共查询到20条相似文献,搜索用时 701 毫秒
1.
Anske Van den Abbeele Sarah De Clercq Ariane De Ganck Veerle De Corte Berlinda Van Loo Sameh Hamdy Soror Vasundara Srinivasan Jan Steyaert Joël Vandekerckhove Jan Gettemans 《Cellular and molecular life sciences : CMLS》2010,67(9):1519-1535
RNA interference has tremendously advanced our understanding of gene function but recent reports have exposed undesirable
side-effects. Recombinant Camelid single-domain antibodies (VHHs) provide an attractive means for studying protein function without affecting gene expression.
We raised VHHs against gelsolin (GsnVHHs), a multifunctional actin-binding protein that controls cellular actin organization
and migration. GsnVHH-induced delocalization of gelsolin to mitochondria or the nucleus in mammalian cells reveals distinct
subpopulations including free gelsolin and actin-bound gelsolin complexes. GsnVHH 13 specifically recognizes Ca2+-activated gelsolin (K
d ~10 nM) while GsnVHH 11 binds gelsolin irrespective of Ca2+ (K
d ~5 nM) but completely blocks its interaction with G-actin. Both GsnVHHs trace gelsolin in membrane ruffles of EGF-stimulated
MCF-7 cells and delay cell migration without affecting F-actin severing/capping or actin nucleation activities by gelsolin.
We conclude that VHHs represent a potent way of blocking structural proteins and that actin nucleation by gelsolin is more
complex than previously anticipated. 相似文献
2.
M. Appel J. Davy G. Durand D. Biou J. Feger J. Agneray 《Cellular and molecular life sciences : CMLS》1984,40(12):1443-1445
Summary A method for measuring proteins in low concentrations applying the zone immunoelectrophoresis assay is reported. The low detection limit makes it possible to measure 1-acid glycoprotein in rat serum and also to quantify the secretion of this protein after concentration of the incubation media containing less than 107 isolated rat hepatocytes. The method is simple and consumes very small quantities of antiserum. 相似文献
3.
C. J. Duncan 《Cellular and molecular life sciences : CMLS》1990,46(1):41-48
Summary The O2– and Ca2+-paradoxes have a number of features in common and it is suggested that release of cytosolic proteins in both paradoxes is initiated by the activation of a sarcolemma NAD(P)H dehydrogenase which can generate a transmembrane flow of H+ and e– and also oxygen radicals or recox cycling which damage ion channels and membrane proteins (phase I). Entry of Ca2+ through the damaged ion channels then exacerbates the damage by further activating this system, either directly or indirectly, and the redox cycling and/or oxygen radicals cause further damage to integral and cytoskeletal proteins of the sarcolemma resulting in microdamage to the integrity of the membrane (phase II) and the consequent release or exocytosis of cytoplasmic proteins and, under specialised condition, the blebbing of the sarcolemma. The system may be primed either by removal of extracellular Ca2+ or by raising [Ca2+]i by a variety of measures, these two actions being synergistic. The system is initially activated in the Ca2+-paradox by the membrane perturbation associated with removal of extracellular Ca2+; prolonged anoxia in the metabolically active cardiac muscle causes a depletion of the ATP supply, particularly in the absence of glucose, and hence a rise in [Ca2+]i in phase I of the oxygen paradox with the consequent activation of the NAD(P)H oxidase at the sarcolemma. Oxygen radicals are probably generated in both paradoxes and may have a partial role in the genesis of damage, but are not essential in the Ca2+-paradox which continues under anoxia. Massive entry of Ca2+ also activates an intracellularly localised dehydrogenase (probably at the SR) which produces myofilament damage by redox cycling. 相似文献
4.
M. Pucéat 《Cellular and molecular life sciences : CMLS》1999,55(10):1216-1229
Intracellular pH (pHi) is a major regulator of various and critical cellular functions. A close regulation of pHi is thus mandatory to maintain normal cellular activity. To this end, all cells express ion transporters that carry across
their plasma membrane H+ or equivalent H+ into and out of the cell. Besides pHi, these ion transporters are under the regulation of neurohormonal stimuli. This review summarises the molecular identity,
regulation and function of the main membrane pH-regulatory ion transporters.
Received 30 December 1998; received after revision 4 February 1999; accepted 9 February 1999 相似文献
5.
M. L. Pressler 《Cellular and molecular life sciences : CMLS》1987,43(10):1084-1091
Summary Internal longitudinal resistance (ri), a determinant of cardiac conduction, is affected by changes in intracellular calcium and protons. However, the role and mechanism by which H+ and Ca2+ may modulate ri is uncertain. Cable analysis was performed in cardiac Purkinje fibers to measure ri during various interventions. In some experiments, intracellular pH (pHi) was recorded simultaneously to study the pHi-ri relation. Both intracellular Ca2+ and H+ independently modified ri. However, internal resistance of cardiac fibers was insensitive to pHi changes compared to other tissues. A latent period preceded the pHi-related changes in ri and the amount of change depended upon methodology. The results suggest that direct action of protons on ri may be subordinate to other regulatory processes. Ionic regulation of internal longitudinal resistance may occur by more than one mechanism: i) direct cationic binding to sites on junctional membrane proteins; and ii) H+- or Ca2+-dependent phosphorylation of junctional proteins. 相似文献
6.
Endomannosidase is a Golgi-localized endoglycosidase, which provides an alternate glucosidase-independent pathway of glucose trimming. Using a protease protection assay we demonstrated that Golgi-endomannosidase is a type II membrane protein. The first 25 amino acids of this protein, containing the cytoplasmic tail and the transmembrane domain, were sufficient for Golgi retention of fused reporter proteins alpha1-antitrypsin or green fluorescent protein. However, shortening or deletion of the transmembrane domain prevented Golgi localization, while lengthening it partially reduced Golgi retention of the enzyme. Substitution of the highly conserved positively charged amino acids within the cytoplasmic tail had neither an effect on type II topology nor on the inherent Golgi localization of the enzyme. In contrast, cytoplasmic tail-deleted rat endomannosidase possessed an inverted topology resulting in endoplasmic reticulum mislocalization. Thus, proper topology rather than the presence of positively charged amino acids in the cytoplasmic tail is critical for Golgi localization of rat endomannosidase. 相似文献
7.
Zhang LJ Wang XE Peng X Wei YJ Cao R Liu Z Xiong JX Yin XF Ping C Liang S 《Cellular and molecular life sciences : CMLS》2006,63(15):1790-1804
To characterize low-copy integral membrane proteins and offer some methods for human liver proteome projects, we fractionated
highly purified rat liver plasma membrane (PM). PM was purified through two sucrose density gradient centrifugations, and
treated with 0.1 M Na2CO3, chloroform/methanol and Triton X-100. Proteins were separated by electrophoresis and submitted to mass spectrometry analysis.
Four hundred and fiftyseven non-redundant membrane proteins were identified, of which 23% (105) were integral membrane proteins
with one or more transmembrane domains. One hundred and fifty-three (33.5%) had no location annotation and 68 were unknown-function
proteins. The proteins from different fractions were complementory. A database search for all identified proteins revealed
that 53 proteins were involved in the cell communication pathway. More interestingly, more than 50% of the proteins had a
protein abundance index concentration of less than 0.1 mol/l, and 12% proteins a concentration 100 times less than that of
arginase 1 and actin.
Received 15 March 2006; received after revision 17 May 2006; accepted 10 June 2006
L.-J. Zhang and X.-e Wang are contributed equally to this work. 相似文献
8.
Dekki N Refai E Holmberg R Köhler M Jörnvall H Berggren PO Juntti-Berggren L 《Cellular and molecular life sciences : CMLS》2012,69(10):1733-1743
Transthyretin (TTR) is a functional protein in the pancreatic β-cell. It promotes insulin release and protects against β-cell
death. We now demonstrate by ligand blotting, adsorption to specific magnetic beads, and surface plasmon resonance that TTR
binds to glucose-regulated proteins (Grps)78, 94, and 170, which are members of the endoplasmic reticulum chaperone family,
but Grps78 and 94 have also been found at the plasma membrane. The effect of TTR on changes in cytoplasmic free Ca2+ concentration ([Ca2+]i) was abolished if the cells were treated with either dynasore, a specific inhibitor of dynamin GTPase that blocks clathrin-mediated
endocytosis, or an antibody against Grp78, that prevents TTR from binding to Grp78. The conclusion is that TTR binds to Grp78
at the plasma membrane, is internalized into the β-cell via a clathrin-dependent pathway, and that this internalization is
necessary for the effects of TTR on β-cell function. 相似文献
9.
The annexins: spatial and temporal coordination of signaling events during cellular stress 总被引:2,自引:0,他引:2
Katia Monastyrskaya Eduard B. Babiychuk Annette Draeger 《Cellular and molecular life sciences : CMLS》2009,66(16):2623-2642
Annexins are a family of structurally related, Ca2+-sensitive proteins that bind to negatively charged phospholipids and establish specific interactions with other lipids and
lipid microdomains. They are present in all eukaryotic cells and share a common folding motif, the “annexin core”, which incorporates
Ca2+- and membrane-binding sites. Annexins participate in a variety of intracellular processes, ranging from the regulation of
membrane dynamics to cell migration, proliferation, and apoptosis. Here we focus on the role of annexins in cellular signaling
during stress. A chronic stress response triggers the activation of different intracellular pathways, resulting in profound
changes in Ca2+ and pH homeostasis and the production of lipid second messengers. We review the latest data on how these changes are sensed
by the annexins, which have the ability to simultaneously interact with specific lipid and protein moieties at the plasma
membrane, contributing to stress adaptation via regulation of various signaling pathways. 相似文献
10.
Sánchez-Margalet V González-Yanes C Santos-Alvarez J Najib S 《Cellular and molecular life sciences : CMLS》1999,55(1):142-147
Insulin action is initiated by binding to its cognate receptor, which then triggers multiple cellular responses by activating
different signaling pathways. There is evidence that insulin receptor signaling may involve G protein activation in different
target cells. We have studied the activation of G proteins in rat hepatoma (HTC) cells. We found that insulin stimulated binding
of guanosine 5′-O-(3-thiotriphosphate) (GTP-γ-35S) to plasma membrane proteins of HTC cells, in a dose-dependent manner. This effect was completely blocked by pertussis toxin
treatment of the membranes, suggesting the involvement of G proteins of the Gα
i/Gα
o family. The expression of these Gα proteins was checked by Western blotting. Next, we used blocking antibodies to sort out the specific Gα protein activated by insulin stimulation. Anti-Gα
il,2 antibodies completely prevented insulin-stimulated GTP binding, whereas anti-Gα
o,i3 did not modify this effect of insulin on GTP binding. Moreover, we found physical association of the insulin receptor with
Gα
i1,2 by copurification studies. These results further support the involvement of a pertussis toxin-sensitive G protein in insulin
receptor signaling and provides some evidence of specific association and activation of Gα
i1,2 protein by insulin. These findings suggest that Gα
i1,2 proteins might be involved in insulin action.
Received 23 September 1998; received after revision 23 November 1998; accepted 25 November 1998 相似文献
11.
Autophagic degradation of cytoplasm (including protein, RNA etc.) is a non-selective bulk process, as indicated by ultrastructural evidence and by the similarity in autophagic sequestration rates of various cytosolic enzymes with different half-lives. The initial autophagic sequestration step, performed by a poorly-characterized organelle called a phagophore, is subject tofeedback inhibition by purines and amino acids, the effect of the latter being potentiated by insulin and antagonized by glucagon. Epinephrine and other adrenergic agonists inhibit autophagic sequestration through a prazosin-sensitive 1-adrenergic mechanism. The sequestration is also inhibited by cAMP and by protein phosphorylation as indicated by the effects of cyclic nucleotide analogues, phosphodiesterase inhibitors and okadaic acid.Asparagine specifically inhibits autophagic-lysosomal fusion without having any significant effects on autophagic sequestration, on intralysosomal degradation or on the endocytic pathway. Autophaged material that accumulates in prelysosomal vacuoles in the presence of asparagine is accessible to endocytosed enzymes, revealing the existence of an amphifunctional organelle, the amphisome. Evidence from several cell types suggests that endocytosis may be coupled to autophagy to a variable extent, and that the amphisome may play a central role as a collecting station for material destined for lysosomal degradation.Protein degradation can also take place in a salvage compartment closely associated with the endoplasmic reticulum (ER). In this compartment unassembled protein chains are degraded by uncharacterized proteinases, while resident proteins roturn to the ER and assembled secretory and membrane proteins proceed through the Golgi apparatus. In thetrans-Golgi network some proteins are proteolytically processed by Ca2+-dependent proteinases; furthermore, this compartment sorts proteins to lysosomes, various membrane domains, endosomes or secretory vesicles/granules. Processing of both endogenous and exogenous proteins can occurr in endosomes, which may play a particularly important role in antigen processing and presentation. Proteins in endosomes or secretory compartments can either be exocytosed, or channeled to lysosomes for degradation. The switch mechanisms which decide between these options are subject to bioregulation by external agents (hormones and growth factors), and may play an important role in the control of protein uptake and secretion. 相似文献
12.
The Membrane Protein Data Bank (MPDB) is an online, searchable, relational database of structural and functional information
on integral, anchored and peripheral membrane proteins and peptides. Data originates from the Protein Data Bank and other
databases, and from the literature. Structures are based on X-ray and electron diffraction, nuclear magnetic resonance and
cryoelectron microscopy. The MPDB is searchable online by protein characteristic, structure determination method, crystallization
technique, detergent, temperature, pH, author, etc. Record entries are hyperlinked to the PDB and Pfam for viewing sequence,
three-dimensional structure and domain architecture, and for downloading coordinates. Links to PubMed are also provided. The
MPDB is updated weekly in parallel with the Protein Data Bank. Statistical analysis of MPDB records can be performed and viewed
online. A summary of the statistics as applied to entries in the MPDB is presented. The data suggest conditions appropriate
for crystallization trials with novel membrane proteins.
Received 3 August 2005; received after revision 18 September 2005; accepted 26 September 2005
This paper and the Membrane Protein Data Bank celebrate the 20th anniversary of the landmark paper in Nature (1985, 318: 618–624) describing the first ‘high-resolution’ three-dimensional structure of a membrane protein, the photosynthetic reaction
center from Rhodopseudomonas (Blastochloris) viridis. 相似文献
13.
Dehydroepiandrosterone sulfate (DHA-S) plays a critical role in cervical dilation at labor. Incubation of cervical fibroblasts with [3H]DHA-S caused a rapid and saturable increase in cellular radioactivity: an apparent equilibrium was reached by 2 min. There was no detectable conversion of DHA-S into DHA or oestradiol. When the fibroblasts loaded with [3H]DHA-S were homogenized and fractionated, the specific radioactivity in the plasma membrane fraction was enriched approximately 8- to 9-fold compared with the whole homogenate; only low amounts of radioactivity were observed in the other subcellular fractions. The binding of DHA-S to plasma membrane preparations showed saturation kinetics with an apparent equilibrium dissociation constant (K
d) of 12 nM, and the binding capacity (B
max) was calculated to be 1.25 fmol/mg protein. Neither DHA nor oestrone sulfate affected [3H]DHA-S binding to the plasma membrane. The plasma membranes of skin fibroblasts did not show specific binding sites for DHA-S. These findings demonstrate the presence of specific binding sites for DHA-S in the plasma membrane of cervical stroma cells. The fetal adrenal steroid may exert its action on cervical ripening at least in part through membrane-associated binding sites, or receptors. 相似文献
14.
Jiraporn Ousingsawat Inês Cabrita Podchanart Wanitchakool Lalida Sirianant Stefan Krautwald Andreas Linkermann Rainer Schreiber Karl Kunzelmann 《Cellular and molecular life sciences : CMLS》2017,74(1):173-181
Activated receptor-interacting protein kinase 3 (RIPK3) and mixed lineage kinase domain like (MLKL) are essential components of the necroptotic pathway. Phosphorylated MLKL (pMLKL) is thought to induce membrane leakage, leading to cell swelling and disintegration of the cell membrane. However, the molecular identity of the necroptotic membrane pore remains unclear, and the role of pMLKL for membrane permeabilization is currently disputed. We observed earlier that the phospholipid scramblase and ion channel TMEM16F/anoctamin 6 cause large membrane currents, cell swelling, and cell death when activated by a strong increase in intracellular Ca2+. We, therefore, asked whether TMEM16F is also central to necroptotic cell death and other cellular events during necroptosis. Necroptosis was induced by TNFα, smac mimetic, and Z-VAD (TSZ) in NIH3T3 fibroblasts and the four additional cell lines HT29, 16HBE, H441, and L929. Time-dependent changes in intracellular Ca2+, cell morphology, and membrane currents were recorded. TSZ induced a small and only transient oscillatory rise in intracellular Ca2+, which was paralleled by the activation of outwardly rectifying Cl? currents, which were typical for TMEM16F/ANO6. Ca2+ oscillations were due to Ca2+ release from endoplasmic reticulum, and were independent of extracellular Ca2+. The initial TSZ-induced cell swelling was followed by cell shrinkage. Using typical channel blockers and siRNA-knockdown, the Cl? currents were shown to be due to the activation of ANO6. However, the knockdown of ANO6 or inhibitors of ANO6 did not inhibit necroptotic cell death. The present data demonstrate the activation of ANO6 during necroptosis, which, however, is not essential for cell death. 相似文献
15.
Senchou V Weide R Carrasco A Bouyssou H Pont-Lezica R Govers F Canut H 《Cellular and molecular life sciences : CMLS》2004,61(4):502-509
The RGD tripeptide sequence, a cell adhesion motif present in several extracellular matrix proteins of mammalians, is involved in numerous plant processes. In plant-pathogen interactions, the RGD motif is believed to reduce plant defence responses by disrupting adhesions between the cell wall and plasma membrane. Photoaffinity cross-linking of [125I]-azido-RGD heptapeptide in the presence of purified plasma membrane vesicles of Arabidopsis thaliana led to label incorporation into a single protein with an apparent molecular mass of 80 kDa. Incorporation could be prevented by excess RGD peptides, but also by the IPI-O protein, an RGD-containing protein secreted by the oomycete plant pathogen Phytophthora infestans. Hydrophobic cluster analysis revealed that the RGD motif of IPI-O (positions 53–56) is readily accessible for interactions. Single amino acid mutations in the RGD motif in IPI-O (of Asp56 into Glu or Ala) resulted in the loss of protection of the 80-kDa protein from labelling. Thus, the interaction between the two proteins is mediated through RGD recognition and the 80-kDa RGD-binding protein has the characteristics of a receptor for IPI-O. The IPI-O protein also disrupted cell wall-plasma membrane adhesions in plasmolysed A. thaliana cells, whereas IPI-O proteins mutated in the RGD motif (D56A and D56E) did not.Received 23 October 2003; received after revision 5 December 2003; accepted 12 December 2003 相似文献
16.
Arada Vinaiphat Visith Thongboonkerd 《Cellular and molecular life sciences : CMLS》2018,75(8):1461-1482
Three isoforms of plasma membrane Ca2+-ATPase (PMCA) are expressed in the kidney. While PMCA1 and PMCA4 play major role in regulating Ca2+ reabsorption, the role for PMCA2 remains vaguely defined. To define PMCA2 function, PMCA2-interacting complex was characterized by immunoprecipitation followed by nanoLC-ESI-Qq-TripleTOF MS/MS (IP-MS). After subtracting non-specific binders using isotype-controlled IP-MS, 474 proteins were identified as PMCA2-interacting partners. Among these, eight were known and 20 were potential PMCA2-interacting partners based on bioinformatic prediction, whereas other 446 were novel and had not been previously reported/predicted. Quantitative immuno-co-localization assay confirmed the association of PMCA2 with these partners. Gene ontology analysis revealed binding activity as the major molecular function of PMCA2-interacting complex. Functional validation using calcium oxalate monohydrate (COM) crystal-protein binding, crystal-cell adhesion, and crystal internalization assays together with neutralization by anti-PMCA2 antibody compared to isotype-controlled IgG and blank control, revealed a novel role of PMCA2 as a COM crystal-binding protein that was crucial for crystal retention and uptake. In summary, a large number of novel PMCA2-interacting proteins have been defined and a novel function of PMCA2 as a COM crystal-binding protein sheds light onto its involvement, at least in part, in kidney stone pathogenesis. 相似文献
17.
Oxysterol-binding protein/OSBP-related proteins (ORPs) constitute a conserved family of sterol/phospholipid-binding proteins with lipid transporter or sensor functions. We investigated the spatial occurrence and regulation of the interactions of human OSBP/ORPs or the S. cerevisiae orthologs, the Osh (OSBP homolog) proteins, with their endoplasmic reticulum (ER) anchors, the VAMP-associated proteins (VAPs), by employing bimolecular fluorescence complementation and pull-down set-ups. The ORP–VAP interactions localize frequently at distinct subcellular sites, shown in several cases to represent membrane contact sites (MCSs). Using established ORP ligand-binding domain mutants and pull-down assays with recombinant proteins, we show that ORP liganding regulates the ORP–VAP association, alters the subcellular targeting of ORP–VAP complexes, or modifies organelle morphology. There is distinct protein specificity in the effects of the mutants on subcellular targeting of ORP–VAP complexes. We provide evidence that complexes of human ORP2 and VAPs at ER–lipid droplet interfaces regulate the hydrolysis of triglycerides and lipid droplet turnover. The data suggest evolutionarily conserved, complex ligand-dependent functions of ORP–VAP complexes at MCSs, with implications for cellular lipid homeostasis and signaling. 相似文献
18.
E. Sick N. Niederhoffer K. Takeda Y. Landry J.-P. Gies 《Cellular and molecular life sciences : CMLS》2009,66(7):1271-1282
Mast cells play pivotal roles in allergic and inflammatory processes via distinct activation pathways. Mucosal and serosal mast cells are activated by the IgE/FcɛRI pathway, while only serosal mast
cells are activated by basic secretagogues. We show that CD47 receptors are expressed on rat peritoneal mast cells. 4N1K,
a peptide agonist of CD47, rapidly caused exocytosis. Such exocytosis required increased intracellular calcium and was inhibited
by pertussis toxin and an antibody against the βγ dimer of a Gi protein. Cooperation with integrins and glycosylphosphatidylinositol-anchored proteins was necessary, since anti-integrin
antibodies and pretreatment with phosphatidylinositol-phospholipase C reduced exocytosis. Depletion of membrane cholesterol
inhibited exocytosis and decreased CD47 in lipid rafts, consistent with a CD47/integrin/Gi protein complex being located in rafts. An anti-CD47 antibody inhibited exocytosis induced by 4N1K and by mastoparan and
spermine, suggesting that basic secretagogues might target CD47. We propose that 4N1K-stimulated mast cell exocytosis involves
a CD47/integrin/Gi protein complex.
Received 8 December 2008; received after revision 12 January 2009; accepted 29 January 2009 相似文献
19.
Liora Lindenboim Elisa Ferrando-May Christoph Borner Reuven Stein 《Cellular and molecular life sciences : CMLS》2013,70(16):3013-3027
Bax and Bak (Bax/Bak) are essential pro-apoptotic proteins of the Bcl-2 family that trigger mitochondrial outer membrane permeabilization (MOMP) in a Bcl-2/Bcl-xL-inhibitable manner. We recently discovered a new stress-related function for Bax/Bak—regulation of nuclear protein redistribution (NPR) from the nucleus to cytoplasm. This effect was independent of Bax/Bak N-terminus exposure and not inhibited by Bcl-xL over-expression. Here, we studied the molecular mechanism governing this novel non-canonical response. Wild-type (WT) and mutant versions of Bax were re-expressed in Bax/Bak double-knockout mouse embryonic fibroblasts and their ability to promote NPR, apoptotic events, and changes in lamin A mobility was examined. Our results show that, in this system, Bax expression was sufficient to restore NPR such as in WT cells undergoing apoptosis. This activity of Bax was uncoupled from cytochrome c release from the mitochondria (indicative of MOMP) and required its membrane localization, α helices 5/6, and the Bcl-2 homology 3 (BH3) domain. Moreover, enrichment of Bax in the nuclear envelope by the so-called Klarsicht/ANC-1/Syne-1 homology domain effectively triggered NPR as in WT Bax, but without inducing MOMP or cell death. Bax-induced NPR was associated with impairment in lamin A mobility, implying a connection between these two nuclear envelope-associated events. Overall, the results indicate a new MOMP-independent, stress-induced Bax function on the nuclear envelope. 相似文献
20.
C. M. Vázquez R. Coleto R. Zanetti V. Ruiz-Gutierrez 《Cellular and molecular life sciences : CMLS》1997,53(5):442-446
In the present study, we have examined the intestinal Na+ transport, through the Na+-H+ exchanger, in ileal brush-border membrane vesicles (BBMV) isolated from spontaneously hypertensive rats (SHR), and normotensive
Wistar Kyoto (WKY) rats as a control group. Na+ uptake into ileal BBMV was stimulated in the presence of a proton gradient (pH 5.5 inside/pH 7.5 outside) in SHR and WKY
rats, resulting in a transient accumulation (overshoot) in both groups of rats. No overshoot was observed in the absence of
a pH gradient. The magnitude of the accumulation was significantly higher in SHR than in WKY rats. Uptake of Na+ at equilibrium was identical in the presence and the absence of a proton gradient and was not changed in SHR. The use of
amiloride inhibited pH gradient-driven Na+ uptake in a dose-dependent manner with a Ki of 90 μM and 100 μM for SHR and WKY rats, respectively. The relationship between
proton gradient-driven Na+ uptake and external Na+ concentration was saturable and conformed to Michaelis-Menten kinetics in both SHR and WKY rats. Lineweaver-Burk analysis
of the pH gradient-driven Na+ uptake indicated values of Vmax that were significantly increased in SHR compared to WKY rats (11.4±0.55 nmol/mg/8 s vs. 4.96±0.78 nmol/mg/8 s for SHR and
WKY rats, respectively). In contrast, similar Km values for Na+ were found between SHR and WKY rats (4.0±0.2 mM vs. 4.9±0.6 mM for SHR and WKY rats, respectively). These studies show derangement
in ileal BBMV Na+ transport of SHR, which is characterized by increased Na+-H+ exchanger activity.
Received 18 December 1996; received after revision 3 February 1997; accepted 7 February 1997 相似文献