scholarly journals Effects of cycloheximide on thermotolerance expression, heat shock protein synthesis, and heat shock protein mRNA accumulation in rat fibroblasts.

1986 ◽  
Vol 6 (4) ◽  
pp. 1088-1094 ◽  
Author(s):  
R B Widelitz ◽  
B E Magun ◽  
E W Gerner
Keyword(s):  
Protein Synthesis ◽  
Heat Shock ◽  
Heat Shock Protein ◽  
Mrna Levels ◽  
Hsp 70 ◽  
Mrna Accumulation ◽  
General Protein Synthesis ◽  
Rat Fibroblasts ◽  
General Protein ◽  
Rat Embryonic Fibroblasts

A single hyperthermic exposure can render cells transiently resistant to subsequent high temperature stresses. Treatment of rat embryonic fibroblasts with cycloheximide for 6 h after a 20-min interval at 45 degrees C inhibits protein synthesis, including heat shock protein (hsp) synthesis, and results in an accumulation of hsp 70 mRNA, but has no effect on subsequent survival responses to 45 degrees C hyperthermia. hsp 70 mRNA levels decreased within 1 h after removal of cycloheximide but then appeared to stabilize during the next 2 h (3 h after drug removal and 9 h after heat shock). hsp 70 mRNA accumulation could be further increased by a second heat shock at 45 degrees C for 20 min 6 h after the first hyperthermic exposure in cycloheximide-treated cells. Both normal protein and hsp synthesis appeared increased during the 6-h interval after hyperthermia in cultures which received two exposures to 45 degrees C for 20 min compared with those which received only one treatment. No increased hsp synthesis was observed in cultures treated with cycloheximide, even though hsp 70 mRNA levels appeared elevated. These data indicate that, although heat shock induces the accumulation of hsp 70 mRNA in both normal and thermotolerant cells, neither general protein synthesis nor hsp synthesis is required during the interval between two hyperthermic stresses for Rat-1 cells to express either thermotolerance (survival resistance) or resistance to heat shock-induced inhibition of protein synthesis.

Effects of cycloheximide on thermotolerance expression, heat shock protein synthesis, and heat shock protein mRNA accumulation in rat fibroblasts

1986 ◽  
Vol 6 (4) ◽  
pp. 1088-1094
Author(s):  
R B Widelitz ◽  
B E Magun ◽  
E W Gerner
Keyword(s):  
Protein Synthesis ◽  
Heat Shock ◽  
Heat Shock Protein ◽  
Mrna Levels ◽  
Hsp 70 ◽  
Mrna Accumulation ◽  
General Protein Synthesis ◽  
Rat Fibroblasts ◽  
General Protein ◽  
Rat Embryonic Fibroblasts

A single hyperthermic exposure can render cells transiently resistant to subsequent high temperature stresses. Treatment of rat embryonic fibroblasts with cycloheximide for 6 h after a 20-min interval at 45 degrees C inhibits protein synthesis, including heat shock protein (hsp) synthesis, and results in an accumulation of hsp 70 mRNA, but has no effect on subsequent survival responses to 45 degrees C hyperthermia. hsp 70 mRNA levels decreased within 1 h after removal of cycloheximide but then appeared to stabilize during the next 2 h (3 h after drug removal and 9 h after heat shock). hsp 70 mRNA accumulation could be further increased by a second heat shock at 45 degrees C for 20 min 6 h after the first hyperthermic exposure in cycloheximide-treated cells. Both normal protein and hsp synthesis appeared increased during the 6-h interval after hyperthermia in cultures which received two exposures to 45 degrees C for 20 min compared with those which received only one treatment. No increased hsp synthesis was observed in cultures treated with cycloheximide, even though hsp 70 mRNA levels appeared elevated. These data indicate that, although heat shock induces the accumulation of hsp 70 mRNA in both normal and thermotolerant cells, neither general protein synthesis nor hsp synthesis is required during the interval between two hyperthermic stresses for Rat-1 cells to express either thermotolerance (survival resistance) or resistance to heat shock-induced inhibition of protein synthesis.

Dissociation of 68,000 M r Heat Shock Protein Synthesis from Thermotolerance Expression in Rat Fibroblasts

1984 ◽  
Vol 99 (2) ◽  
pp. 433 ◽  
Author(s):  
Randall B. Widelitz ◽  
Bruce E. Magun ◽  
Eugene W. Gerner
Keyword(s):  
Protein Synthesis ◽  
Heat Shock ◽  
Heat Shock Protein ◽  
Rat Fibroblasts

Heat shock gene expression in Xenopus laevis A6 cells in response to heat shock and sodium arsenite treatments

1988 ◽  
Vol 66 (8) ◽  
pp. 862-870 ◽  
Author(s):  
S. Darasch ◽  
D. D. Mosser ◽  
N. C. Bols ◽  
J. J. Heikkila
Keyword(s):  
Heat Shock ◽  
Xenopus Laevis ◽  
Sodium Arsenite ◽  
Polyacrylamide Gel Electrophoresis ◽  
Heat Shock Gene ◽  
Epithelial Cell Line ◽  
Continuous Exposure ◽  
Mrna Levels ◽  
Hsp 70 ◽  
Mrna Accumulation

Continuous exposure of a Xenopus laevis kidney epithelial cell line, A6, to either heat shock (33 °C) or sodium arsenite (50 μM) resulted in transient but markedly different temporal patterns of heat-shock protein (HSP) synthesis and HSP 70 and 30 mRNA accumulation. Heat-shock-induced synthesis of HSPs was detectable within 1 h and reached maximum levels by 2–3 h. While sodium arsenite induced the synthesis of some HSPs within 1 h, maximal HSP synthesis did not occur until 12 h. The pattern of HSP 70 and 30 mRNA accumulation was similar to the response observed at the protein level. During recovery from heat shock, a coordinate decline in HSPs and HSP 70 and 30 mRNA was observed. During recovery from sodium arsenite, a similar phenomenon occurred during the initial stages. However, after 6 h of recovery, HSP 70 mRNA levels persisted in contrast to the declining HSP 30 mRNA levels. Two-dimensional polyacrylamide gel electrophoresis revealed the presence of 5 HSPs in the HSP 70 family, of which two were constitutive, and 16 different stress-inducible proteins in the HSP 30 family. In conclusion, heat shock and sodium arsenite induce a similar set of HSPs but maximum synthesis of the HSP is temporally separated by 12–24 h.

scholarly journals Biochemical analysis of the stress protein response in human oesophageal epithelium

Gut ◽  
1997 ◽  
Vol 41 (2) ◽  
pp. 156-163 ◽  
Author(s):  
D Hopwood ◽  
S Moitra ◽  
B Vojtesek ◽  
D A Johnston ◽  
J F Dillon ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Heat Shock ◽  
Stress Responses ◽  
Biochemical Analysis ◽  
Ex Vivo ◽  
Stress Protein ◽  
Hsp70 Protein ◽  
Biopsy Specimens ◽  
General Protein Synthesis ◽  
General Protein

Background—The oesophageal epithelium is exposed routinely to noxious agents in the environment, including gastric acid, thermal stress, and chemical toxins. These epithelial cells have presumably evolved effective protective mechanisms to withstand tissue damage and repair injured cells. Heat shock protein or stress protein responses play a central role in protecting distinct cell types from different types of injury.Aim—To determine (i) whether biochemical analysis of stress protein responses in pinch biopsy specimens from human oesophageal epithelium is feasible; (ii) whether undue stresses are imposed on cells by the act of sample collection, thus precluding analysis of stress responses; and (iii) if amenable to experimentation, the type of heat shock protein (Hsp) response that operates in the human oesophageal epithelium.Methods—Tissue from the human oesophagus comprised predominantly of squamous epithelium was acquired within two hours of biopsy and subjected to an in vitro heat shock. Soluble tissue cell lysates derived from untreated or heat shocked samples were examined using denaturing polyacrylamide gel electrophoresis for changes in: (i) the pattern of general protein synthesis by labelling epithelial cells with 35S-methionine and (ii) the levels of soluble Hsp70 protein and related isoforms using immunochemical protein blots.Results—A single pinch biopsy specimen is sufficient to extract and analyse specific sets of polypeptides in the oesophageal epithelium. After ex vivo heat shock, a classic inhibition of general protein synthesis is observed and correlates with the increased synthesis of two major proteins of molecular weight of 60 and 70 kDa. Notably, cells from unheated controls exhibit a “stressed” biochemical state 22 hours after incubation at 37°C, as shown by inhibition of general protein synthesis and increased synthesis of the 70 kDa protein. These data indicate that only freshly acquired specimens are suitable for studying stress responses ex vivo. No evidence was found that the two heat induced polypeptides are previously identified Hsp70 isoforms. In fact, heat shock results in a reduction in the steady state concentrations of Hsp70 protein in the oesophageal epithelium.Conclusion—Systematic and highly controlled studies on protein biochemistry are possible on epithelial biopsy specimens from the human oesophagus. These technical innovations have permitted the discovery of a novel heat shock response operating in the oesophageal epithelium. Notably, two polypeptides were synthesised after heat shock that seem to differ from Hsp70 protein. In addition, the striking reduction in steady state concentrations of Hsp70 protein after heat shock suggests that oesophageal epithelium has evolved an atypical biochemical response to thermal stress.

Hydrogen peroxide induces heat shock protein and proto-oncogene mRNA accumulation in Xenopus laevis A6 kidney epithelial cells

2004 ◽  
Vol 82 (7) ◽  
pp. 523-529 ◽  
Author(s):  
M Muller ◽  
J Gauley ◽  
John J Heikkila
Keyword(s):  
Hydrogen Peroxide ◽  
Heat Shock ◽  
Epithelial Cells ◽  
Heat Shock Protein ◽  
Mrna Levels ◽  
Mrna Accumulation ◽  
Disease States ◽  
Hydrogen Peroxide Treatment ◽  
Kidney Epithelial Cells ◽  
Shock Proteins

In this study, we examined the effect of hydrogen peroxide on the accumulation of various mRNAs encoding heat shock proteins (hsps) and proto-oncogenes in Xenopus A6 kidney epithelial cells. Hydrogen peroxide treatment enhanced the accumulation of hsp90, hsp70, hsp30, c-jun, c-fos, and actin mRNAs with distinct temporal patterns. Although hsp70, c-fos, and c-jun mRNA levels peaked at 1–2 h before declining, hsp30 and hsp90 mRNA levels were maximal at 4–6 h. Other mRNAs, including heat shock cognate hsc70, immunoglobulin binding protein, and ribosomal L8, were unaffected. Treatment of kidney cells with a combination of mild heat shock plus hydrogen peroxide resulted in a synergistic increase in the relative levels of both hsp70 and hsp30 mRNA, but not hsp90, c-fos, c-jun, or actin. This study suggests that analysis of hsp and proto-oncogene mRNA levels may be of value as molecular biomarkers of oxidative stress associated with various disease states and nephrotoxicity in kidney.Key words: Xenopus, kidney, mRNA, heat shock protein, hydrogen peroxide.

Examination of heat shock protein mRNA accumulation in early Xenopus laevis embryos

1987 ◽  
Vol 65 (2) ◽  
pp. 87-94 ◽  
Author(s):  
J. J. Heikkila ◽  
N. Ovsenek ◽  
P. Krone
Keyword(s):  
Heat Shock ◽  
Xenopus Laevis ◽  
Heat Shock Protein ◽  
Incubation Temperature ◽  
Effect Of Temperature ◽  
Continuous Exposure ◽  
Hsp 70 ◽  
Mrna Accumulation ◽  
Mrna Induction ◽  
Xenopus Laevis Embryos

Elevation of the incubation temperature of Xenopus laevis neurulae from 22 to 33–35 °C induced the accumulation of heat shock protein (hsp) 70 mRNA (2.7 kilobases (kb)) and a putative hsp 87 mRNA (3.2 kb). While constitutive levels of both hsp mRNAs were detectable in unfertilized eggs and cleavage-stage embryos, heat-induced accumulation was not observed until after the mid-blastula stage. Exposure of Xenopus laevis embryos to other stressors, such as sodium arsenite or ethanol, also induced a developmental stage-dependent accumulation of hsp 70 mRNA. To characterize the effect of temperature on hsp 70 mRNA induction, neurulae were exposed to a range of temperatures (27–37 °C) for 1 h. Heat-induced hsp 70 mRNA accumulation was first detectable at 27 °C, with relatively greater levels at 30–35 °C and lower levels at 37 °C. A more complex effect of temperature on hsp 70 mRNA accumulation was observed in a series of time course experiments. While continuous exposure of neurulae to heat shock (27–35 °C) induced a transient accumulation of hsp 70 mRNA, the temporal pattern of hsp 70 mRNA accumulation was temperature dependent. Exposure of embryos to 33–35 °C induced maximum relative levels of hsp 70 mRNA within 1–1.5 h, while at 30 and 27 °C peak hsp 70 mRNA accumulation occurred at 3 and 12 h, respectively. Finally, placement of Xenopus neurulae at 22 °C after a 1-h heat shock at 33 °C produced an initial decrease in hsp 70 mRNA within 15–30 min, followed by a transient increase in hsp 70 mRNA at 1–2 h before decaying to background levels by 7 h.

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