Small Hsps as regulators of apoptosis

sHsp (small stress proteins) are molecular chaperones involved in cellular defence mechanisms against several different types of stress (heat shock, oxidative stress) which also participate in essential physiological processes, such as regulation of cell cycle, differentiation and programmed cell death. For example, sHsp are transiently expressed during the cell division to differentiation transition and this phenomenon prevents differentiating cells from undergoing apoptosis. sHsp also protect against apoptosis induced by different conditions or agents, particularly anti-cancer drugs. Of interest, tumor cells usually express high levels of sHsp and several anti-cancer drugs trigger the accumulation of sHsp. sHsp expression also reduce the apoptotic death induced by TNFalpha and FasL and enhance the growth of tumors in vim. sHsp expression is supposed to allow cancerous cells to escape the immunosurveillance mediated by death ligands and to render these cells resistant to therapy. sHsp are therefore prime targets for therapeutical interventions. The protective mechanism by which sHsp protect against apoptosis has been analyzed in murine fibroblasts that express either human Hsp27 or murine Hsp25. We report here that the expression of these sHsp strongly delayed the release of cytochrome c from mitochondria in response to staurosporine or etoposide treatments. This suggests that an antiapoptotic effect mediated by sHsp is localized upstream of cytochrome c release. sHsp were also found to act downstream of cytochrome c release but upstream of caspase 3 activation. This second activity requires a lower level of sHsp expression compared to the first one which takes place upstream of cytochrome Concomitantly with these effects, sHsp phosphorylation status increased and these proteins transiently formed two distinct oligomeric populations. The importance of this structural reorganization was supported by the fact that a Hsp25 unphosphorylatable (Ser 15, 86 to Ala) mutant, unable to generate two populations of oligomers, did not protect against staurosporine but on the contrary stimulated caspase 9 and 3 activation and cell death. We also analyzed HeLa cells that underexpress Hsp27. It was observed that these cells are more sensitive to apoptosis than their normal counterparts; a phenomenon characterized by a more rapid release of cytochrome c from mitochondria and activation of caspases. Our results therefore suggest that mammalian sHsp negatively regulate the apoptotic process by acting both upstream and downstream of cytochrome c release from mitochondria.