Abstract
For many years, there has been uncertainty concerning the reason for Hsp70 translocation to the nucleus and nucleolus. Herein, we propose that Hsp70 translocates to the nucleus and nucleoli in order to participate in pathways related to the protection of the nucleoplasmic DNA or ribosomal DNA from single-strand breaks. The absence of Hsp70 in HeLa cells, via Hsp70 gene silencing (knockdown), indicated the essential role of Hsp70 in DNA integrity. Therefore, HeLa Hsp70 depleted cells were very sensitive in heat treatment and their DNA breaks were multiple compared to that of control HeLa cells. The molecular mechanism with which Hsp70 performs its role at the level of nucleus and nucleolus during stress was examined. Hsp70 co-localizes with PARP1 in the nucleus/nucleoli as was observed in confocal studies and binds to the BCRT domain of PARP1 as was revealed with protein–protein interaction assays. It was also found that Hsp70 binds simultaneously to XRCC1 and PARP-1, indicating that Hsp70 function takes place at the level of DNA repair and possibly at the base excision repair system. Making a hypothetical model, we have suggested that Hsp70 is the molecule that binds and interrelates with PARP1 creating the repair proteins simultaneously, such as XRCC1, at the single-strand DNA breaks. Our data partially clarify a previously unrecognized cellular response to heat stress. Finally, we can speculate that Hsp70 plays a role in the quality and integrity of DNA.
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Abbreviations
- NCA:
-
Nucleolar comet assay
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Acknowledgments
We are very thankful to Professor of Biochemistry, Molecular Biology, and Cell Biology, R. Morimoto., Professor of Biological Chemistry D. Galaris and Emeritus Professor Pagoulatos Gerassimos for their support with reagents or useful discussions. We also thank Dr. Doulias P-T., Dr. Skyrlas A., Dr. Markopoulos G., Dr. Noutsopoulos D., and Dr. Zerikiotis S. for their technical and scientific support. This research was partially co-funded by the European Union and the Hellenic Ministry of Education (program “Herakleitos”) within the “Operational Program for Education and Initial Vocational Training. It was also partially supported by a grant to P. Vezyraki from the Empeirikio Institution, Athens.
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Outlining prior scientific knowledge on the subject and novel information: The role of Hsp70 translocation to the nucleus and nucleolus during heat stress has been nearly unknown. It has been proposed that this biological phenomenon is correlated to Hsp70-chaperoning activity. Furthermore, some previous observations in yeast have revealed that Rad9 complexes—Rad9 being the prototype DNA-damage checkpoint gene—contain Ssa1 and or Ssa2 chaperone proteins, both reconstituting the functions of the corresponding Hsp70 in mammalian cells. Here, we propose that Hsp70 translocates to the nuclei/nucleoli during heat stress, binds to PARP-1 and/or XRCC1, and protects HeLa cells from increased single-strand DNA breaks.
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Kotoglou, P., Kalaitzakis, A., Vezyraki, P. et al. Hsp70 translocates to the nuclei and nucleoli, binds to XRCC1 and PARP-1, and protects HeLa cells from single-strand DNA breaks. Cell Stress and Chaperones 14, 391–406 (2009). https://doi.org/10.1007/s12192-008-0093-6
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DOI: https://doi.org/10.1007/s12192-008-0093-6