembly, and cell surface expression of functional pumps. Immunoprecipitations of Xenopus βHK from metabolically labeled oocytes show that it is well expressed and, when synthesized without α-subunits, can leave the ER and become fully glycosylated. Xenopus βHK can associate with both coexpressed αHK and αNK, but the α-β complexes formed are degraded rapidly in or close to the ER and do not produce functional pumps at the cell surface as assessed by86Rb uptake. A possible explanation of these results is that Xenopus βHK may contain a tissue-specific signal that is important in the formation or correct targeting of functional α-β complexes in the stomach but that cannot be recognized in Xenopusoocytes and in consequence leads to cellular degradation of the α-β complexes in this experimental system.

Influence of Calcium Ions on the Plant Cell Surface: Membrane Fusions and Conformational Changes

1974 ◽  
Vol 32 ◽  
pp. 154-155
Author(s):  
D. James Morré ◽  
Charles E. Bracker ◽  
William J. VanDerWoude
Keyword(s):  
Cell Wall ◽  
Cell Surface ◽  
Conformational Changes ◽  
Calcium Ions ◽  
Surface Membrane ◽  
Carboxyl Groups ◽  
Cross Linking ◽  
Ultrastructural Evidence ◽  
Glycine Max L ◽  
Wall Extensibility

Calcium ions in the concentration range 5-100 mM inhibit auxin-induced cell elongation and wall extensibility of plant stems. Inhibition of wall extensibility requires that the tissue be living; growth inhibition cannot be explained on the basis of cross-linking of carboxyl groups of cell wall uronides by calcium ions. In this study, ultrastructural evidence was sought for an interaction of calcium ions with some component other than the wall at the cell surface of soybean (Glycine max (L.) Merr.) hypocotyls.

scholarly journals BAK1 Is Not a Target of the Pseudomonas syringae Effector AvrPto

2011 ◽  
Vol 24 (1) ◽  
pp. 100-107 ◽  
Author(s):  
Tingting Xiang ◽  
Na Zong ◽  
Jie Zhang ◽  
Jinfeng Chen ◽  
Mingsheng Chen ◽  
...  
Keyword(s):  
Cell Surface ◽  
Immune Responses ◽  
Pseudomonas Syringae ◽  
Plant Cell ◽  
Crucial Role ◽  
Pathogenic Bacteria ◽  
Effector Proteins ◽  
Receptor Like Kinase ◽  
Receptor Kinases ◽  
New Evidence

Plant cell surface-localized receptor kinases such as FLS2, EFR, and CERK1 play a crucial role in detecting invading pathogenic bacteria. Upon stimulation by bacterium-derived ligands, FLS2 and EFR interact with BAK1, a receptor-like kinase, to activate immune responses. A number of Pseudomonas syringae effector proteins are known to block immune responses mediated by these receptors. Previous reports suggested that both FLS2 and BAK1 could be targeted by the P. syringae effector AvrPto to inhibit plant defenses. Here, we provide new evidence further supporting that FLS2 but not BAK1 is targeted by AvrPto in plants. The AvrPto-FLS2 interaction prevented the phosphorylation of BIK1, a downstream component of the FLS2 pathway.

scholarly journals Vesicular Transport Is Not Required for the Cytoplasmic Pool of Cholera Toxin To Interact with the Stimulatory Alpha Subunit of the Heterotrimeric G Protein

2004 ◽  
Vol 72 (12) ◽  
pp. 6826-6835 ◽  
Author(s):  
Ken Teter ◽  
Michael G. Jobling ◽  
Randall K. Holmes
Keyword(s):  
Plasma Membrane ◽  
Cell Surface ◽  
Cholera Toxin ◽  
G Protein ◽  
Vesicular Transport ◽  
Cytotoxic Action ◽  
Anterograde Transport ◽  
Heterotrimeric G Protein ◽  
Α Subunit ◽  
Vesicular Traffic

ABSTRACT Cholera toxin (CT) moves from the cell surface to the endoplasmic reticulum (ER) by retrograde vesicular transport. The catalytic A1 polypeptide of CT (CTA1) then crosses the ER membrane, enters the cytosol, ADP-ribosylates the stimulatory α subunit of the heterotrimeric G protein (Gsα) at the cytoplasmic face of the plasma membrane, and activates adenylate cyclase. The cytosolic pool of CTA1 may reach the plasma membrane and its Gsα target by traveling on anterograde-directed transport vesicles. We examined this possibility with the use of a plasmid-based transfection system that directed newly synthesized CTA1 to either the ER lumen or the cytosol of CHO cells. Such a system allowed us to bypass the CT retrograde trafficking itinerary from the cell surface to the ER. Previous work has shown that the ER-localized pool of CTA1 is rapidly exported from the ER to the cytosol. Expression of CTA1 in either the ER or the cytosol led to the activation of Gsα, and Gsα activation was not inhibited in transfected cells exposed to drugs that inhibit vesicular traffic. Thus, anterograde transport from the ER to the plasma membrane is not required for the cytotoxic action of CTA1.

scholarly journals Natural Cytotoxicity Receptors: Pattern Recognition and Involvement of Carbohydrates

2005 ◽  
Vol 5 ◽  
pp. 151-154 ◽  
Author(s):  
Angel Porgador
Keyword(s):  
Pattern Recognition ◽  
Cell Membrane ◽  
Cell Surface ◽  
Innate Immune ◽  
Target Cells ◽  
Natural Cytotoxicity ◽  
Tumor Cell Membrane ◽  
Infected Cells ◽  
Natural Cytotoxicity Receptors ◽  
New Evidence

Natural cytotoxicity receptors (NCRs), expressed by natural killer (NK) cells, trigger NK lysis of tumor and virus-infected cells on interaction with cell-surface ligands of these target cells. We have determined that viral hemagglutinins expressed on the surface of virus-infected cells are involved in the recognition by the NCRs, NKp44 and NKp46. Recognition of tumor cells by the NCRs NKp30 and NKp46 involves heparan sulfate epitopes expressed on the tumor cell membrane. Our studies provide new evidence for the identity of the ligands for NCRs and indicate that a broader definition should be applied to pathological patterns recognized by innate immune receptors. Since nonmicrobial endogenous carbohydrate structures contribute significantly to this recognition, there is an imperative need to develop appropriate tools for the facile sequencing of carbohydrate moieties.

Structural and Functional State of Erythrocyte Membranes at Drug Addiction

2021 ◽  
Vol 18 (1) ◽  
pp. 13-19
Author(s):  
A.I. Rabadanova ◽  
Keyword(s):  
Amino Acids ◽  
Drug Addiction ◽  
Functional State ◽  
Conformational Changes ◽  
Structural Integrity ◽  
Fluorescence Analysis ◽  
Heroin Addiction ◽  
Dimensional Structure ◽  
Erythrocyte Membranes ◽  
Drug Addicts

The steady growth in the number of drug addicts, especially among young people, dictates the need to find ways to prevent and treat this disease. In this regard, there is a need for a more detailed study of the mechanisms of the course of this disease using modern research methods, such as atomic force microscopy and fluorescence analysis of amino acid residues. Purpose of the work: to reveal the structural and functional state of erythrocyte membranes in drug addiction. Materials and methods. The studies were carried out on the erythrocyte membranes of 60 subjects suffering from heroin addiction. The shape and topography of the erythrocyte surface were studied, and spectral analysis of the proteins of the erythrocyte membranes was carried out. Results. The conducted AFM studies of erythrocyte membranes indicate the heterogeneity of the surface mechanical properties of the erythrocyte membranes of drug addicts. The data obtained indicate an acceleration of the aging process of erythrocytes in drug addiction, which goes in two ways: the formation of outgrowths on the plasmolemma, which subsequently die off (echinocytes) and invagination of the plasmolemma of erythrocytes (spherocytes). The fluorescence spectrum of amino acids in erythrocytes of drug addicts is characterized by a significant decrease in the intensity of almost all peaks and a shift of the fluorescence peak to the short-wave region. Findings. With drug addiction, changes in the structural integrity of red blood cells are noted. In people with drug addiction, in comparison with healthy people, there is a higher variability of the morphology of erythrocytes, which is expressed in a significant increase in the proportion of echinocytes and spherocytes against the background of a significant decrease in the number of discocytes. For the membrane proteins of erythrocytes of drug addicts, conformational changes are characteristic, manifested in a decrease in the intensity of fluorescence of aromatic amino acids, which indicates their structural modification and significant vulnerability of the hematopoietic system. They are largely determined by changes in the fluorescence intensity of tryptophan and, to a lesser extent, tyrosine, which indicates the preservation of the three-dimensional structure of the protein.

Neural induction and the structure of the target cell surface

Development ◽  
1982 ◽  
Vol 70 (1) ◽  
pp. 171-187
Author(s):  
A. M. Duprat ◽  
L. Gualandris ◽  
P. Rouge
Keyword(s):  
Cell Surface ◽  
Structural Integrity ◽  
Neural Induction ◽  
Active Role ◽  
Target Cells ◽  
Similar Treatment ◽  
Structural Modifications ◽  
Pleurodeles Waltl

Lectins (SBA and PSA) were used to provoke crowding and structural modifications of the presumptive ectoderm cell surface in order to investigate the role of the membrane organization of the competent target cells in neural induction. Are specific characteristics of the cell surface essential for this phenomenon to occur? From amphibian gastrulae, it is possible to obtain neural induction in vitro by association of presumptive ectoderm (target cells) with chordamesoderm (inductor tissue): 4 h of contact is sufficient in Pleurodeles waltl for transmission of the inductive signal. Very quickly, the treatment of the normal ectoderm by lectins (SBA-FITC or PSA-FITC) provoked surface modifications. Lectin-treatment (50 µg ml1−, 30 min) of presumptive ectoderm did not result in any neural induction. Lectin-treatment (50 µg ml1−, 30 min) of presumptive ectoderm previous to its association with the natural inductor for 4 h, disturbed the phenomenon: no induction. Similar treatment followed by association with the inductor for 24 h: induction. Treatment of SBA or PSA with their respective hapten inhibitors prior to addition to ectodermal cells completely blocked the suppressive effects on induction. The structural integrity of the membrane of competent target cells is necessary for neural induction to occur. The cell membrane could thus play, directly or indirectly, an active role in the specificity of this process

scholarly journals Conformational epitopes at cadherin calcium-binding sites and p120-catenin phosphorylation regulate cell adhesion

2012 ◽  
Vol 23 (11) ◽  
pp. 2092-2108 ◽  
Author(s):  
Yuliya I. Petrova ◽  
MarthaJoy M. Spano ◽  
Barry M. Gumbiner
Keyword(s):  
Cell Surface ◽  
Conformational Changes ◽  
Binding Sites ◽  
Calcium Binding ◽  
Cell Types ◽  
P120 Catenin ◽  
Physiological Regulation ◽  
Conformational Epitopes ◽  
Calcium Binding Sites ◽  
E Cadherin

We investigated changes in cadherin structure at the cell surface that regulate its adhesive activity. Colo 205 cells are nonadhesive cells with a full but inactive complement of E-cadherin–catenin complexes at the cell surface, but they can be triggered to adhere and form monolayers. We were able to distinguish the inactive and active states of E-cadherin at the cell surface by using a special set of monoclonal antibodies (mAbs). Another set of mAbs binds E-cadherin and strongly activates adhesion. In other epithelial cell types these activating mAbs inhibit growth factor–induced down-regulation of adhesion and epithelial morphogenesis, indicating that these phenomena are also controlled by E-cadherin activity at the cell surface. Both types of mAbs recognize conformational epitopes at different interfaces between extracellular cadherin repeat domains (ECs), especially near calcium-binding sites. Activation also induces p120-catenin dephosphorylation, as well as changes in the cadherin cytoplasmic domain. Moreover, phospho-site mutations indicate that dephosphorylation of specific Ser/Thr residues in the N-terminal domain of p120-catenin mediate adhesion activation. Thus physiological regulation of the adhesive state of E-cadherin involves physical and/or conformational changes in the EC interface regions of the ectodomain at the cell surface that are mediated by catenin-associated changes across the membrane.

scholarly journals Minimal mechanistic component of proteasome activation and prevention of impairment by pathological oligomers

2021 ◽  
Author(s):  
Janelle Chuah ◽  
Tifffany Thibaudeau ◽  
David Smith
Keyword(s):  
Small Molecules ◽  
Conformational Changes ◽  
Bound State ◽  
Proof Of Concept ◽  
Α Subunit ◽  
Gate Opening ◽  
Allosteric Mechanism ◽  
Dependent Mechanism ◽  
Degradation Of Peptides

Abstract Impairment of proteasomal function has been implicated in neurodegenerative diseases, justifying the need to understand how the proteasome is activated for protein degradation. Here, using biochemical and structural (cryo-EM) strategies in both archaeal and mammalian proteasomes, we further determine the HbYX(-motif)-dependent mechanism of proteasomal activation used by multiple proteasome-activating complexes including the 19S Particle. We identify multiple proteasome α subunit residues involved in HbYX-dependent activation, a point mutation that activates the proteasome by partially mimicking a HbYX-bound state, and conformational changes involved in gate-opening with a 2.0A structure. Through an iterative process of peptide synthesis, we successfully design a HbYX-like dipeptide mimetic as a robust tool to elucidate how the motif autonomously activates the proteasome. The mimetic induces near complete gate-opening at saturating concentration, activating mammalian proteasomal degradation of peptides and proteins. Findings using our peptide mimetic suggest the HbYX-dependent mechanism requires cooperative binding in at least two intersubunit pockets of the α ring. Collectively, the results presented here unambiguously demonstrate the lone role of the HbYX tyrosine in the allosteric mechanism of proteasome activation and offer proof of concept for the robust potential of HbYX-like small molecules to activate the proteasome.

scholarly journals Biophysical and functional properties of purified glucose-6-phosphatase catalytic subunit 1

2021 ◽  
pp. 101520
Author(s):  
Derek P. Claxton ◽  
Emily M. Overway ◽  
James K. Oeser ◽  
Richard M. O’Brien ◽  
Hassane S. Mchaourab
Keyword(s):  
Functional Properties ◽  
Catalytic Subunit
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