Stress-Related Protein Synthesis in Mammalian Cells Exposed to Hydrostatic Pressure
Professor Timo Ylikomi, University of Tampere, Finland
Emeritus Professor Pekka Mäenpää, University of Kuopio, Finland
Docent Tuula Nyman, Finnish Institute of Occupational Health Helsinki, Finland
Docent Mikko Lammi, University of Kuopio, Finland
Professor Heikki Helminen, University of Kuopio, Finland
Articles of the thesis
In many cases, adequate mechanical stress is required by cells and tissues to maintain their normal functional structure and com position. On the other hand, mechanical forces of unphysiological mode or magnitude may damage or even kill their target cells. This work was designed to study and compare the stress response in two mammalian continuous cell lines after exposures to hydrostatic pressure (HP), to elevated temperature or to various chemicals.
SV40-immortalized T/C28a4 humanchondrocytic cells and human HeLa cervical carcinoma cells were pressurized in a custom-made pressurization chamber. The results were compared to those observed when the cells were exposed to elevated temperature (HS) or chemicals known to interfere intracellular Ca2+ homeostasis. When the changes in protein synthesis were analyzed with two-dimensional electrophoresis and mass spectrometry, it was found that the synthesis of certain stress proteins had been changed. Of the different pressure treatments, only continuous 30 MPa HP resulted in a marked down-regulation of total protein synthesis, and simultaneous up-regulation of heat shock protein 70 (Hsp70) and the Hsp90beta isoform. This up-regulation of stress proteins differed from that seen after HS or treatments with chemical agents, suggesting that disturbed Ca2+ homeostasis was not responsible for the stress response seen after high continuous HP. Expression of Hsp90alpha isoform remained at a constant level, possibly indicating functional differences between these two isoforms during HP loading.
Hsp90 is known to influence the Hsp70 gene expression, e.g., by repressing the activity of HSF1 transcription factor. The DNA-binding activity of HSF1 was increased in Hsp90 inhibitor geldanamycin (GA)-treated cells but, in line with previous studies, it remained unchanged in HP-treated cells. However, simultaneous treatment with GA and HP decreased the HSF1 DNA-binding activity. When Hsp70 mRNA and protein levels were elevated, the effect of GA alone or together with HP loading was stronger compared with that seen in cultures treated only with HP. The increased expression of Hsp70 was not only due to transriptional activation of the Hsp70 gene, but also to the fact that Hsp70 mRNA was stabilized during HP as well as GA treatments.
We further investigated how HP could change the synthesis of protein unlikely to be major stress proteins. A protein whose synthesis was decreased in both of the cell line during continuous 30 MPa HP was identified as eukaryotic elongation factor-2 (eEF-2). The reduced total levels of eEF-2 protein may be related to a decrease in protein synthesis since there was no change in the phosphorylation-regulated activity of this elongation factor.
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Elo MA, Sironen RK, Karjalainen HM, Kaarniranta K, Helminen HJ, Lammi MJ: Specific induction of heat shock protein 90beta by high hydrostatic pressure. Biorheology 40(1-3):141-146, 2003 [Pubmed] [Full text]
Elo MA, Kaarniranta K, Helminen HJ, Lammi MJ: Hsp90 inhibitor geldanamycin increases hsp70 mRNA stabilisation but fails to activate HSF1 in cells exposed to hydrostatic pressure. Biochim Biophys Acta - Mol Cell Res 1743(1-2):115-119, 2005 [Pubmed] [Full text]
Elo MA, Karjalainen HM, Sironen RK, Valmu L, Redpath NT, Browne GJ, Kalkkinen N, Helminen HJ, Lammi MJ: High hydrostatic pressure inhibits the biosynthesis of eukaryotic elongation factor-2. J Cell Biochem 94(3):497-507, 2005 [Pubmed] [Full text]
Kopakkala-Tani M, Elo MA, Sironen RK, Helminen HJ, Lammi MJ: High hydrostatic pressure induces ERK and PI3 kinase phosphorylation in human HCS-2/8 chondrosarcoma cells. Cell Mol Biol 50(4):485-490, 2004 [Pubmed] [Full text]
Last updated January 29, 2019