Abstract
Rac1 has been implicated in a wide variety of biological processes, including actin remodeling and various signaling cascades. Here we have examined whether Rac1 might be involved in heat shock-induced cell signaling. We found that Rat2 stable cells expressing a dominant negative Rac1 mutant, RacN17 (Rat2-RacN17), were significantly more tolerant to heat shock than control Rat2 cells, and simultaneously inhibited the activation of SAPK/JNK by heat shock compared to control Rat2 cells. However, no discernible effect was observed in typical heat shock responses including total protein synthesis and heat shock protein synthesis. To identify the proteins involved in this difference, we separated the proteins of both Rat2 and Rat2-RacN17 cell lines after heat shock using two-dimensional gel electrophoresis and identified the differentially expressed proteins by matrix assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF MS) after in-gel trypsin digestion. Differentially expressed proteins between two cell lines were identified as vimentin. Rat2-RacN17 cells showed significant changes in vimentin as well as marked changes in vimentin reorganization by heat shock. The vimentin changes were identified as N-terminal head domain cleavage. These results suggest that Rac1 plays a pivotal role in the heat shock-induced signaling cascade by modifying intermediate vimentin filaments. Cell Death and Differentiation (2001) 8, 1093–1102
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Abbreviations
- HSF:
-
heat shock factor
- HSP:
-
heat shock protein
- SAPK:
-
stress-activated protein kinase
- JNK:
-
c-Jun N-terminal kinase
- GST:
-
glutathione-S-transferase
- PARP:
-
poly(ADP-ribose) polymerase
- SRF:
-
serum response factor
- TNF:
-
tumor necrosis factor
- MALDI-TOF MS:
-
matrix assisted laser desorption/ionization time of flight mass spectrometry
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Acknowledgements
We thank Dr. James R Woodgett for providing GST-c-jun 1-89/pGex-2T plasmid and Dr. M Inagaki for antibody against head domain of vimentin. We thank Dr. HJ Kim in Hyundai Pharm.Ind. Co. for letting us to use MALDI-TOF MS. This work was supported by KOSEF through the Center for Cell Signaling Research (CCSR) at Ewha Womans University and by the Research fund for Womans University (KISTEP, 1998). Students (S-Y Lee, EJ Song, H-J Kim, H-J Kang) were financially supported by Brain Korea 21 program.
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Lee, SY., Song, E., Kim, HJ. et al. Rac1 regulates heat shock responses by reorganization of vimentin filaments: Identification using MALDI-TOF MS. Cell Death Differ 8, 1093–1102 (2001). https://doi.org/10.1038/sj.cdd.4400923
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DOI: https://doi.org/10.1038/sj.cdd.4400923
