Changes in Na+/K+-ATPase expression during adaptive cell differentiation in avian nasal salt gland

1997 ◽  
Vol 200 (13) ◽  
pp. 1895-1904 ◽  
Author(s):  
JP Hildebrandt
Keyword(s):  
Salt Stress ◽  
Cyclic Amp ◽  
Atpase Activity ◽  
Time Course ◽  
Atp Hydrolysis ◽  
Salt Gland ◽  
Secretory Cells ◽  
Salt Glands ◽  
Atpase Subunit

Chronic salt stress in ducklings (Anas platyrhynchos) resulted in a sustained accumulation of cyclic AMP in the secretory cells of the nasal salt glands. Adaptive increases in the activity of the Na+/K+-ATPase, measured as ATP hydrolysis rates in freshly isolated tissue, were observed after 12 h of salt stress. This change in enzyme activity was associated with increases in protein abundance in the - as well as in the ss-subunit of Na+/K+-ATPase and an increase in ss-subunit glycosylation. We investigated whether the increase in the cytosolic cyclic AMP concentration and the adaptive changes in Na+/K+-ATPase activity were causally related. Using an organotypic tissue culture system for salt gland slices from unstressed (naive) ducklings, we produced similar changes in Na+/K+-ATPase activity and subunit abundance by treating cultured tissue with drugs that elevate cytosolic cyclic AMP levels (forskolin, 8-CPT-cAMP) during a 15 h culture period. Protein synthesis assays using cultured tissue revealed that elevations in cytosolic cyclic AMP level mediate increases in Na+/K+-ATPase subunit abundance by slowing down the degradation of ATPase subunits. This increase in the amount of enzyme protein was associated with a significant increase in Na+/K+-ATPase activity in tissue homogenates. The time course of these changes in cyclic-AMP-treated cultured tissue resembled that observed in salt-stressed intact animals, indicating that the elevation in cyclic AMP level in salt gland tissue may constitute a portion of the signalling events ultimately leading to the adaptive increase in Na+/K+-ATPase activity in vivo.

Potassium secretion by nasal salt glands of desert lizard Sauromalus obesus

1987 ◽  
Vol 253 (1) ◽  
pp. R83-R90 ◽  
Author(s):  
T. J. Shuttleworth ◽  
J. L. Thompson ◽  
W. H. Dantzler
Keyword(s):  
Atpase Activity ◽  
Intraperitoneal Injection ◽  
Salt Gland ◽  
Salt Glands ◽  
Fourfold Increase ◽  
Gland Weight ◽  
Desert Lizard ◽  
Potassium Secretion ◽  
Sauromalus Obesus

Potassium secretion by the nasal salt glands of the herbivorous desert lizard Sauromalus obesus was determined in vivo by a new technique. Intraperitoneal injection of KCl rapidly increased the potassium secretion rate from 0.28 to 15.35 mumol X 100 g-1 X h-1. A second identical intraperitoneal injection, given 15 h after the first, further increased potassium secretion to 50.09 mumol X 100 g-1 X h-1. This was associated with a doubling of plasma K+ concentration and salt gland Na+-K+-adenosinetriphosphatase (ATPase) activity. Neither salt gland weight or residual (Mg2+) ATPase activity were affected. In an isolated perfused head preparation, potassium secretion from the nasal salt glands was stimulated from 0.99 to 10.76 mumol X 100 g-1 X h-1 by methacholine and to 14.68 mumol X 100 g-1 X h-1 by forskolin. In this perfused preparation, simultaneous determination of salt gland perfusion flow (using radiolabeled microspheres) and the rate of potassium secretion revealed that the secreting glands removed 68% of the perfusing potassium ions. Calculations indicated that secretion at the maximal rate observed in vivo would necessitate a fourfold increase in the rate of blood flow to the gland.

Salt stress increases abundance and glycosylation of CFTR localized at apical surfaces of salt gland secretory cells

1994 ◽  
Vol 267 (4) ◽  
pp. C990-C1001 ◽  
Author(s):  
S. A. Ernst ◽  
K. M. Crawford ◽  
M. A. Post ◽  
J. A. Cohn
Keyword(s):  
Salt Stress ◽  
Single Cells ◽  
Salt Gland ◽  
Cell Protein ◽  
Secretory Cells ◽  
Salt Glands ◽  
Cl Secretion ◽  
Peptide Antibody ◽  
Nacl Secretion ◽  
Apical Membranes

Osmotic stress elicits hypertonic NaCl secretion and promotes structural and biochemical differentiation in avian salt glands. In addition to cholinergic control, Cl- secretion is stimulated by vasoactive intestinal peptide (VIP), suggesting that the cystic fibrosis transmembrane conductance regulator (CFTR) may be present and that its expression may be regulated by chronic salt stress. Anion efflux, assayed by 6-methoxy-N-(3-sulfopropyl)quinolinium fluorescence changes in single cells, was stimulated by VIP or 8-(4-chlorophenylthio)adenosine 3',5'-cyclic monophosphate. Immunoblots with a COOH-terminal peptide antibody to human CFTR revealed approximately 170- and approximately 180-kDa bands in lysates from control and salt-stressed glands, respectively. Both variants reduced to approximately 140 kDa after N-glycanase digestion and gave identical tryptic phosphopeptide maps after immunoprecipitation and phosphorylation by protein kinase A. CFTR was localized to apical membranes by immunofluorescence and, additionally, to subapical vesicles by immunoelectron microscopy. Salt stress induced an approximately twofold increase in CFTR abundance/cell protein (approximately 5-fold/cell) and intensified apical membrane immunofluorescence. For comparison, Na+ pump expression increased approximately fourfold per cell protein with little change in actin. Thus differentiation induced by salt stress is accompanied by alteration in CFTR abundance and glycosylation. Upregulation of CFTR likely contributes to increased efficiency of Cl- secretion.

scholarly journals PRESERVATION OF Na-K-ACTIVATED AND Mg-ACTIVATED ADENOSINE TRIPHOSPHATASE ACTIVITIES OF AVIAN SALT GLAND AND TELEOST GILL WITH FORMALDEHYDE AS FIXATIVE

1970 ◽  
Vol 18 (4) ◽  
pp. 251-263 ◽  
Author(s):  
STEPHEN A. ERNST ◽  
CHARLES W. PHILPOTT
Keyword(s):  
Atpase Activity ◽  
Adenosine Triphosphatase ◽  
Electron Microscopic Examination ◽  
Salt Gland ◽  
Chloride Cells ◽  
Secretory Cells ◽  
Salt Glands ◽  
Electron Microscopic ◽  
Fixed Tissue ◽  
Gill Filaments

The effect of glutaraldehyde and formaldehyde fixation on the level of biochemically demonstrable Na-K-adenosine triphosphatase (Na-K-ATPase) and Mg-ATPase of avian salt glands and teleost gill filaments was studied. Sections, 100-200 µ, prepared with the Smith-Farquhar tissue chopper, were fixed for varying periods, homogenized and assayed for ATPase activity. Fixation of salt gland tissue with 0.5% glutaraldehyde for 40-60 min completely inhibited the Na-K-ATPase activity and reduced the level of Mg-ATPase by 85%. In contrast, fixation with 2 or 3% formaldehyde, prepared from paraformaldehyde, for 60-90 min resulted in a loss of only 30% of the Na-K-ATPase activity and 65% of the Mg-ATPase activity. Similar results were obtained with gill filaments. In addition, Na-K-ATPase of formaldehyde-fixed tissue retained an obligatory requirement for Na+ and K+ and was fully sensitive to ouabain. Electron microscopic examination of formaldehyde-fixed tissue, sectioned with either the tissue chopper or in the cryostat and incubated in the Wachstein-Meisel medium, showed excellent morphologic preservation. Reaction product deposition (presumably due to Mg-ATPase) was associated with the extracellular side of the plasma membrane in the secretory cells of the salt gland and over the mitochondrial matrix of chloride cells present in the gill epithelium.

Morphology of isolated crustacean larval salt glands

1985 ◽  
Vol 248 (6) ◽  
pp. R709-R716
Author(s):  
R. J. Lowy ◽  
F. P. Conte
Keyword(s):  
Plasma Membrane ◽  
Salt Gland ◽  
Artemia Salina ◽  
Apical Plasma Membrane ◽  
Secretory Cells ◽  
Cell Contact ◽  
Salt Glands ◽  
Transmission Electron

Larval salt glands isolated from the naupliar brine shrimp (Artemia salina) were examined using light microscopy and scanning and transmission electron microscopy. These methods demonstrated that most cellular and subcellular features of the in vitro organ compared favorably with those seen in vivo. This salt gland measures 130 micron in diameter and is comprised of 50-70 secretory cells, which are of a single epithelial cell type. Characteristic ultrastructural features that are well preserved include apical to basal cell polarity, apical plasma membrane projections, and the extent of the basolateral tubular labyrinth and its association with numerous mitochondria. Some features that have been altered are a decrease in cell-cell contact, separation of septate junctions, and expansion of tubular labyrinth lumens and mitochondrial cristae. Use of this preparation has allowed examination of the salt gland cell's hemocoelic surface for the first time and provided information about the ultrastructure of the tufts formed by the apical plasma membrane.

Biogenesis of Plasma Membranes in Salt Glands of Salt-Stressed Domestic Ducklings: Localization of Acyltransferase Activity

1972 ◽  
Vol 11 (3) ◽  
pp. 855-873
Author(s):  
A. M. LEVINE ◽  
JOAN A. HIGGINS ◽  
R. J. BARRNETT
Keyword(s):  
Plasma Membrane ◽  
Plasma Membranes ◽  
Specific Activity ◽  
Salt Gland ◽  
Secretory Cells ◽  
Salt Glands ◽  
Acyltransferase Activity ◽  
Structural Localization ◽  
Differentiated Cells ◽  
Golgi Membranes

In response to salt water stress there is a marked increase in the plasma membranes of the epithelial secretory cells of the salt glands of domestic ducklings. In the present study, the fine-structural localization of the acyltransferases involved in synthesis of phospholipids has been investigated in this tissue during this increased biogenesis of plasma membranes. The specific activity of the acyltransferases of the salt gland rose in response to salt stress, and this preceded the rapid increase in weight and cellular differentiation. After the weight increase of the gland became established, the specific activity of the acyltransferases declined, but the total activity remained constant. Salt gland tissue fixed in a mixture of glutaraldehyde and formaldehyde retained 35% of the acyltransferase activity of unfixed tissue. Cytochemical studies of the localization of acyltransferase activity in fixed and unfixed salt gland showed reaction product associated only with the lamellar membranes of the Golgi complex. This localization occurred in partially differentiated cells from salt-stressed glands to the greatest extent; and to only a small extent in cells of control tissue from unstressed salt glands. Omission of substrates resulted in absence of reaction product in association with the Golgi membranes. In addition, vesicles having limiting membranes morphologically similar to the plasma membrane occurred between the Golgi region and the plasma membrane in the partially differentiated cells. The phospholipid component of the plasma membrane appears therefore to be synthesized in association with the Golgi membranes and the membrane packaged at this site from which it moves in the form of vesicles to fuse with the pre-existing plasma membrane.

Methyl jasmonate improves tolerance to high salt stress in the recretohalophyte Limonium bicolor

2019 ◽  
Vol 46 (1) ◽  
pp. 82 ◽  
Author(s):  
Fang Yuan ◽  
Xue Liang ◽  
Ying Li ◽  
Shanshan Yin ◽  
Baoshan Wang
Keyword(s):  
Salt Stress ◽  
Salt Tolerance ◽  
Methyl Jasmonate ◽  
Salt Gland ◽  
Underlying Mechanism ◽  
Photosynthetic Parameters ◽  
Salt Glands ◽  
Plasma Membrane Permeability ◽  
Limonium Bicolor ◽  
Relative Electrical Conductivity

Limonium bicolor is a typical recretohalophyte with salt glands in the epidermis, which shows maximal growth at moderate salt concentrations (100mM NaCl) but reduced growth in the presence of excess salt (more than 200mM). Jasmonic acid (JA) alleviates the reduced growth of L. bicolor under salt stress; however, the underlying mechanism is unknown. In this study we investigated the effects of exogenous methyl jasmonate (MeJA) application on L. bicolor growth at high NaCl concentrations. We found that treatment with 300mM NaCl led to dramatic inhibition of seedling growth that was significantly alleviated by the application of 0.03mM MeJA, resulting in a biomass close to that of plants not subjected to salt stress. To determine the parameters that correlate with MeJA-induced salt tolerance (assessed as the biomass production in saline and control conditions), we measured 14 physiological parameters relating to ion contents, plasma membrane permeability, photosynthetic parameters, salt gland density, and salt secretion. We identified a correlation between individual indicators and salt tolerance: the most positively correlated indicator was net photosynthetic rate, and the most negatively correlated one was relative electrical conductivity. These findings provide insights into a possible mechanism underlying MeJA-mediated salt stress alleviation.

scholarly journals Measurement of the dynamics of stimulation and inhibition of steroidogenesis in isolated rat adrenal cells by using column perfusion

1974 ◽  
Vol 142 (2) ◽  
pp. 287-294 ◽  
Author(s):  
P. J. Lowry ◽  
Colin McMartin
Keyword(s):  
Cyclic Amp ◽  
Time Course ◽  
Acth Stimulation ◽  
Duration Of Action ◽  
Adrenal Cells ◽  
Small Column ◽  
Long Duration ◽  
Full Effect ◽  
Rapid Fall

Isolated adrenal cells were perfused in a small column by using Bio-Gel polyacrylamide beads as an inert supporting matrix, and the time-course of the response to various stimuli was observed by measuring fluorogenic 11-hydroxycorticosteroids in the effluent. A small but significant response was observed 1 min after stimulation with physiological concentrations of ACTH (adrenocorticotrophin), but the response did not start to build up rapidly for 3–4min and eventually reached a plateau after 9–10min. A similar pattern of events was observed for the decay of the steroid output on removal of ACTH. ACTH analogues, including one with a long duration of action in vivo, were found to produce responses with similar kinetics. However, cyclic AMP caused a more rapid increase in steroidogenesis and its effects were more short-lived after withdrawal. If, as present evidence suggests, cyclic AMP is produced rapidly after ACTH stimulation the delayed build-up of the steroidogenic response to ACTH would indicate that cyclic AMP may not be the intracellular mediator. When inhibitors were applied during ACTH stimulation, aminoglutethimide, which blocks mitochondrial conversion of cholesterol into pregnenolone (3β-hydroxypregn-5-en-20-one), caused a rapid fall in steroid output (1 min), whereas cycloheximide took longer to achieve its full effect. Nevertheless, the response had fallen by 50% in 2 min, indicating a much shorter half-life than that previously reported for the labile protein implicated in steroidogenesis. In addition the rapid response to cyclic AMP makes it unlikely that steroid production is induced as a result of initiation of protein synthesis. This suggests that the labile protein plays an obligatory but permissive role in the development of the response. Column perfusion has proved to be a simple technique which can readily yield accurate data on responses of cells to stimulants and inhibitors.

Possible role of cyclic GMP in stimulus-secretion coupling by salt gland of the duck

1979 ◽  
Vol 237 (5) ◽  
pp. C200-C204 ◽  
Author(s):  
D. J. Stewart ◽  
J. Sax ◽  
R. Funk ◽  
A. K. Sen
Keyword(s):  
Cyclic Amp ◽  
Guanylate Cyclase ◽  
Cyclic Gmp ◽  
Salt Gland ◽  
Domestic Ducks ◽  
Stimulus Secretion Coupling ◽  
Stimulation Of

Stimulation of salt galnd secretion in domestic ducks in vivo increased the cyclic GMP concentration of the tissue, but had no effect on cyclic AMP levels. Methacholine, which is known to stimulate sodium transport by the glands both in vivo and in vitro, stimulated ouabain-sensitive respiration in salt gland slices. Cyclic GMP stimulated ouabain-sensitive respiration to the same extent as methacholine. Guanylate cyclase stimulators, hydroxylamine and sodium azide, also stimulated ouabain-sensitive respiration. The stimulation of ouabain-sensitive respiration by methacholine was blocked either by atropine or by removal of calcium from the incubation medium. The stimulation of ouabain-sensitive respiration by cyclic GMP still occurred in the absence of calcium. The above observations seem to indicate that cyclic GMP acts as a tertiary link in the process of stimulus-secretion coupling in the tissue.

scholarly journals The Walker B motif of the second nucleotide-binding domain (NBD2) of CFTR plays a key role in ATPase activity by the NBD1–NBD2 heterodimer

2006 ◽  
Vol 401 (2) ◽  
pp. 581-586 ◽  
Author(s):  
Fiona L. L. Stratford ◽  
Mohabir Ramjeesingh ◽  
Joanne C. Cheung ◽  
Ling-JUN Huan ◽  
Christine E. Bear
Keyword(s):  
Atpase Activity ◽  
Atp Hydrolysis ◽  
Nucleotide Binding ◽  
Vital Role ◽  
Atp Binding ◽  
Primary Role ◽  
Binding Domains ◽  
Membrane Spanning ◽  
Atp Binding And Hydrolysis

CFTR (cystic fibrosis transmembrane conductance regulator), a member of the ABC (ATP-binding cassette) superfamily of membrane proteins, possesses two NBDs (nucleotide-binding domains) in addition to two MSDs (membrane spanning domains) and the regulatory ‘R’ domain. The two NBDs of CFTR have been modelled as a heterodimer, stabilized by ATP binding at two sites in the NBD interface. It has been suggested that ATP hydrolysis occurs at only one of these sites as the putative catalytic base is only conserved in NBD2 of CFTR (Glu1371), but not in NBD1 where the corresponding residue is a serine, Ser573. Previously, we showed that fragments of CFTR corresponding to NBD1 and NBD2 can be purified and co-reconstituted to form a heterodimer capable of ATPase activity. In the present study, we show that the two NBD fragments form a complex in vivo, supporting the utility of this model system to evaluate the role of Glu1371 in ATP binding and hydrolysis. The present studies revealed that a mutant NBD2 (E1371Q) retains wild-type nucleotide binding affinity of NBD2. On the other hand, this substitution abolished the ATPase activity formed by the co-purified complex. Interestingly, introduction of a glutamate residue in place of the non-conserved Ser573 in NBD1 did not confer additional ATPase activity by the heterodimer, implicating a vital role for multiple residues in formation of the catalytic site. These findings provide the first biochemical evidence suggesting that the Walker B residue: Glu1371, plays a primary role in the ATPase activity conferred by the NBD1–NBD2 heterodimer.

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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