Cytosolic Accumulation of HSP60 during Apoptosis with or without Apparent Mitochondrial Release

Abstract

Most heat shock proteins (HSPs) have pro-survival functions. However, the role of HSP60, a mitochondrial matrix protein, is somewhat controversial with both pro-survival and pro-apoptotic functions reported. Here we show that in numerous apoptotic systems HSP60 protein accumulates in the cytosol. In BMD188-induced cell death, HSP60 accumulates in the cytosol with significant mitochondrial release. In contrast, in apoptosis induced by multiple other inducers, the cytosolic HSP60 accumulates without an apparent mitochondrial release. The short interfering RNA-mediated knockdown experiments revealed that in BMD188-induced apoptosis, HSP60 has a pro-death function and that the pro-death role of HSP60 seems to involve caspase-3 maturation and activation in the cytosol. In contrast, HSP60 appears to play a pro-survival role in other apoptotic systems where there is no apparent mitochondrial release as its knockdown promotes cell death. In these latter apoptotic systems HSP60 does not associate with active caspase-3. In both cases, HSP60 does not appreciably interact with Bax. Taken together, our results suggest the following: 1) cytosolic accumulation of HSP60 represents a common phenomenon during apoptosis induction; 2) cytosolic HSP60 accumulation during apoptosis occurs either with or without apparent mitochondrial release; and 3) the cytosolically accumulated HSP60 possesses either pro-survival or pro-death functions, which involves differential interactions with caspase-3.

The dissociation of the Hsp60/pro-Caspase-3 complex by bis(pyridyl)oxadiazole copper complex ( CubipyOXA ) leads to cell death in NCI-H292 cancer cells

“…Hsp60 has been implicated in carcinogenesis via its interaction with components of the Caspase cascade that leads to apoptosis. However, opinions about the role of this chaperone in carcinogenesis, and in apoptosis in particular, differ and it is not yet clear when and how Hsp60 is pro-apoptotic (i.e., it acts as an anti-cancer factor) or the contrary, namely it works in favor of the tumor by interfering with apoptosis [5][6][7][8][9][10].…”

Mitochondrial DNA variability modulates mRNA and intra-mitochondrial protein levels of HSP60 and HSP75: experimental evidence from cybrid lines

“…HSP60 (Cpn60) and HSP75 (TRAP1) are mitochondrial chaperones that assist, in both stressed and non-stressed cells, in the folding, unfolding, or disaggregating of proteins either imported from the cytosol or synthesized within mitochondria (HSP60) (Cheng et al 1989;Frydman 2001;Itoh et al 2002;Saibil 2008) and in the reallocation of cytosolic protein into mitochondria (HSP75) (Felts et al 2000;Mokranjac and Neupert 2005). In addition to its chaperone activity, HSP60 has welldocumented anti-or pro-apoptotic roles (Arya et al 2007;Chandra et al 2007) as well as immunoregulatory properties (Habich and Burkart 2007;Pockley et al 2008). HSP75 acts as an antagonist of ROS and exhibits anti-apoptotic activity (Hua et al 2007;Pridgeon et al 2007).…”

Resveratrol Induces p53-independent, X-linked Inhibitor of Apoptosis Protein (XIAP)-mediated Bax Protein Oligomerization on Mitochondria to Initiate Cytochrome c Release and Caspase Activation

“…Prostate cancer (PC3 and LNCaP), breast cancer (MDA-MB231, MCF-7, and MDA-MB435), immortalized normal human fibroblast (GM701), Jurkat wild type (WT), Jurkat caspase-8 Ϫ/Ϫ , mouse embryonic fibroblast WT, and mouse embryonic fibroblast Apaf-1 Ϫ/Ϫ cells were obtained from the ATCC or from various investigators and were subcultured as described previously (31)(32)(33)(34)(35)(36). The primary antibodies against cytochrome c (monoclonal antibody (mAb)), Apaf-1 and Bax (rabbit polyclonal antibody (Rb pAb)), Bid, caspase-8, XIAP, and Bcl-xL were purchased from BD Pharmingen.…”

“…Finally, the beads were washed thoroughly and analyzed by the TrueBlot (eBioscience) Western blotting system (34).…”

“…Cytosolic and mitochondrial fractions were purified as described previously (31,32,34) and loaded onto a Superdex 200 HR10/30 column (GE Healthcare). Proteins were eluted at 0.5 ml/min, and fractions (0.5 ml) were collected for Western analyses (38).…”