Phosphorylation-dependent cellular localization and thermoprotective role of heat shock protein 25 in hippocampal progenitor cells.
The present study examined phosphorylation-dependent cellular localization and the thermoprotective role of heat shock protein (HSP) 25 in hippocampal HiB5 cells. HSP25 was induced and phosphorylated by heat shock (at 43 degrees C for 3 h). HSP25, which was located in the cytoplasm in the normal condition, translocated into the nucleus after the heat shock. Transfection experiments with hsp27 mutants in which specific serine phosphorylation residues (Ser(78) and Ser(82)) were substituted with alanines or aspartic acids showed that phosphorylation of HSP27 is accompanied by its nuclear translocation. Phosphorylation of mitogen-activated protein kinases (MAPKs) such as p38 MAPK and ERK was markedly increased by the heat shock, and SB203580 (a p38 MAPK kinase inhibitor) and/or PD098059 (a MEK inhibitor) inhibited the phosphorylation of HSP25, indicating that p38 MAPK and ERK are upstream regulators of HSP25 phosphorylation in the heat shock condition. In the absence of heat shock, actin filament stability was not affected by SB203580 and/or PD098059. Heat shock caused disruption of the actin filament and cell death when phosphorylation of HSP25 was inhibited by SB203580 and/or PD098059. In addition, actin filament was more stable in Asp(78,82)-hsp27 (mimics the phosphorylated form) transfected HiB5 cells than in the normal and Ala(78,82)-hsp27 (nonphosphorylative form) transfected cells. In accordance with actin filament stability, the survival rate against the heat shock increased markedly in Asp(15,78,82)-hsp27 expressing HiB5 cells but decreased in Ala(15,78,82)-hsp27 expressing cells. These results support the idea that phosphorylation of HSP25 is critical for the maintenance of actin filament and enhancement of thermoresistance. Interestingly, HSP25 was dephosphorylated and returned to cytoplasm in a recovery time-dependent manner. This phenomenon was accompanied by an increment of apoptotic cell death as determined by nuclear and DNA fragmentation and fluorescence-activated cell sorter analysis. These results suggest that nuclear-translocated HSP25 might function to protect nuclear structure, thereby preventing apoptotic cell death.
Geum
Dongho
D
School of Biological Sciences, Seoul National University, Seoul 151-742, Korea.
Son
Gi Hoon
GH
Kim
Kyungjin
K
D021581:Active Transport, Cell Nucleus; D000409:Alanine / Q000737:chemistry; D000818:Animals; D017209:Apoptosis; D001224:Aspartic Acid; D015152:Blotting, Northern; D015153:Blotting, Western; D002460:Cell Line; D002467:Cell Nucleus / Q000378:metabolism; D015638:Cytochalasin D / Q000494:pharmacology; D003593:Cytoplasm / Q000378:metabolism; D053938:DNA Fragmentation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005419:Flavonoids / Q000494:pharmacology; D005434:Flow Cytometry; D055551:HSP27 Heat-Shock Proteins; D006360:Heat-Shock Proteins; D006624:Hippocampus / Q000166:cytology*; D006358:Hot Temperature; D007093:Imidazoles / Q000494:pharmacology; D007150:Immunohistochemistry; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D008957:Models, Genetic; D009154:Mutation; D009363:Neoplasm Proteins / Q000378:metabolism* / Q000502:physiology*; D010766:Phosphorylation; D011233:Precipitin Tests; D011419:Propidium / Q000494:pharmacology; D011725:Pyridines / Q000494:pharmacology; D051381:Rats; D012694:Serine / Q000378:metabolism; D015398:Signal Transduction; D013234:Stem Cells / Q000378:metabolism*; D013696:Temperature; D013997:Time Factors; D014162:Transfection; D048051:p38 Mitogen-Activated Protein Kinases
D004791:Enzyme Inhibitors; D005419:Flavonoids; D055551:HSP27 Heat-Shock Proteins; D006360:Heat-Shock Proteins; C486114:Hspb1 protein, rat; D007093:Imidazoles; D009363:Neoplasm Proteins; D011725:Pyridines; D015638:Cytochalasin D; D001224:Aspartic Acid; D011419:Propidium; D012694:Serine; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D000409:Alanine; C093642:SB 203580; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one