Stressful preconditioning and HSP70 overexpression attenuate proteotoxicity of cellular ATP depletion.
Rat H9c2 myoblasts were preconditioned by heat or metabolic stress followed by recovery under normal conditions. Cells were then subjected to severe ATP depletion, and stress-associated proteotoxicity was assessed on 1) the increase in a Triton X-100-insoluble component of total cellular protein and 2) the rate of inactivation and insolubilization of transfected luciferase with cytoplasmic or nuclear localization. Both heat and metabolic preconditioning elevated the intracellular heat shock protein 70 (HSP70) level and reduced cell death after sustained ATP depletion without affecting the rate and extent of ATP decrease. Each preconditioning attenuated the stress-induced insolubility among total cellular protein as well as the inactivation and insolubilization of cytoplasmic and nuclear luciferase. Transient overexpression of human HSP70 in cells also attenuated both the cytotoxic and proteotoxic effects of ATP depletion. Quercetin, a blocker of stress-responsive HSP expression, abolished the effects of stressful preconditioning but did not influence the effects of overexpressed HSP70. Analyses of the cellular fractions revealed that both the stress-preconditioned and HSP70-overexpressing cells retain the soluble pool of HSP70 longer during ATP depletion. Larger amounts of other proteins coimmunoprecipitated with excess HSP70 compared with control cells deprived of ATP. This is the first demonstration of positive correlation between chaperone activity within cells and their viability in the context of ischemia-like stress.
Kabakov
Alexander E
AE
Medical Radiology Research Center, Obninsk 249020, Russia. aekabakov@hotmail.com
Budagova
Karina R
KR
Latchman
David S
DS
Kampinga
Harm H
HH
D000255:Adenosine Triphosphate / Q000172:deficiency*; D000818:Animals; D016923:Cell Death; D002460:Cell Line; D019610:Cytoprotection; D018840:HSP70 Heat-Shock Proteins / Q000502:physiology*; D006358:Hot Temperature; D006801:Humans; D019157:Ischemic Preconditioning, Myocardial / Q000379:methods; D009124:Muscle Proteins / Q000502:physiology*; D009206:Myocardium / Q000378:metabolism / Q000473:pathology; D018384:Oxidative Stress; D051381:Rats
D018840:HSP70 Heat-Shock Proteins; D009124:Muscle Proteins; D000255:Adenosine Triphosphate