Caspase-2 function in response to DNA damage.

Caspase-2 is one of the best conserved caspases across species. This enzyme is unique among caspases in that it has features of both initiator and effector caspases. Caspase-2 appears to be necessary for the onset of apoptosis triggered by several insults, including DNA damage, administration of TNF, and different pathogens and viruses.

PKC 412 sensitizes U1810 non-small cell lung cancer cells to DNA damage.

Non-small cell lung carcinoma (NSCLC) is characterized by resistance to drug-induced apoptosis, which might explain the survival of lung cancer cells following treatment. Recently we have shown that the broad-range kinase inhibitor staurosporine (STS) reactivates the apoptotic machinery in U1810 NSCLC cells [Joseph et al., Oncogene 21 (2002) 65].

Aberrant mitochondrial iron distribution and maturation arrest characterize early erythroid precursors in low-risk myelodysplastic syndromes.

Early erythroblasts from patients with refractory anemia (RA) and RA with ringed sideroblasts (RARS) show constitutive mitochondrial release of cytochrome c. Moreover, mature erythroblasts in RARS, but not in RA, display aberrant accumulation of mitochondrial ferritin (MtF). We analyzed cytochrome c release, MtF expression, and gene expression during erythroid differentiation in bone marrow cells from myelodysplastic syndrome (MDS) patients and healthy controls.

The HPV-16 E7 oncogene sensitizes malignant cells to IFN-alpha-induced apoptosis.

Interferons (IFNs) exert antitumor effects in several human malignancies, but their mechanism of action is unclear. There is a great variability in sensitivity to IFN treatment depending on both tumor type and the individual patient. The reason for this variable sensitivity is not known.

Caspase-2 permeabilizes the outer mitochondrial membrane and disrupts the binding of cytochrome c to anionic phospholipids.

Caspases are cysteine proteases that play a central role in the execution of apoptosis. Recent evidence indicates that caspase-2 is activated early in response to genotoxic stress and can function as an upstream modulator of the mitochondrial apoptotic pathway. In particular, we have shown previously that fully processed caspase-2 can permeabilize the outer mitochondrial membrane and cause cytochrome c and Smac/DIABLO release from these organelles.

Apoptosis and genomic instability.

Genomic instability is intrinsically linked to significant alterations in apoptosis control. Chromosomal and microsatellite instability can cause the inactivation of pro-apoptotic pathways. In addition, the inhibition of apoptosis itself can be permissive for the survival and ongoing division of cells that have failed to repair DNA double-strand breaks, experience telomere dysfunction or are in an abnormal polyploid state.

Downregulation of peroxiredoxin V stimulates formation of etoposide-induced double-strand DNA breaks.

Antioxidant protein Peroxiredoxin V (PrxV) is located in mitochondria and peroxisomes but is also present in the nucleus. Here, we show that nuclear PrxV associates with coilin-containing bodies suggesting possible interaction of this protein with transcription complexes. We also studied etoposide-induced phosphorylation of histone H2AX (gamma-H2AX) in human cells in which PrxV activity was downregulated (knockdown, KD-clones) or compromised by overexpression of redox-negative (RD) protein.

Apoptosis-inducing factor determines the chemoresistance of non-small-cell lung carcinomas.

Non-small-cell lung carcinomas (NSCLCs) are resistant to the induction of apoptosis by conventional anticancer treatment. However, NSCLC cell lines are sensitive to the action of the broad protein kinase inhibitor, staurosporine (STS). In the NSCLC cell line U1810, STS induced the mitochondrial release of apoptosis-inducing factor (AIF) and cytochrome c (Cyt c) followed by activation of caspases, nuclear condensation, DNA fragmentation and finally cell death.

Endogenously released Smac is insufficient to mediate cell death of human lung carcinoma in response to etoposide.

Cytotoxic agents eliminate tumor cells via different mechanisms including apoptosis, although this process is not equally efficient in all kinds of cancer cells. Thus, small cell lung carcinomas (SCLCs) are more sensitive than non-small cell lung carcinomas (NSCLCs) to therapy-induced killing. During apoptosis, several apoptogenic proteins release from the mitochondria.

Processed caspase-2 can induce mitochondria-mediated apoptosis independently of its enzymatic activity.

The mechanism by which caspase-2 executes apoptosis remains obscure. Recent findings indicate that caspase-2 is activated early in response to DNA-damaging antineoplastic agents and may be important for the engagement of the mitochondrial apoptotic pathway. We demonstrate here that fully processed caspase-2 stimulates mitochondrial release of cytochrome c and Smac/DIABLO, but not apoptosis-inducing factor (AIF).

Endothelial cell surface ATP synthase-triggered caspase-apoptotic pathway is essential for k1-5-induced antiangiogenesis.

We have recently reported the identification of kringle 1-5 (K1-5) of plasminogen as a potent and specific inhibitor of angiogenesis and tumor growth. Here, we show that K1-5 bound to endothelial cell surface ATP synthase and triggered caspase-mediated endothelial cell apoptosis. Induction of endothelial apoptosis involved sequential activation of caspases-8, -9, and -3.

Analysis of mitochondrial dysfunction during cell death.

Attempts to identify a common underlying step in the apoptotic program in response to various cytotoxic stimuli have focused on the role of mitochondria in this form of cell death. This unit contains a family of protocols that can be used to assess mitochondrial functions during apoptotic responses. Protocols are included for the collection and analysis of released proteins, for detection of the mitochondrial permeability transition, for measurement of mitochondrial membrane potential, and for preparation of mitochondria from different tissue sources.

DNA damage-induced apoptosis.

Unicellular organisms respond to the presence of DNA lesions by activating cell cycle checkpoint and repair mechanisms, while multicellular animals have acquired the further option of eliminating damaged cells by triggering apoptosis. Defects in DNA damage-induced apoptosis contribute to tumorigenesis and to the resistance of cancer cells to a variety of therapeutic agents. The intranuclear mechanisms that signal apoptosis after DNA damage overlap with those that initiate cell cycle arrest and DNA repair, and the early events in these pathways are highly conserved.

Interferon alpha-induced apoptosis in tumor cells is mediated through the phosphoinositide 3-kinase/mammalian target of rapamycin signaling pathway.

Interferon (IFN) alpha induces a caspase-dependent apoptosis that is associated with activation of the proapoptotic Bak and Bax, loss of mitochondrial membrane potential, and release of cytochrome c. In addition to the onset of the classical Jak-STAT pathway, IFNalpha also induced phosphoinositide 3-kinase (PI3K) activity. Pharmacological inhibition of PI3K activity by Ly294002 disrupted IFN-induced apoptosis upstream of mitochondria.

The transcriptosomal response of human A549 lung cells to a hydrogen peroxide-generating system: relationship to DNA damage, cell cycle arrest, and caspase activation.

Intracellular oxidative stress is a dynamic situation characterized by the accumulation of reactive oxygen metabolites, such as hydrogen peroxide. This is traditionally associated with both macromolecular damage and adaptive changes in gene expression, aimed at preventing cellular demise. However, the overall extent of such genetic changes is not well characterized.

Adiponectin-induced antiangiogenesis and antitumor activity involve caspase-mediated endothelial cell apoptosis.

Obesity is a risk factor for the development of many severe human diseases such as cardiovascular disorders, diabetes, and cancer, which are tightly linked to angiogenesis. The adipose tissue produces several growth factors/hormones including leptin, tumor necrosis factor alpha, and adiponectin. It has been found that adiponectin levels are reduced in obesity.

RasGTPase-activating protein is a target of caspases in spontaneous apoptosis of lung carcinoma cells and in response to etoposide.

p120 RasGTPase-activating protein (RasGAP), the main regulator of Ras GTPase family members, is cleaved at low caspase activity into an N-terminal fragment that triggers potent anti-apoptotic signals via activation of the Ras/PI-3 kinase/Akt pathway. When caspase activity is increased, RasGAP fragment N is further processed into two fragments that effectively potentiate apoptosis. Expression of RasGAP protein and its cleavage was assessed in human lung cancer cells with different histology and responsiveness to anticancer drug-induced apoptosis.

Heat shock protein 72 does not modulate ionizing radiation-induced apoptosis in U1810 non-small cell lung carcinoma cells.

Heat shock proteins (HSP) have been shown to interfere with apoptosis signaling, suggesting that there might be a role for these proteins as mediators of resistance to ionizing radiation (IR)-induced apoptosis. Protein expression of the stress inducible heat shock proteins, HSP72 and HSP27, was analyzed in a panel of lung carcinoma cell lines displaying various degrees of radiosensitivity. Expression of HSP72 was high in all cell lines investigated while HSP27 was present in all non-small cell lung carcinoma (NSCLC) and 6/9 small cell lung carcinoma (SCLC) cell lines.

Mitochondrial cytochrome c release may occur by volume-dependent mechanisms not involving permeability transition.

The mechanisms regulating mitochondrial outer-membrane permeabilization and the release of cytochrome c during apoptosis remain controversial. In the present study, we show in an in vitro model system that the release of cytochrome c may occur via moderate modulation of mitochondrial volume, irrespective of the mechanism leading to the mitochondrial swelling. In contrast with mitochondrial permeability transition-dependent release of cytochrome c, in the present study mitochondria remain intact and functionally active.

Caspases: the enzymes of death.

The caspases are a unique family of cysteine proteases, which cleave proteins next to an aspartate residue. Among all known mammalian proteases, only the serine protease granzyme B has similar substrate specificity. In addition to a central role of caspases in the initiation and execution phases of apoptosis, these enzymes have some other non-apoptotic functions in living cells.

VEIDase is a principal caspase-like activity involved in plant programmed cell death and essential for embryonic pattern formation.

Plant embryogenesis is intimately associated with programmed cell death. The mechanisms of initiation and control of programmed cell death during plant embryo development are not known. Proteolytic activity associated with caspase-like proteins is paramount for control of programmed cell death in animals and yeasts.