Inhibition of heat-induced apoptosis in human tumor cells by heat shock protein 70 occurs upstream of mitochondrial membrane permeabilization through suppression ofc-Jun N-terminal kinase activity

Exposure of cells to hyperthermia induces the synthesis of heat shock proteins. These molecular chaperones help cells cope with protein-damaging stresses by preventing and repairing misfoided proteins. Overexpression of heat shock protein 70 (Hsp70) can protect cells from stress-induced apoptosis. Several studies have revealed multiple mechanisms of apoptosis suppression by Hsp70, including effects occurring both upstream and downstream of mitochondrial membrane permeabilization. Hsp70 is often overexpressed in tumor cells and inhibition of stress-induced apoptosis by Hsp70 is believed to be a contributing factor in tumorigenesis. Suppression of this ability could increase the effectiveness of anti-tumor therapy. Tumor cells exist in an acidic environment, although their intracellular pH is maintained in a physiological range. Acute acidification can sensitize tumor cells to heat-induced cell death. However, the ability of Hsp70 to prevent apoptosis under these conditions has not been examined. The objectives of this thesis were to examine the role of Hsp70 in suppressing heat-induced apoptosis and to evaluate the ability of Hsp70 to suppress apoptosis under conditions of acute acidification. A human acute lymphoblastic T cell line with tetracycline-regulated expression of Hsp70 was used for these studies. Hsp70 was found to inhibit stress-induced c-jun N-terminal kinase (JNK) activation and this inhibition prevented apoptosis by blocking the translocation of the pro-apoptotic protein Bax (B cell lymphoma 2 associated protein X) to mitochondria. Inhibition of JNK with SP600125 or by expression of a dominant negative mutant of JNK blocked Bax translocation as effectively as Hsp70 overexpression. Furthermore, acute acidification sensitized Hsp70-expressing cells to heat-induced apoptosis and this was correlated with a reduced ability to suppress JNK activation. These results indicate that Hsp70 blocks heat-induced apoptosis by inhibiting JNK mediated Bax translocation. As well, the ability of Hsp70 to suppress JNK is compromised when cells are exposed to hyperthermia in an acidic environment resulting in elevated levels of apoptosis. This suggests that the tumor cell environment may be exploited to increase the effectiveness of thermal therapy of tumor cells with elevated levels of Hsp70.