Induction of gene expression by heat shock versus osmotic stress

1994 ◽  
Vol 267 (1) ◽  
pp. F28-F34 ◽  
Author(s):  
D. Sheikh-Hamad ◽  
A. Garcia-Perez ◽  
J. D. Ferraris ◽  
E. M. Peters ◽  
M. B. Burg
Keyword(s):  
High Temperature ◽  
Elevated Temperature ◽  
Heat Shock ◽  
Mdck Cells ◽  
Organic Osmolytes ◽  
Hsp 70 ◽  
Expression Of Genes ◽  
High Concentrations ◽  
Shock Proteins ◽  
Darby Canine Kidney

Elevated temperature rapidly increases expression of genes for heat shock proteins (HSP), including HSP-70. The response is presumably triggered by denaturation of cell proteins and helps in their renaturation. Hypertonicity may also denature proteins, but the protective response, which is accumulation of compatible organic osmolytes [including betaine and inositol in Madin-Darby canine kidney (MDCK) cells], apparently differs and is slow. Recently, hypertonicity was found also to increase expression of HSP-70 in MDCK cells, a response proposed to provide protection until organic osmolytes can accumulate. Our purpose was to examine whether 1) a gene involved in accumulation of organic osmolytes also responds to heat stress and 2) whether accumulation of organic osmolytes affects expression of HSP-70. We find that 1) the betaine transporter mRNA, which is greatly increased by hypertonicity (515 vs. 315 mosmol), is unaffected by high temperature (42 degrees C vs. 37 degrees C); 2) hypertonicity-induced increases in HSP-70 and betaine transporter mRNA are much greater when the medium (and cell) contain no betaine and no inositol than when high concentrations of these are present; and 3) high betaine greatly inhibits the increase in HSP-70 mRNA at high temperature. We conclude the following. 1) Although heat shock and betaine transporter genes both respond to hypertonicity, the betaine transporter is not a HSP. 2) Accumulation of organic osmolytes attenuates the HSP-70 response to hypertonicity, as it might if the HSP-70 expression were a temporizing response. 3) Betaine inhibits HSP-70 response to elevated temperature, presumably by its known effect of stabilizing proteins.

Osmoregulation of glycerophosphorylcholine content of mammalian renal cells

1989 ◽  
Vol 257 (4) ◽  
pp. C795-C801 ◽  
Author(s):  
T. Nakanishi ◽  
M. B. Burg
Keyword(s):  
Mdck Cells ◽  
Urea Concentration ◽  
Organic Osmolytes ◽  
Renal Cells ◽  
Madin Darby Canine Kidney ◽  
Cells In Culture ◽  
Nacl Concentration ◽  
High Concentrations ◽  
Darby Canine Kidney ◽  
Canine Kidney

Renal medullary cells contain high concentrations of "compatible" organic osmolytes such as glycerophosphorylcholine (GPC), betaine, myo-inositol, and sorbitol. The organic osmolytes occur as an osmoregulatory response to the high and variable interstitial NaCl concentration that is part of the urinary concentrating mechanism. Madin-Darby canine kidney (MDCK) cells in culture were previously shown to accumulate GPC in response to increased osmolality. We demonstrate here that this accumulation occurs in response to elevated extracellular urea concentration as well as to elevated NaCl. GPC does not accumulate unless either choline or GPC is present in the medium. Thus the accumulation results from osmoregulated synthesis of GPC from choline and, possibly, also osmoregulated uptake of extracellular GPC. When the osmolality is decreased from high to normal levels, cell GPC concentration decreases greatly over 24 h, accompanied by efflux of GPC and choline into the medium.

Osmoregulatory fluxes of myo-inositol and betaine in renal cells

1989 ◽  
Vol 257 (5) ◽  
pp. C964-C970 ◽  
Author(s):  
T. Nakanishi ◽  
M. B. Burg
Keyword(s):  
Mdck Cells ◽  
Initial Response ◽  
Organic Osmolytes ◽  
Madin Darby Canine Kidney ◽  
Initial Value ◽  
Nacl Concentration ◽  
Sodium Dependent ◽  
High Concentrations ◽  
Darby Canine Kidney ◽  
Canine Kidney

Renal medullary cells contain high concentrations of "compatible" organic osmolytes, such as myo-inositol, betaine, sorbitol, and glycero-phosphorylcholine. These organic osmolytes accumulate as an osmoregulatory response to the high and variable interstitial NaCl concentration that is part of the urinary concentrating mechanism. Madin-Darby canine kidney (MDCK) cells in culture were previously shown to accumulate myo-inositol and betaine in response to increased NaCl. These organic osmolytes are taken up by sodium-dependent active transport into the cells from the medium. The maximum concentration is not reached until 2-4 days after an increase in medium osmolality. The purpose of this study was to characterize the response to a decrease in medium osmolality of cells that had been grown at a high osmolality. The initial response to decreased osmolality was a rapid, transient efflux of both myo-inositol and betaine from the cells. Efflux was greatest during the first 15 min and resulted in a reduction of cell myo-inositol and betaine by almost 13 and 22%, respectively, after 3 h. Active myo-inositol and betaine influx fell more slowly, reaching a lower limit after approximately 1-2 days. The reduced influx was followed by progressive decrease in cell myo-inositol and betaine to approximately 30% of the initial value after 6 days. Thus, after a decrease in medium osmolality, MDCK cell myo-inositol and betaine fell because of a rapid, transient increase in efflux and a slow, sustained decrease in active influx.

Accumulation of natural and synthetic betaines by a mammalian renal cell line

1996 ◽  
Vol 74 (2) ◽  
pp. 283-287 ◽  
Author(s):  
K. Randall ◽  
M. Lever ◽  
B. A. Peddie ◽  
S. T. Chambers
Keyword(s):  
Osmotic Stress ◽  
Glycine Betaine ◽  
Mdck Cells ◽  
Intracellular Accumulation ◽  
Madin Darby Canine Kidney ◽  
Renal Cell Line ◽  
High Concentrations ◽  
Inhibition Studies ◽  
Darby Canine Kidney ◽  
Canine Kidney

Intracellular accumulation of different betaines was compared in osmotically stressed Madin Darby canine kidney (MDCK) cells to model the betaine accumulation specificity of the mammalian inner medulla and to show how this accumulation differed from that of bacteria. All betaines accumulated less than glycine betaine. Arsenobetaine (the arsenic analogue of glycine betaine) accumulated to 12% of the glycine betaine levels and the sulphur analogue dimethylthetin accumulated to >80%. Most substituted glycine betaine analogues accumulated to 2–5% of intracellular glycine betaine concentrations, however, serine betaine accumulated to <0.5% of glycine betaine levels. Inhibition studies to distinguish the betaine ports were performed by the addition of proline. Butyrobetaine and carnitine accumulation was not proline sensitive, whereas that of omer betaines was. As with glycine betaine, the accumulation of propionobetaine and dimethylthetin was proline sensitive and osmoregulated. Pyridinium betaine was accumulated by both proline-sensitive and -insensitive systems, with a small increase under osmotic stress. High concentrations (10 times that of glycine betaine) of the dietary betaines proline betaine and trigonelline inhibited total betaine accumulation. Because α-substituted betaines are accumulated by bacteria and not by MDCK cells, these betaines may be the basis for design of antimicrobial agents.Key words: MDCK cells, betaine accumulation, osmolytes, betaine analogues.

scholarly journals Regulated Overexpression of Heat Shock Protein 72 Protects Madin-Darby Canine Kidney Cells from the Detrimental Effects of High Urea Concentrations

2001 ◽  
Vol 12 (12) ◽  
pp. 2565-2571 ◽  
Author(s):  
Wolfgang Neuhofer ◽  
Karin Lugmayr ◽  
Maria-Luisa Fraek ◽  
Franz-X Beck
Keyword(s):  
Heat Shock ◽  
Heat Shock Protein ◽  
Mdck Cells ◽  
Heat Shock Protein 72 ◽  
Madin Darby Canine Kidney ◽  
Lactate Dehydrogenase Activity ◽  
Hsp72 Expression ◽  
Darby Canine Kidney ◽  
Canine Kidney ◽  
Medullary Cells

ABSTRACT. Exposure of renal medullary cells to elevated extracellular NaCl concentrations is associated with increased heat shock protein 72 (HSP72) expression and improved resistance to subsequent exposure to a high urea concentration (600 mM). To establish a causal relationship between HSP72 expression and protection against high urea concentrations, HSP72 was inducibly overexpressed in Madin-Darby canine kidney (MDCK) cells, in the absence of hypertonic stress before urea exposure. For this purpose, the human stress-inducible HSP72 gene was cloned downstream from a dexamethasone (DEX)-inducible promoter in the eukaryotic expression vector pLKneo. This construct allowed robust induction of HSP72 by exposure of stably transfected MDCK cells (MDCK-LK72) to 0.1 μM DEX. Increased HSP72 abundance significantly improved survival rates after 24-h exposure of the cells to medium containing 600 mM urea (14 versus 43%). In mock-transfected or wild-type cells, DEX had no significant effect on HSP72 abundance or urea resistance. In accordance with those findings, lactate dehydrogenase activity in the supernatant was significantly reduced, compared with appropriate control samples, only in MDCK-LK72 cells overexpressing HSP72. Labeling with annexin V-FITC and propidium iodide, followed by flow cytometry, revealed that overexpression of HSP72 was associated with a reduction in the number of apoptotic-lysed cells, a concomitant retardation of apoptosis, and an increase in the number of viable cells. These data support the view that HSP72, which is very abundant in the renal inner medulla, is an important component of the defense mechanism of medullary cells against extreme concentrations of urea.

Morphological and molecular modifications induced by heat shock in Drosophila melanogaster embryos

Development ◽  
1983 ◽  
Vol 77 (1) ◽  
pp. 167-182
Author(s):  
Giorgio Graziosi ◽  
Franco de Cristini ◽  
Angelo di Marcotullio ◽  
Roberto Marzari ◽  
Fulvio Micali ◽  
...  
Keyword(s):  
Drosophila Melanogaster ◽  
Heat Shock ◽  
Dna Synthesis ◽  
Histological Analysis ◽  
Developmental Stages ◽  
Early Embryo ◽  
Direct Relation ◽  
Hsp 70 ◽  
Survival Pattern ◽  
Shock Proteins

The early embryo of Drosophila melanogaster did not survive treatment at 37 °C (heat shock) for 25 min. The histological analysis of eggs treated in this way showed that the heat shock caused disintegration of nuclei and of cytoplasmic islands, displacement and swelling of nuclei and blocked mitoses. These effects were not observed in embryos treatedafter blastoderm formation. After this stage, we noticed that development was slowed down. The heat shock proteins (hsp 83,70 and 68) were, under shock, synthesized at all developmental stages. There was little or no synthesis of hsp 70 and 68 in unfertilized eggs, but synthesis increased in proportion to the number of nuclei present. Most probably, hsp 70 synthesis was directed by zygotic mRNA. DNA synthesis was not blocked by the heat shock though the overall incorporation of [3H]thymidine was substantially reduced, presumably because of the block of mitoses. We did not find a direct relation between survival pattern and hsp synthesis. We concluded that some, at least, of the heat shock genes can be activated at all developmental stages and that heat shock could be used for synchronizing mitoses.

scholarly journals Species-specific collaboration of heat shock proteins (Hsp) 70 and 100 in thermotolerance and protein disaggregation

2011 ◽  
Vol 108 (17) ◽  
pp. 6915-6920 ◽  
Author(s):  
M. Miot ◽  
M. Reidy ◽  
S. M. Doyle ◽  
J. R. Hoskins ◽  
D. M. Johnston ◽  
...  
Keyword(s):  
Heat Shock ◽  
Heat Shock Proteins ◽  
Hsp 70 ◽  
Species Specific ◽  
Shock Proteins

The Acquisition and Maintenance of Thermotolerance in Australian Wheats

1990 ◽  
Vol 17 (1) ◽  
pp. 37 ◽  
Author(s):  
C Blumenthal ◽  
F Bekes ◽  
CW Wrigley ◽  
EWR Barlow
Keyword(s):  
Heat Treatment ◽  
Triticum Aestivum ◽  
Protein Synthesis ◽  
High Temperature ◽  
Heat Shock ◽  
High Temperature Stress ◽  
Time Interval ◽  
Degree Of Protection ◽  
Concomitant Reduction ◽  
Shock Proteins

The exposure of wheat (Triticum aestivum) coleoptiles to a transient high temperature stress results in the synthesis of a group of proteins known as the heat shock proteins (hsps). The appearance of these proteins is associated with a concomitant reduction in normal protein synthesis and has been correlated with the acquisition of thermotolerance (assessed as growth of coleoptiles). Pretreatment with a sublethal heat shock confers protection to a subsequent heat shock that would otherwise have been lethal. In addition, we find that increasing the time interval between the sublethal heat treatment and the subsequent heat shock from 0 to 72 h reduces the protective effect of the sublethal heat treatment considerably. The five cultivars examined (Sunelg, Sunco, Hartog, Vulcan, Halberd) showed differences in the degree of protection acquired, and in the length of time for which protection was maintained. Hartog was found to be the most thermotolerant, and acquired the greatest degree of protection from exposure to a sublethal heat treatment, but the duration of this acquired protection was shorter than in the remaining cultivars. Sunelg was most susceptible to a heat shock but the duration of acquired protection was the greatest.

Osmoregulation of GPC:choline phosphodiesterase in MDCK cells: different effects of urea and NaCl

1995 ◽  
Vol 269 (1) ◽  
pp. C35-C41 ◽  
Author(s):  
E. D. Kwon ◽  
K. Zablocki ◽  
K. Y. Jung ◽  
E. M. Peters ◽  
A. Garcia-Perez ◽  
...  
Keyword(s):  
Enzymatic Degradation ◽  
Cell Function ◽  
Specific Activity ◽  
Mdck Cells ◽  
Absolute Rate ◽  
Madin Darby Canine Kidney ◽  
Organic Osmolyte ◽  
The Absolute ◽  
High Concentrations ◽  
Darby Canine Kidney

The organic osmolyte, glycerophosphocholine (GPC), accumulates in renal cells in response to high concentrations of either NaCl or urea, despite the very different effects of these solutes on cell function and volume. Together, high levels of these solutes increase GPC amount in Madin-Darby canine kidney cells by inhibiting its enzymatic degradation. The present study tests the effects of NaCl and urea, individually, on GPC accumulation and its degradation. A technique was developed to determine the absolute rate of GPC degradation by measuring the initial rate of disappearance of [3H]GPC (pulsed into the cells by hypotonic shock) and the specific activity of GPC in the cells. The mass of GPC in the cells was measured by another newly developed method, a sensitive chemiluminescent assay. We find that exposure to high NaCl or urea decreases the absolute rate of cellular GPC degradation by approximately one-half during the first 20.5 h. Reductions in GPC degradation are accompanied by commensurate decreases in the activity of GPC:choline phosphodiesterase (GPC:PDE; EC 3.1.4.2), an enzyme that catalyzes degradation of GPC. Activity of GPC:PDE falls > 50% in cells exposed for 2 h to high osmolality. Inhibition is sustained for 7 days with high urea alone. In contrast, with high NaCl alone, GPC:PDE activity reverts to control values by 7 days, by which time synthesis of GPC is increased, accounting for sustained GPC accumulation. Collectively, these data suggest that GPC accumulation in response to either high NaCl or urea occurs initially by inhibition of its degradation but that the effect of NaCl on degradation differs, in that it is transient, while that of urea is sustained.

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