Putative Determinants of the Cellular Response to Hyperthermia

Laszlo, Andrei; Davidson, Teri; Hu, Anzi; Landry, Jacques; Bedford, Joel S.

doi:10.1080/09553009314450751

Cited by 21 publications

(11 citation statements)

References 26 publications

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“…In some cells following heat shock and recovery, we observed translocation from the cytoplasm to nucleus as previously reported by others (Laszlo et al, 1993;Snoeckx et al, 1991;Welch and Feramisco, 1984). HSP70 translocation from the nucleus back to the cytoplasm was found in fibroblasts at about 8 h (Welch and Suhan, 1986), and the redistribution occurred more rapidly in transient or permanently heat-resistant cells than their normal counterpart (Ohtsuka and Laszlo, 1992).…”

Section: Conclusion and Discussionsupporting

confidence: 81%

HSP70 kinetics study by continuous observation of HSP–GFP fusion protein expression on a perfusion heating stage

Wang

Xie

Rylander

et al. 2007

Biotech & Bioengineering

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The direct correlation between levels of heat shock protein expression and efficiency of its tissue protection function motivates this study of how thermal doses can be used for an optimal stress protocol design. Heat shock protein 70 (HSP70) expression kinetics were visualized continuously in cultured bovine aortic endothelial cells (BAECs) on a microscope heating stage using green fluorescent protein (GFP) as a reporter. BAECs were transfected with a DNA vector, HSP(p)-HSP70-GFP which expresses an HSP70-GFP fusion protein under control of the HSP70 promoter. Expression levels were validated by western blot analysis. Transfected cells were heated on a controlled temperature microscope stage at 42 degrees C for a defined period, then shifted to 37 degrees C for varied post-heating times. The expression of HSP70-GFP and its sub-cellular localization were visualized via fluorescence microscopy. The progressive expression kinetics were measured by quantitative analysis of serial fluorescence images captured during heating protocols from 1 to 2 h and post-heating times from 0 to 20 h. The results show two sequential peaks in HSP70 expression at approximately 3 and 12 h post-heat shock. A progressive translocation of HSP70 from the cytoplasm to the nucleus was observed from 6 to 16 h. We conclude that we have successfully combined molecular cloning and optical imaging to study HSP70 expression kinetics. The kinetic profile for HSP70-GFP fusion protein is consistent with the endogenous HSP70. Furthermore, information on dynamic intracellular translocation of HSP70 was extracted from the same experimental data.

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Section: Conclusion and Discussionsupporting

confidence: 81%

HSP70 kinetics study by continuous observation of HSP–GFP fusion protein expression on a perfusion heating stage

Wang

Xie

Rylander

et al. 2007

Biotech & Bioengineering

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“…p38 hog activation may therefore generate protective mechanisms in response to cellular stresses. This is consistent with data showing that p38 hog , as an activator MAP kinase-associated protein kinase II (MAPKKAP kinase II), phosphorylates, and activates small molecular weight heat shock proteins providing protection from heat shock (33)(34)(35)(36). A subtle balance may thus be struck between repair and growth arrest after stress stimuli that may be mediated differentially through each of these two pathways.…”

Section: Discussionsupporting

confidence: 77%

Mammalian Mitogen-activated Protein Kinase Pathways Are Regulated through Formation of Specific Kinase-Activator Complexes

Zanke

Rubie

Winnett

et al. 1996

Journal of Biological Chemistry

105

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Mammalian cells contain at least three signaling systems which are structurally related to the mitogen-activated protein kinase (MAPK) pathway. Growth factors acting through Ras primarily stimulate the Raf/MEK/ MAPK cascade of protein kinases. In contrast, many stress-related signals such as heat shock, inflammatory cytokines, and hyperosmolarity induce the MEKK/SEK-(MKK4)/SAPK(JNK) and/or the MKK3 or MKK6/p38 hog pathways. Physiological agonists of these pathway types are either qualitatively or quantitatively distinct, suggesting few common proximal signaling elements, although past studies performed in vitro, or in cells using transient over-expression, reveal interaction between the components of all three pathways. These studies suggest a high degree of cross-talk apparently not seen in vivo. We have examined the possible molecular basis of the differing agonist profiles of these three MAPK pathways. We report preferential association between MAP kinases and their activators in eukaryotic cells. Furthermore, using the yeast 2-hybrid system, we show that association between these components can occur independent of additional eukaryotic proteins. We show that SAPK(JNK) or p38 hog activation is specifically impaired by co-expression of cognate dominant negative MAP kinase kinase mutants, demonstrating functional specificity at this level. Further divergence and insulation of the stress pathways occurs proximal to the MAPK kinases since activation of the MAPK kinase kinase MEKK results in SAPK(JNK) activation but does not cause p38 hog phosphorylation. Therefore, in intact cells, the three MAPK pathways may be independently regulated and their components show specificity in their interaction with cognate cascade members. The degree of intermolecular specificity suggests that mammalian MAPK signaling pathways may remain distinct without the need for specific scaffolding proteins to sequester components of individual pathways.Genetic studies in yeast first revealed the existence of multiple distinct mitogen-activated protein kinase (MAPK) 1 related signal transduction pathways containing structurally similar protein kinase cascades mediating responses to mating factor, hyperosmolarity, and cell wall integrity (1). Pathways in mammals, which likely have different physiologic significance, have been discovered which utilize related protein kinase cascades (2-4). The archetypal MAPK pathway is activated in response to Ras-GTP loading as well as other processes such as phosphatidylinositol turnover (5-7). The pathway comprises a series of protein kinases such that activation of the Raf protooncogene causes phosphorylation and activation of MEK which, in turn, phosphorylates and activates the MAPKs, Erk1, and -2 (4, 8). The targets of these kinases include other protein kinases and transcription factors such as p62 tcf (9 -11). While mitogens and growth factors commonly stimulate this MAPK pathway, cells respond to cellular stress agents by induction of two structurally related but distinct pathways (2, 12-16). Stres...

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“…In contrast to the reported more rapid recovery from heat-induced radiosensitization in transiently thermotolerant cells [29], recovery was similar in wild type and heat-resistant cells (Figures 4a and b). This observation also supports the notion that the permanently heat-resistant state and the transiently thermotolerant state are different [30,33]. Since, in the above described examples, resistance to heat-induced cell killing does not lead to resistance to heat-induced radiosensitization and, conversely, sensitization to heatinduced cell killing does not increase heat-induced radiosensitization, the pathways that lead to resistance to heat-induced radiosensitization do not appear to be identical to those that lead to resistance to heat-induced cell killing.…”

Section: Discussionsupporting

confidence: 83%

“…Two sets of results support the possibility that faster recovery from heat-induced radiosensitization could account for the differences seen above. Faster recovery from heat-induced perturbations in cellular physiology has been found to be associated with the heat resistance of HR-1 and OC-14 cells [25,30,33] and faster recovery from heat-induced radiosensitization has been observed in transiently thermotolerant cells [29,34]. The recovery from heat-induced radiosensitization was determined from treatments involving isodoses of ionizing radiation ( Figure 4a) and from doses of radiation with similar initial survival (Figure 4b), all in combination with an initial exposure of 1 h to 438C.…”

Section: Effect Of Hyperthermia Combined With Radiationmentioning

confidence: 99%

Alterations in heat-induced radiosensitization accompanied by nuclear structure alterations in Chinese hamster cells

Laszlo

Davidson

Harvey

et al. 2006

International Journal of Hyperthermia

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This paper examined heat-induced radiosensitization in two Chinese hamster heat-resistant cell lines, HR-1 and OC-14, that were isolated from the same wild-type HA-1 cell line. It found a reduction of the magnitude of heat-induced radiosensitization after exposure to 43 degrees C in both HR-1 and OC-14 cells and a similar reduction after exposure to 45 degrees C in HR-1 cells, but not in OC-14 cells. The effect of heat exposure on a class of ionizing radiation-induced DNA damage that inhibits the ability of nuclear DNA to undergo super-coiling changes was also studied using the fluorescent halo assay in these three cell lines. Wild type cells exposed to either 43 or 45 degrees C before irradiation had a DNA rewinding ability that was intermediate between control and unheated cells, a phenomenon previously described as a masking effect. This masking effect was significantly reduced in HR-1 cells exposed to either 43 or 45 degrees C or in OC-14 cells exposed to 43 degrees C under conditions that heat-induced radiosensitization was reduced. In contrast, the masking effect was not altered in OC-14 cells exposed to 45 degrees C, conditions under which heat-induced radiosensitization was similar to that observed in wild-type HA-1 cells. These results suggest that a reduction in the masking effect is associated with a reduction of the magnitude of heat-induced radiosensitization in the HR-1 and OC-14 heat-resistant cell lines. The reduction of the masking effect in the cell lines resistant to heat-induced radiosensitization was associated with neither a reduction in the magnitude of the heat-induced increase in total nuclear protein content nor major differences in the protein profiles of the nucleoids isolated from heated cells.

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Putative Determinants of the Cellular Response to Hyperthermia

Cited by 21 publications

References 26 publications

HSP70 kinetics study by continuous observation of HSP–GFP fusion protein expression on a perfusion heating stage

HSP70 kinetics study by continuous observation of HSP–GFP fusion protein expression on a perfusion heating stage

Mammalian Mitogen-activated Protein Kinase Pathways Are Regulated through Formation of Specific Kinase-Activator Complexes

Alterations in heat-induced radiosensitization accompanied by nuclear structure alterations in Chinese hamster cells

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