The role of Hsp70 chaperone in the reaction of human leukemic cells to anticancer drugs

Most of anti-tumor factors are designed to kill selectively cancer cells; in most cases this action is related to the ability of the above substances to induce apoptosis. One of potent anti-apoptotic mechanisms is based on Hsp70 protein. Since the level of this protein is often higher in malignant tumors than in normal tissues, the aim of this study was to establish whether the elevated Hsp70 content may influence the process of apoptosis induced by anti-tumor drugs in cancer cells population. The increase of intracellular content of Hsp70 in human leukemia U-937 cells was attained by a mild heat stress or by transfection of cells with the human hsp70 gene. The elevation of Hsp70 quantity, irrespective of the way it was performed, leads to the inhibition of apoptosis in cells treated with two substances, etoposide or adriamycin. The inhibition of apoptosis was accompanied with the reducing of the share of cells with fragmented nuclei and with the delay in caspase activation. It is suggested that in addition to the previously discovered targets, whose activity is suppressed by the Hsp70 chaperone, this protein can inhibit the activity of caspase-3 and -7; this delays the onset of apoptosis in part of a cancer cell population.     

Downstream caspases are novel targets for the antiapoptotic activity of the molecular chaperone hsp70

The response of cancer cells to apoptosis-inducing agents can be characterized by 2 opposing factors, the proapoptotic caspase cascade and the antiapoptotic stress protein Hsp70. We show here that these factors interact in U-937 leukemia cells induced to apoptosis with anticancer drugs, etoposide and adriamycin (ADR). The protective effect of Hsp70 was verified using 2 approaches: mild heat stress and transfection-mediated overexpression of the Hsp70 gene. The increase in Hsp70 levels attained by these 2 methods was found to postpone caspase activation for 12-18 hours. An in vitro assay was developed using mouse myeloma NS0/1 cells, which lack the expression of Hsp70. Measurement of DEVD-ase activity in extracts of apoptotic NS0/1 cells incubated with purified Hsp70 showed that Hsp70 reduced caspase activity by up to 50% of its control value in a dose-dependent manner. The hypothesis that the inhibitory effect of Hsp70 on caspase-3/7 activity related to a direct interaction between Hsp70 and the caspases was tested by reciprocal immunoprecipitations and Far-western analyses. These tests were performed with extracts of Hsp70-overexpressing, control, and ADR-treated U-937 cells and using anti-caspase-3, caspase-7, and anti-Hsp70 antibodies, and the data clearly showed that Hsp70 was able to interact with the proforms of these caspases in cell lysates and with reconstituted purified proteins but did not bind the activated forms of either caspase-3 or -7. This association was also corroborated by a novel, enzyme-linked immunosorbent assay-like assay, protein interaction assay, that combined the advantages of immunoprecipitation and immunoblotting in a 96-well microplate-based assay. Thus, Hsp70 may act to suppress caspase-dependent apoptotic signaling through binding the precursor forms of both caspase-3 and caspase-7 and preventing their maturation.     

18β-Glycyrrhetinic acid potentiates Hsp90 inhibition-induced apoptosis in human epithelial ovarian carcinoma cells via activation of death receptor and mitochondrial pathway

The Hsp90 inhibition has been shown to induce apoptosis in various cancer cells. The licorice compounds may enhance the anti-cancer drug effect. However, effect of the licorice compounds on the Hsp90 inhibition-induced apoptosis in ovarian cancer cells has not been studied. To assess the ability of 18β-glycyrrhetinic acid to promote apoptosis, we examined whether 18β-glycyrrhetinic acid potentiated the Hsp90 inhibitor-induced apoptosis in the human epithelial ovarian carcinoma cell lines OVCAR-3 and SK-OV-3. Radicicol and geldanamycin induced a decrease in Bid, Bcl-2, Bcl-xL and survivin protein levels, an increase in Bax levels, the mitochondrial transmembrane potential loss, cytochrome c release, activation of caspases (-8, -9, and -3), cleavage of PARP-1, and an increase in the tumor suppressor p53 levels. 18β-Glycyrrhetinic acid enhanced Hsp90 inhibitor-induced apoptosis-related protein activation, nuclear damage, and cell death. The results suggest that 18β-glycyrrhetinic acid may potentiate the Hsp90 inhibition-induced apoptosis in ovarian carcinoma cell lines via the activation of the caspase-8- and Bid-dependent pathways and the mitochondria-mediated cell death pathway, leading to activation of caspases. Combination of Hsp90 inhibitors and 18β-glycyrrhetinic acid may confer a benefit in the treatment of epithelial ovarian adenocarcinoma.     

Radicicol, an inhibitor of Hsp90, enhances TRAIL-induced apoptosis in human epithelial ovarian carcinoma cells by promoting activation of apoptosis-related proteins

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) induces apoptosis in various cancer cells. Hsp90 is known to be involved in cell survival and growth in tumor cells. Nevertheless, Hsp90 inhibitors exhibit a variable effect on the cytotoxicity of anticancer drugs. Furthermore, the combined effect of Hsp90 inhibitors on TRAIL-induced apoptosis in epithelial ovarian cancer cells has not been determined. To assess the ability of an inhibitor of Hsp90 inhibitor radicicol to promote apoptosis, we investigated the effect of radicicol on TRAIL-induced apoptosis in the human epithelial ovarian carcinoma cell lines OVCAR-3 and SK-OV-3. TRAIL induced a decrease in Bid, Bcl-2, Bcl-xL, and survivin protein levels, increase in Bax levels, loss of the mitochondrial transmembrane potential, cytochrome c release, activation of caspases (-8, -9, and -3), cleavage of PARP-1 and an increase in the tumor suppressor p53 levels. Radicicol enhanced TRAIL-induced apoptosis-related protein activation, nuclear damage and cell death. These results suggest that radicicol may potentiate the apoptotic effect of TRAIL on ovarian carcinoma cell lines by increasing the activation of the caspase-8- and Bid-dependent pathway and the mitochondria-mediated apoptotic pathway, leading to caspase activation. Radicicol may confer a benefit in the TRAIL treatment of epithelial ovarian adenocarcinoma.     

Hsp70 inhibits heat-induced apoptosis upstream of mitochondria by preventing Bax translocation

Hsp70 overexpression can protect cells from stress-induced apoptosis. Our previous observation that Hsp70 inhibits cytochrome c release in heat-stressed cells led us to examine events occurring upstream of mitochondrial disruption. In this study we examined the effects of heat shock on the proapoptotic Bcl-2 family member Bax because of its central role in regulating cytochrome c release in stressed cells. We found that heat shock caused a conformational change in Bax that leads to its translocation to mitochondria, stable membrane association, and oligomerization. All of these events were inhibited in cells that had elevated levels of Hsp70. Hsp70 did not physically interact with Bax in control or heat-shocked cells, indicating that Hsp70 acts to suppress signals leading to Bax activation. Hsp70 inhibited stress-induced JNK activation and inhibition of JNK with SP600125 or by expression of a dominant negative mutant of JNK-blocked Bax translocation as effectively as Hsp70 overexpression. Hsp70 did not protect cells expressing a mutant form of Bax that has constitutive membrane insertion capability or cells treated with a small molecule activator of apoptosome formation, indicating that it is unable to prevent cell death after mitochondrial disruption and caspase activation have occurred. These results indicate that Hsp70 blocks heat-induced apoptosis primarily by inhibiting Bax activation and thereby preventing the release of proapoptotic factors from mitochondria. Hsp70, therefore, inhibits events leading up to mitochondrial membrane permeabilization in heat-stressed cells and thereby controls the decision to die but does not interfere with cell death after this event has occurred.     

Splicing reprogramming of TRAIL/DISC-components sensitizes lung cancer cells to TRAIL-mediated apoptosis

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) selective killing of cancer cells underlines its anticancer potential. However, poor tolerability and resistance underscores the need to identify cancer-selective TRAIL-sensitizing agents. Apigenin, a dietary flavonoid, sensitizes lung cancer cell lines to TRAIL. It remains unknown, however, whether apigenin sensitizes primary lung cancer cells to TRAIL and its underlying mechanisms. Here we show that apigenin reprograms alternative splicing of key TRAIL/death-inducing-signaling-complex (DISC) components: TRAIL Death Receptor 5 (DR5) and cellular-FLICE-inhibitory-protein (c-FLIP) by interacting with the RNA-binding proteins hnRNPA2 and MSI2, resulting in increased DR5 and decreased c-FLIP_S protein levels, enhancing TRAIL-induced apoptosis of primary lung cancer cells. In addition, apigenin directly bound heat shock protein 70 (Hsp70), promoting TRAIL/DISC assembly and triggering apoptosis. Our findings reveal that apigenin directs alternative splicing and inhibits Hsp70 enhancing TRAIL anticancer activity. These findings underscore impactful synergies between diet and cancer treatments opening new avenues for improved cancer treatments.Subject terms: Cancer, Molecular biology     

Isorhamnetin in Tsoong blocks Hsp70 expression to promote apoptosis of colon cancer cells

The roots of Codonopis bulleynana Forest ex diels (cbFed), locally known as Tsoong, have been used as a tonic food. Tsoong has wide range of pharmacological effects, including anticancer efficacy. In the present study, the anticancer activity of Tsoong and its potential molecular mechanisms were investigated. Isorhamnetin, a flavonol aglycone, is important compound and metabolite in Tsoong. It can promote apoptosis of colon cancer cells through up-regulating apoptosis-related genes (Apaf1, Casp3 and Casp9) because it blocks Hsp70 genes (Hspa1a, Hspa1b and Hspa8). These were verified by in vitro and in vivo experiments. In vitro, cell counting kit-8 (CCK-8) assays and flow cytometry in HCT116 and SW480 colon cancer cell were used to assess the anti-proliferation and apoptosis-promoting activities of Tsoong. In vivo, the antitumor effect of Tsoong was assessed in colon cancer-bearing nude mice as a xenograft model. These results show that Isorhamnetin is very critical in Tsoong because Tsoong can down-regulate Hsp70 genes and promote apoptosis of colon cancer cells by inhibiting Hsp70 largely due to the efficacy of Isorhamnetin. Our results may ultimately help in the development of diagnostic and therapeutic strategies to control this devastating disease.     

Heat Shock Protein 70 Prevents Hyperoxia-Induced Disruption of Lung Endothelial Barrier via Caspase-Dependent and AIF-Dependent Pathways

Exposure of pulmonary artery endothelial cells (PAECs) to hyperoxia results in a compromise in endothelial monolayer integrity, an increase in caspase-3 activity, and nuclear translocation of apoptosis-inducing factor (AIF), a marker of caspase-independent apoptosis. In an endeavor to identify proteins involved in hyperoxic endothelial injury, we found that the protein expression of heat-shock protein 70 (Hsp70) was increased in hyperoxic PAECs. The hyperoxia-induced Hsp70 protein expression is from hspA1B gene. Neither inhibition nor overexpression of Hsp70 affected the first phase barrier disruption of endothelial monolayer. Nevertheless, inhibition of Hsp70 by using the Hsp70 inhibitor KNK437 or knock down Hsp70 using siRNA exaggerated and overexpression of Hsp70 prevented the second phase disruption of lung endothelial integrity. Moreover, inhibition of Hsp70 exacerbated and overexpression of Hsp70 prevented hyperoxia-induced apoptosis, caspase-3 activation, and increase in nuclear AIF protein level in PAECs. Furthermore, we found that Hsp70 interacted with AIF in the cytosol in hyperoxic PAECs. Inhibition of Hsp70/AIF association by KNK437 correlated with increased nuclear AIF level and apoptosis in KNK437-treated PAECs. Finally, the ROS scavenger NAC prevented the hyperoxia-induced increase in Hsp70 expression and reduced the interaction of Hsp70 with AIF in hyperoxic PAECs. Together, these data indicate that increased expression of Hsp70 plays a protective role against hyperoxia-induced lung endothelial barrier disruption through caspase-dependent and AIF-dependent apoptotic pathways. Association of Hsp70 with AIF prevents AIF nuclear translocation, contributing to the protective effect of Hsp70 on hyperoxia-induced endothelial apoptosis. The hyperoxia-induced increase in Hsp70 expression and Hsp70/AIF interaction is contributed to ROS formation.     

Spontaneous apoptosis in primary cultures of human and rat hepatocytes: molecular mechanisms and regulation by dexamethasone

To elucidate the biochemical pathways leading to spontaneous apoptosis in primary cultures of human and rat hepatocytes, we examined the activation of the caspase cascade, the expression of Bcl-2-related-proteins and heat shock proteins. Comparisons were made before and after dexamethasone (DEX) treatment. We show that DEX inhibited spontaneous apoptosis in a dose-dependent manner. DEX increases the expression of anti-apoptotic Bcl-2 and Bcl-x(L) proteins, decreases the expression of pro-apoptotic Bax and inhibits Bad translocation thereby preventing the release of cytochrome c, the activation of caspases, and cell death. Although, the expression of Hsp27 and Hsp70 proteins remained unchanged, the oncogenic protein c-Myc is upregulated upon DEX-treatment. These results indicate that DEX mediates its survival effect against spontaneous apoptosis by acting upstream of the mitochondrial changes. Thus, the mitochondrial apoptotic pathway plays a major role in regulating spontaneous apoptosis in these cells. Blocking this pathway therefore may assist with organ preservation for transplant, drug screening, and other purposes.     

Hsp70 Regulates Immune Response in Experimental Autoimmune Encephalomyelitis

Heat shock protein (Hsp)70 is one of the most important stress-inducible proteins. Intracellular Hsp70 not only mediates chaperone-cytoprotective functions but can also block multiple steps in the apoptosis pathway. In addition, Hsp70 is actively released into the extracellular milieu, thereby promoting innate and adaptive immune responses. Thus, Hsp70 may be a critical molecule in multiple sclerosis (MS) pathogenesis and a potential target in this disease due to its immunological and cytoprotective functions. To investigate the role of Hsp70 in MS pathogenesis, we examined its immune and cytoprotective roles using both in vitro and in vivo experimental procedures. We found that Hsp70.1-deficient mice were more resistant to developing experimental autoimmune encephalomyelitis (EAE) compared with their wild-type (WT) littermates, suggesting that Hsp70.1 plays a critical role in promoting an effective myelin oligodendrocyte glycoprotein (MOG)-specific T cell response. Conversely, Hsp70.1-deficient mice that developed EAE showed an increased level of autoreactive T cells to achieve the same production of cytokines compared with the WT mice. Although a neuroprotective role of HSP70 has been suggested, Hsp70.1-deficient mice that developed EAE did not exhibit increased demyelination compared with the control mice. Accordingly, Hsp70 deficiency did not influence the vulnerability to apoptosis of oligodendrocyte precursor cells (OPCs) in culture. Thus, the immunological role of Hsp70 may be relevant in EAE, and specific therapies down-regulating Hsp70 expression may be a promising approach to reduce the early autoimmune response in MS patients.     

Hsp60 regulation of tumor cell apoptosis

Molecular chaperones may promote cell survival, but how this process is regulated, especially in cancer, is not well understood. Using high throughput proteomics screening, we identified the cell cycle regulator and apoptosis inhibitor survivin as a novel protein associated with the molecular chaperone Hsp60. Acute ablation of Hsp60 by small interfering RNA destabilizes the mitochondrial pool of survivin, induces mitochondrial dysfunction, and activates caspase-dependent apoptosis. This response involves disruption of an Hsp60-p53 complex, which results in p53 stabilization, increased expression of pro-apoptotic Bax, and Bax-dependent apoptosis. In vivo, Hsp60 is abundantly expressed in primary human tumors, as compared with matched normal tissues, and small interfering RNA ablation of Hsp60 in normal cells is well tolerated and does not cause apoptosis. Therefore, Hsp60 orchestrates a broad cell survival program centered on stabilization of mitochondrial survivin and restraining of p53 function, and this process is selectively exploited in cancer. Hsp60 inhibitors may function as attractive anticancer agents by differentially inducing apoptosis in tumor cells.     

The in vitro and in vivo apoptotic effects of Mahonia oiwakensis on human lung cancer cells

Both the root and stem bark of Mahonia species were popular folk medicines. The plant has several proven biological activities including anti-bacterial, anti-fungal, and anti-inflammatory effects. However, Mahonia has not been studied for its anticancer effects. In the present study, we made extracts from Mahonia oiwakensis (MOE), a selected species in Taiwan, and investigated their effects on various human lung cells. We found that MOE-induced apoptotic death in human A549 non-small-cell lung carcinoma (NSCLC) cells in a dose- and time-dependent manner. Treatment with the extracts also caused an increase in the sub-G1 fraction of cells, chromosome condensation, and DNA fragmentation. The mitochondrial-mediated pathway was implicated in this MOE-induced apoptosis as evidenced by the activation of the caspase cascade, cleavage of poly (ADP-ribose) polymerase (PARP), disruption of mitochondrial membrane potential, and release of cytochrome C. A higher ratio of Bax/Bcl-2 proteins and cleavage of Bid were also observed in MOE-induced cell apoptosis. In A549 tumor-xenografted nude mice, MOE also retarded in vivo proliferation (P < 0.05) and induced apoptosis in tumor cells, as shown by a decrease in Ki-67-positive staining (P < 0.05) and increased transferase-mediated dUTP nick-end labeling (TUNEL)-positive staining (P < 0.05). In conclusion, MOE inhibits the growth of human lung cancer cells in vitro and in vivo, suggesting that it may have therapeutic potential against human lung cancer.     

p53 Is Required for Cisplatin-induced Processing of the Mitochondrial Fusion Protein L-Opa1 That Is Mediated by the Mitochondrial Metallopeptidase Oma1 in Gynecologic Cancers

Mitochondria are highly dynamic organelles, and mitochondrial fission is a crucial step of apoptosis. Although Oma1 is believed to be responsible for long form Opa1 (L-Opa1) processing during mitochondrial fragmentation, whether and how Oma1 is involved in L-Opa1 processing and participates in the regulation of chemoresistance is unknown. Chemosensitive and chemoresistant ovarian (OVCA) and cervical (CECA) cancer cells were treated with cisplatin (CDDP). Mitochondrial dynamics and protein contents were assessed by immunofluorescence and Western blot, respectively. The requirements of Oma1 and p53 for CDDP-induced L-Opa1 processing, mitochondrial fragmentation, and apoptosis were examined by siRNA or cDNA. CDDP induces L-Opa1 processing and mitochondrial fragmentation in chemosensitive but not in chemoresistant cells. CDDP induced Oma1 40-kDa form increases in OV2008 cells, not in C13* cells. Oma1 knockdown inhibited L-Opa1 processing, mitochondrial fragmentation, and apoptosis. Silencing p53 expression attenuated the effects of CDDP in Oma1 (40 kDa) increase, L-Opa1 processing, mitochondrial fragmentation, and apoptosis in chemosensitive OVCA cells, whereas reconstitution of p53 in p53 mutant or null chemoresistant OVCA cells induced Oma1 (40 kDa) increase, L-Opa1 processing, mitochondrial fragmentation, and apoptosis irrespective of the presence of CDDP. Prohibitin 1 (Phb1) dissociates from Opa1-Phb1 complex and binds phosphorylated p53 (serine 15) in response to CDDP in chemosensitive but not chemoresistant CECA cells. These findings demonstrate that (a) p53 and Oma1 mediate L-Opa1 processing, (b) mitochondrial fragmentation is involved in CDDP-induced apoptosis in OVCA and CECA cells, and (c) dysregulated mitochondrial dynamics may in part be involved in the pathophysiology of CDDP resistance.     

Salicylic acid-mediated potentiation of Hsp70 induction correlates with reduced apoptosis in tobacco protoplasts.

BACKGROUND: Elevated temperatures jeopardize plant disease resistance, as mediated by salicylic acid (SA). SA potentiates heat-induced expression of the 70-kDa heat shock protein (Hsp70) in tomato cells. In mammalian cells, Hsp70 suppresses apoptosis. We hypothesized that potentiation of heat-induced Hsp70 by SA contributes to a reduction in apoptosis in tobacco protoplasts. METHODS: Tobacco protoplasts (Nicotiana tabacum) were exposed to SA (70 microM) at normal temperatures or in combination with heat shock. Hsp70/Hsc70 accumulation and phosphatidylserine (PS) exposure, DNA fragmentation, as well as loss of mitochondrial membrane potential were quantified by flow cytometry. RESULTS AND CONCLUSIONS: SA at normal temperatures did not influence Hsp70/Hsc70 accumulation, but were found to induce apoptosis. In contrast, SA in combination with HS potentiated heat-induced Hsp70/Hsc70 accumulation in tobacco protoplasts that correlated negatively with apoptosis, illustrated by decreased PS exposure and DNA fragmentation and enhanced mitochondrial membrane potential. We propose that this correlation supports a possible role for apoptosis suppression by Hsp70 under elevated temperatures during pathogen infection.     

Crystal structure of the nucleotide‐binding domain of mortalin,the mitochondrial Hsp70 chaperone

Mortalin, a member of the Hsp70‐family of molecular chaperones, functions in a variety of processes including mitochondrial protein import and quality control, Fe‐S cluster protein biogenesis, mitochondrial homeostasis, and regulation of p53. Mortalin is implicated in regulation of apoptosis, cell stress response, neurodegeneration, and cancer and is a target of the antitumor compound MKT‐077. Like other Hsp70‐family members, Mortalin consists of a nucleotide‐binding domain (NBD) and a substrate‐binding domain. We determined the crystal structure of the NBD of human Mortalin at 2.8 Å resolution. Although the Mortalin nucleotide‐binding pocket is highly conserved relative to other Hsp70 family members, we find that its nucleotide affinity is weaker than that of Hsc70. A Parkinson's disease‐associated mutation is located on the Mortalin‐NBD surface and may contribute to Mortalin aggregation. We present structure‐based models for how the Mortalin‐NBD may interact with the nucleotide exchange factor GrpEL1, with p53, and with MKT‐077. Our structure may contribute to the understanding of disease‐associated Mortalin mutations and to improved Mortalin‐targeting antitumor compounds.     

Caveolin-1 knockdown increases the therapeutic sensitivity of lung cancer to cisplatin-induced apoptosis by repressing Parkin-related mitophagy and activating the ROCK1 pathway

Chemotherapy is the first-line treatment option for patients with lung cancer. However, therapeutic resistance occurs through an incompletely understood mechanism. Our research wants to investigate the influence of Caveolin-1 (Cav-1) on the therapeutic sensitivity of lung cancer in vitro. Results in this study demonstrated that Cav-1 levels were markedly inhibited in A549 lung cancer cells after exposure to cisplatin. Knockdown of caveolin further enhanced cisplatin-triggered cancer death in A549 cells. The functional investigation demonstrated that Cav-1 inhibition amplified the mitochondrial stress signaling induced by cisplatin, as evidenced by the mitochondrial reactive oxygen species burst, cellular metabolic disruption, mitochondrial membrane potential reduction, and mitochondrial caspase-9-related apoptosis activation. At the molecular level, cav-1 augmented cisplatin-mediated mitochondrial damage by inhibiting Parkin-related mitochondrial autophagy. Mitophagy activation effectively attenuated the promotive impact of Cav-1 knockdown on mitochondrial damage and cell death. Furthermore, our data indicated that Cav-1 affected Parkin-related mitophagy by activating the Rho-associated coiled-coil kinase 1 (ROCK1) pathway; inhibition of the ROCK1 axis prevented cav-1 knockdown-mediated cell death and mitochondrial damage. Taken together, our results provide ample data illuminate the necessary action exerted by Cav-1 on affecting cisplatin-related therapeutic resistance. Silencing of Cav-1 inhibited Parkin-related mitophagy, thus amplifying cisplatin-mediated mitochondrial apoptotic signaling. This finding identifies the Cav-1/ROCK1/Parkin/mitophagy axis as a potential target to overcome cisplatin-related resistance in lung cancer cells.     

Geldanamycin-Induced Osteosarcoma Cell Death Is Associated with Hyperacetylation and Loss of Mitochondrial Pool of Heat Shock Protein 60 (Hsp60)

Osteosarcoma is one of the most malignant tumors of childhood and adolescence that is often resistant to standard chemo- and radio-therapy. Geldanamycin and geldanamycin analogs have been recently studied as potential anticancer agents for osteosarcoma treatment. Here, for the first time, we have presented novel anticancer mechanisms of geldanamycin biological activity. Moreover, we demonstrated an association between the effects of geldanamycin on the major heat shock proteins (HSPs) and the overall survival of highly metastatic human osteosarcoma 143B cells. We demonstrated that the treatment of 143B cells with geldanamycin caused a subsequent upregulation of cytoplasmic Hsp90 and Hsp70 whose activity is at least partly responsible for cancer development and drug resistance. On the other hand, geldanamycin induced upregulation of Hsp60 gene expression, and a simultaneous loss of hyperacetylated Hsp60 mitochondrial protein pool resulting in decreased viability and augmented cancer cell death. Hyperacetylation of Hsp60 seems to be associated with anticancer activity of geldanamycin. In light of the fact that mitochondrial dysfunction plays a critical role in the apoptotic signaling pathway, the presented data may support a hypothesis that Hsp60 can be another functional part of mitochondria-related acetylome being a potential target for developing novel anticancer strategies.     

Hsp70 exerts its anti-apoptotic function downstream of caspase-3-like proteases.

The major heat shock protein, Hsp70, is an effective inhibitor of apoptosis. To study its mechanism of action, we created tumor cell lines with altered Hsp70 levels. The expression levels of Hsp70 in the cells obtained correlated well with their survival following treatments with tumor necrosis factor, staurosporine and doxorubicin. Surprisingly, the surviving Hsp70-expressing cells responded to the apoptotic stimuli by activation of stress-activated protein kinases, generation of free radicals, early disruption of mitochondrial transmembrane potential, release of cytochrome c from mitochondria and activation of caspase-3-like proteases in a manner essentially similar to that of the dying cells with low Hsp70 levels. However, Hsp70 inhibited late caspase-dependent events such as activation of cytosolic phospholipase A2 and changes in nuclear morphology. Furthermore, Hsp70 conferred significant protection against cell death induced by enforced expression of caspase-3. Thus, Hsp70 rescues cells from apoptosis later in the death signaling pathway than any known anti-apoptotic protein, making it a tempting target for therapeutic interventions.