Role of mitochondrial network stabilisation by a human small heat shock protein in tumour malignancy.

Previously, we found that the unconventional small human heat-shock protein HSPB11 inhibits cell death by HSP90 mediated cholesterol-rich membrane microdomain dependent activation of phosphatidylinositol-3 kinase/protein kinase B pathway and by stabilising the mitochondrial membrane systems. Also, progressive cytoplasmic expression of HSPB11 correlated with brain tumor malignancy. In the present study we investigated how cytoplasmic abundance of HSPB11 augments tumor malignancy. We up- and downregulated the cytoplasmic level of HSPB11 before paclitaxel exposure in NIH3T3 and HeLa cells, which normally express low and high levels, respectively, of the HSPB11 protein. We examined the paclitaxel-mediated induction of cell death, mitochondrial fission, HSPB11 mitochondrial translocation and inhibitory phosphorylation of dynamin-like protein-1 (DLP1). We found that increasing cytoplasmic abundance of HSPB11 in NIH3T3 cells protected against paclitaxel-induced apoptosis, while suppressing HSPB11 in HeLa cells sensitised the cells toward paclitaxel. Also, paclitaxel enhanced mitochondrial translocation of HSPB11 in wild type HeLa but not in NIH3T3 cells. More importantly, increased cytoplasmic level of HSPB11 in NIH3T3 cells enhanced the inhibitory phosphorylation of DLP1 and attenuated paclitaxel-induced mitochondrial fission. All these results suggest that increased cytoplasmic abundance of HSPB11 augments inhibitory phosphorylation of DLP1 thereby reduces mitochondrial fission that eventually leads to decreased apoptosis. This novel mechanism may explain the resistance to apoptosis and increased malignancy of HSPB11-overexpressing tumours. The clinical significance of this mechanism has already been highlighted by the finding that the kinase inhibitor tyrphostin A9 induces cancer cell death by DLP1-mediated mitochondrial fragmentation.

Protective effect of the poly(ADP-ribose) polymerase inhibitor PJ34 on mitochondrial depolarization-mediated cell death in hepatocellular carcinoma cells involves attenuation of c-Jun N-terminal kinase-2 and protein kinase B/Akt activation.

BACKGROUND: 2,4-Dimethoxyphenyl-E-4-arylidene-3-isochromanone (IK11) was previously described to induce apoptotic death of A431 tumor cells. In this report, we investigated the molecular action of IK11 in the HepG2 human hepatocellular carcinoma cell line to increase our knowledge of the role of poly (ADP-ribose)-polymerase (PARP), protein kinase B/Akt and mitogen activated protein kinase (MAPK) activation in the survival and death of tumor cells and to highlight the possible role of PARP-inhibitors in co-treatments with different cytotoxic agents in cancer therapy. RESULTS: We found that sublethal concentrations of IK11 prevented proliferation, migration and entry of the cells into their G2 phase. At higher concentrations, IK11 induced reactive oxygen species (ROS) production, mitochondrial membrane depolarization, activation of c-Jun N-terminal kinase 2 (JNK2), and substantial loss of HepG2 cells. ROS production appeared marginal in mediating the cytotoxicity of IK11 since N-acetyl cysteine was unable to prevent it. However, the PARP inhibitor PJ34, although not a ROS scavenger, strongly inhibited both IK11-induced ROS production and cell death. JNK2 activation seemed to be a major mediator of the effect of IK11 since inhibition of JNK resulted in a substantial cytoprotection while inhibitors of the other kinases failed to do so. Inhibition of Akt slightly diminished the effect of IK11, while the JNK and Akt inhibitor and ROS scavenger trans-resveratrol completely protected against it. CONCLUSIONS: These results indicate significant involvement of PARP, a marginal role of ROS and a pro-apoptotic role of Akt in this system, and raise attention to a novel mechanism that should be considered when cancer therapy is augmented with PARP-inhibition, namely the cytoprotection by inhibition of JNK2.

Induction of mitochondrial destabilization and necrotic cell death by apolar mitochondria-directed SOD mimetics.

In this paper, we present evidence, for the first time, that increasing the lipophilicity of mitochondria targeting SOD mimetics reverses their cytoprotective properties, destabilizing the mitochondrial membrane system and promoting cell death. A new mitochondria-directed apolar SOD mimetic, HO-3814, was found to provoke mitochondrial swelling and loss of mitochondrial membrane potential, and these effects were not inhibited by cyclosporine A. HO-3814-induced cell death was predominantly necrotic, caspase-independent, and not affected by mitochondrial permeability transition inhibitors or cyclophilin D-suppression, inhibitors of mitogen-activated protein kinases or Akt, or various antioxidants. In contrast, Bcl-2 overexpression diminished the effects of HO-3814.