Distribution, phosphorylation, and activities of Hsp25 in heat-stressed H9c2 myoblasts: a functional link to cytoprotection.
The behavior of the endogenous heat shock protein 25 (Hsp25) in heat-stressed rat H9c2 myoblasts was studied. After mild or severe heating, this protein became less extractable with Triton X-100 and displayed characteristic immunofluorescence patterns, namely (1) granules in the nucleus, and (2) association with F-actin bundles in the cytoplasm. The intranuclear granulation of Hsp25 and its association with F-actin were sensitive to drugs affecting Hsp25 phosphorylation (cantharidin, sodium orthovanadate, SB203580, SB202190). Isoform analysis of Hsp25 translocated to the nucleus-free cytoskeletal fraction revealed only mono- and biphosphorylated Hsp25 and no unphosphorylated Hsp25. Transfected luciferase with initial localization in the nucleosol became colocalized with the Hsp25-containing granules after a heat shock treatment that denatured the enzyme in the cells. The association of Hsp25 with actin filaments after a mild heat stress conferred protection from subsequent F-actin-damaging treatments with cytochalasins (D and B) or severe heat stress. We hypothesize that (1) the binding of heat-denatured nucleosolic proteins to the Hsp25 contained in specific granular structures may serve for the subsequent chaperoning or degradation of the bound proteins, and (2) the actin cytoskeleton is stabilized by the direct targeting of phosphorylated Hsp25 to microfilament bundles.
Bryantsev
Anton L
AL
Molecular and Cellular Cardiology Lab, Institute of Experimental Cardiology, Cardiology Research Centre, Moscow, Russia. cardiocell@cardio.ru
Loktionova
Svetlana A
SA
Ilyinskaya
Olga P
OP
Tararak
Eduard M
EM
Kampinga
Harm H
HH
Kabakov
Alexander E
AE
D000818:Animals; D002467:Cell Nucleus / Q000378:metabolism; D002470:Cell Survival; D003599:Cytoskeleton / Q000378:metabolism; D055551:HSP27 Heat-Shock Proteins; D006360:Heat-Shock Proteins; D006358:Hot Temperature; D007093:Imidazoles / Q000378:metabolism; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D032446:Myoblasts / Q000378:metabolism*; D009363:Neoplasm Proteins / Q000378:metabolism*; D010766:Phosphorylation; D011725:Pyridines / Q000378:metabolism; D051381:Rats; D048051:p38 Mitogen-Activated Protein Kinases
D055551:HSP27 Heat-Shock Proteins; D006360:Heat-Shock Proteins; C486114:Hspb1 protein, rat; D007093:Imidazoles; D009363:Neoplasm Proteins; D011725:Pyridines; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; C093642:SB 203580