Cytosolic heat shock protein 60, apoptosis, and myocardial injury.
BACKGROUND
Heat shock proteins (HSPs) are well known for their ability to "protect" the structure and function of native macromolecules, particularly as they traffic across membranes. Considering the role of key mitochondrial proteins in apoptosis and the known antiapoptotic effects of HSP27 and HSP72, we postulated that HSP60, primarily a mitochondrial protein, also exerts an antiapoptotic effect. Methods and Results- To test this hypothesis, we used an antisense phosphorothioate oligonucleotide to effect a 50% reduction in the levels of HSP60 in cardiac myocytes, a cell type that has abundant mitochondria. The induced decrease in HSP60 precipitated apoptosis, as manifested by the release of cytochrome c, activation of caspase 3, and induction of DNA fragmentation. Antisense treatment was associated with an increase in bax and a decrease in bcl-2 secondary to increased synthesis of bax and degradation of bcl-2. A control oligonucleotide had no effect on these measurements. We further demonstrated that cytosolic HSP60 forms a macromolecular complex with bax and bak in vitro suggesting that complex formation with HSP60 may block the ability of bax and bak to effect apoptosis in vivo. Lastly, we show that as cytosolic (nonmitochondrial) HSP60 decreases, a small unbound fraction of bax appears and that the amount of bax associated with the mitochondria and cell membranes increases.


CONCLUSIONS
These results support a key antiapoptotic role for cytosolic HSP60. To our knowledge, this is the first report suggesting that interactions of HSP60 with bax and/or bak regulate apoptosis.
Kirchhoff
S R
SR
Division of Cardiology Research, Veterans Affairs Medical Center and Baylor College of Medicine, Houston, Texas, USA.
Gupta
S
S
Knowlton
A A
AA
D000818:Animals; D017209:Apoptosis; D002478:Cells, Cultured; D018834:Chaperonin 60 / Q000235:genetics / Q000502:physiology*; D003600:Cytosol / Q000378:metabolism; D008565:Membrane Proteins / Q000378:metabolism; D024101:Mitochondrial Proteins / Q000235:genetics / Q000502:physiology; D009206:Myocardium / Q000166:cytology / Q000378:metabolism*; D016376:Oligonucleotides, Antisense / Q000494:pharmacology; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D050998:bcl-2 Homologous Antagonist-Killer Protein; D051028:bcl-2-Associated X Protein
C494774:Bak1 protein, rat; C494828:Bax protein, rat; D018834:Chaperonin 60; D008565:Membrane Proteins; D024101:Mitochondrial Proteins; D016376:Oligonucleotides, Antisense; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D050998:bcl-2 Homologous Antagonist-Killer Protein; D051028:bcl-2-Associated X Protein