Phosphorylation at tyrosine-524 influences nuclear accumulation of HSP72 with heat stress.
Nuclear accumulation of heat shock protein (HSP) 72 occurs after cardiac ischemia. This nuclear accumulation of HSP72 with stress occurs in other tissues and species. We postulated that nuclear accumulation of HSP72 was important for the protective effect of HSP72 and that phosphorylation of a single tyrosine (Y(524)) regulated nuclear accumulation of HSP72. Western blots of immunoprecipitated HSP72 from Cos-1 cells demonstrated that tyrosine becomes phosphorylated after heat shock. Treatment with the tyrosine kinase inhibitor geldanamycin blocked nuclear accumulation of HSP72 with heat shock. Two epitope-tagged constructs were made: M17 converting Y(524) to aspartic acid (pseudophosphorylation) and M18 converting Y(524) to phenylalanine. When transfected into Cos-1 cells, M17 accumulates more rapidly and M18 less rapidly than wild-type (WT) HSP72 in the nucleus following heat shock. Cells expressing M18 had less viability after heat shock at 43.5 degrees C than other constructs. After heat shock at 45 degrees C, cells expressing M17 had superior survival compared with WT and M18. These data suggest that phosphorylation at Y(524) facilitates nuclear accumulation of HSP72 following heat stress, and substitution of aspartic acid at Y(524) enhances resistance to heat-shock injury.
Knowlton
A A
AA
Veterans Administration Medical Center, and Baylor College of Medicine, Houston 77030, USA. annek@bcm.tmc.edu
Grenier
M
M
Kirchhoff
S R
SR
Salfity
M
M
D000595:Amino Acid Sequence; D000818:Animals; D019556:COS Cells; D002467:Cell Nucleus / Q000378:metabolism*; D002470:Cell Survival / Q000502:physiology; D050884:HSP72 Heat-Shock Proteins; D018882:Heat Stress Disorders / Q000378:metabolism*; D006360:Heat-Shock Proteins / Q000235:genetics* / Q000378:metabolism*; D009154:Mutation / Q000502:physiology; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors; D014443:Tyrosine / Q000378:metabolism
D050884:HSP72 Heat-Shock Proteins; D006360:Heat-Shock Proteins; D014443:Tyrosine; D011505:Protein-Tyrosine Kinases