Endothelial apoptotic activity of angiocidin is dependent on its polyubiquitin binding activity

We recently cloned the full-length cDNA of a tumour-associated protein. The recombinant protein expressed in bacteria and referred to as angiocidin has potent antitumour activity in vivo and in vitro. Angiocidin inhibits tumour growth and angiogenesis by inducing apoptosis in endothelial cells. Based on the sequence similarity of angiocidin to S5a, one of the major polyubiquitin recognition proteins in eukaryotic cells, we postulated that the antiendothelial activity of angiocidin could be due in part to its polyubiquitin binding activity. In support of this hypothesis, we show that angiocidin binds polyubiquitin in vivo with high affinity and colocalises with ubiquitinated proteins on the surface of endothelial cells. Binding is blocked with an antiubiquitin antibody. Angiocidin treatment of endothelial cells transfected with a proteasome fluorescent reporter protein showed a dose-dependent inhibition of proteasome activity and accumulation of polyubiquitinated proteins. Full-length angiocidin bound polyubiquitin while three angiocidin recombinant proteins whose putative polyubiquitin binding sites were mutated either failed to bind polyubiquitin or had significantly diminished binding activity. The in vitro apoptotic activity of these mutants correlated with their polyubiquitin binding activity. These data strongly argue that the apoptotic activity of angiocidin is dependent on its polyubiquitin binding activity

The role of the ubiquitination-proteasome pathway in breast cancer: Applying drugs that affect the ubiquitin-proteasome pathway to the therapy of breast cancer

The ubiquitin-proteasome pathway is responsible for most eukaryotic intracellular protein degradation. This pathway has been validated as a target for antineoplastic therapy using both in vitro and preclinical models of human malignancies, and is influenced as part of the mechanism of action of certain chemotherapeutic agents. Drugs whose primary action involves modulation of ubiquitin-proteasome activity, most notably the proteasome inhibitor PS-341, are currently being evaluated in clinical trials, and have already been found to have significant antitumor efficacy. On the basis of the known mechanisms by which these agents work, and the available clinical data, they would seem to be well suited for the treatment of breast neoplasms. Such drugs, alone and especially in combination with current chemotherapeutics, may well represent important advances in the therapy of patients with breast cancer

Activation and modulation of antiviral and apoptotic genes in pigs infected with classical swine fever viruses of high, moderate or low virulence

The immune response to CSFV and the strategies of this virus to evade and suppress the pigs’ immune system are still poorly understood. Therefore, we investigated the transcriptional response in the tonsils, median retropharyngeal lymph node (MRLN), and spleen of pigs infected with CSFV strains of similar origin with high, moderate, and low virulence. Using a porcine spleen/intestinal cDNA microarray, expression levels in RNA pools prepared from infected tissue at 3 dpi (three pigs per virus strain) were compared to levels in pools prepared from uninfected homologue tissues (nine pigs). A total of 44 genes were found to be differentially expressed. The genes were functionally clustered in six groups: innate and adaptive immune response, interferon-regulated genes, apoptosis, ubiquitin-mediated proteolysis, oxidative phosphorylation and cytoskeleton. Significant up-regulation of three IFN-γ-induced genes in the MRLNs of pigs infected with the low virulence strain was the only clear qualitative difference in gene expression observed between the strains with high, moderate and low virulence. Real-time PCR analysis of four response genes in all individual samples largely confirmed the microarray data at 3 dpi. Additional PCR analysis of infected tonsil, MRLN, and spleen samples collected at 7 and 10 dpi indicated that the strong induction of expression of the antiviral response genes chemokine CXCL10 and 2′–5′ oligoadenylate synthetase 2, and of the TNF-related apoptosis-inducing ligand (TRAIL) gene at 3 dpi, decreased to lower levels at 7 and 10 dpi. For the highly and moderately virulent strains, this decrease in antiviral and apoptotic gene expression coincided with higher levels of virus in these immune tissues

Turnover of BRCA1 Involves in Radiation-Induced Apoptosis

Background: Germ-line mutations of the breast cancer susceptibility gene-1 (BRCA1) increase the susceptibility to tumorigenesis. The function of BRCA1 is to regulate critical cellular processes, including cell cycle progression, genomic integrity, and apoptosis. Studies on the regulation of BRCA1 have focused intensely on transcription and phosphorylation mechanisms. Proteolytic regulation of BRCA1 in response to stress signaling remains largely unknown. The manuscript identified a novel mechanism by which BRCA1 is regulated by the ubiquitin-dependent degradation in response to ionization. Methodology/Principal Findings: Here, we report that severe ionization triggers rapid degradation of BRCA1, which in turn results in the activation of apoptosis. Ionization-induced BRCA1 turnover is mediated via an ubiquitin-proteasomal pathway. The stabilization of BRCA1 significantly delays the onset of ionization-induced apoptosis. We have mapped the essential region on BRCA1, which mediates its proteolysis in response to ionization. Moreover, we have demonstrated that BRCA1 protein is most sensitive to degradation when ionization occurs during G2/M and S phase. Conclusions/Significance: Our results suggest that ubiquitin-proteasome plays an important role in regulating BRCA1 during genotoxic stress. Proteolytic regulation of BRCA1 involves in ionization-induced apoptosis. © 2010 Liu et al

The Antidiabetic Drug Ciglitazone Induces High Grade Bladder Cancer Cells Apoptosis through the Up-Regulation of TRAIL

International audienceBACKGROUND: Ciglitazone belongs to the thiazolidinediones class of antidiabetic drug family and is a high-affinity ligand for the Peroxisome Proliferator-Activated Receptor γ (PPARγ). Apart from its antidiabetic activity, this molecule shows antineoplastic effectiveness in numerous cancer cell lines. METHODOLOGY/PRINCIPAL FINDINGS: Using RT4 (derived from a well differentiated grade I papillary tumor) and T24 (derived from an undifferentiated grade III carcinoma) bladder cancer cells, we investigated the potential of ciglitazone to induce apoptotic cell death and characterized the molecular mechanisms involved. In RT4 cells, the drug induced G2/M cell cycle arrest characterized by an overexpression of p53, p21(waf1/CIP1) and p27(Kip1) in concomitance with a decrease of cyclin B1. On the contrary, in T24 cells, it triggered apoptosis via extrinsic and intrinsic pathways. Cell cycle arrest and induction of apoptosis occurred at high concentrations through PPARγ activation-independent pathways. We show that in vivo treatment of nude mice by ciglitazone inhibits high grade bladder cancer xenograft development. We identified a novel mechanism by which ciglitazone kills cancer cells. Ciglitazone up-regulated soluble and membrane-bound TRAIL and let TRAIL-resistant T24 cells to respond to TRAIL through caspase activation, death receptor signalling pathway and Bid cleavage. We provided evidence that TRAIL-induced apoptosis is partially driven by ciglitazone-mediated down-regulation of c-FLIP and survivin protein levels through a proteasome-dependent degradation mechanism. CONCLUSIONS/SIGNIFICANCE: Therefore, ciglitazone could be clinically relevant as chemopreventive or therapeutic agent for the treatment of TRAIL-refractory high grade urothelial cancers

Caspase-2 Mediated Apoptotic and Necrotic Murine Macrophage Cell Death Induced by Rough Brucella abortus

Brucella species are Gram-negative, facultative intracellular bacteria that cause zoonotic brucellosis. Survival and replication inside macrophages is critical for establishment of chronic Brucella infection. Virulent smooth B. abortus strain 2308 inhibits programmed macrophage cell death and replicates inside macrophages. Cattle B. abortus vaccine strain RB51 is an attenuated rough, lipopolysaccharide O antigen-deficient mutant derived from smooth strain 2308. B. abortus rough mutant RA1 contains a single wboA gene mutation in strain 2308. Our studies demonstrated that live RB51 and RA1, but not strain 2308 or heat-killed Brucella, induced both apoptotic and necrotic cell death in murine RAW264.7 macrophages and bone marrow derived macrophages. The same phenomenon was also observed in primary mouse peritoneal macrophages from mice immunized intraperitoneally with vaccine strain RB51 using the same dose as regularly performed in protection studies. Programmed macrophage cell death induced by RB51 and RA1 was inhibited by a caspase-2 inhibitor (Z-VDVAD-FMK). Caspase-2 enzyme activation and cleavage were observed at the early infection stage in macrophages infected with RB51 and RA1 but not strain 2308. The inhibition of macrophage cell death promoted the survival of rough Brucella cells inside macrophages. The critical role of caspase-2 in mediating rough B. abortus induced macrophage cell death was confirmed using caspase-2 specific shRNA. The mitochondrial apoptosis pathway was activated in macrophages infected with rough B. abortus as demonstrated by increase in mitochondrial membrane permeability and the release of cytochrome c to cytoplasm in macrophages infected with rough Brucella. These results demonstrate that rough B. abortus strains RB51 and RA1 induce apoptotic and necrotic murine macrophage cell death that is mediated by caspase-2. The biological relevance of Brucella O antigen and caspase-2-mediated macrophage cell death in Brucella pathogenesis and protective Brucella immunity is discussed

Hippocampal Atrophy as a Quantitative Trait in a Genome-Wide Association Study Identifying Novel Susceptibility Genes for Alzheimer's Disease

With the exception of APOE ε4 allele, the common genetic risk factors for sporadic Alzheimer's Disease (AD) are unknown., which can be considered potential “new” candidate loci to explore in the etiology of sporadic AD. These candidates included EFNA5, CAND1, MAGI2, ARSB, and PRUNE2, genes involved in the regulation of protein degradation, apoptosis, neuronal loss and neurodevelopment. Thus, we identified common genetic variants associated with the increased risk of developing AD in the ADNI cohort, and present publicly available genome-wide data. Supportive evidence based on case-control studies and biological plausibility by gene annotation is provided. Currently no available sample with both imaging and genetic data is available for replication.Using hippocampal atrophy as a quantitative phenotype in a genome-wide scan, we have identified candidate risk genes for sporadic Alzheimer's disease that merit further investigation

Nat. Rev. Mol. Cell Biol.

The ubiquitin/proteasome pathway is the main non-lysosomal route for intracellular protein degradation in eukaryotes, It is instrumental to various cellular processes, such as cell- cycle progression, transcription and antigen processing. Recent findings also substantiate a pivotal role of the ubiquitin/proteasome pathway in the regulation of apoptosis. Regulatory molecules that are involved in programmed cell death have been identified as substrates of the proteasome. Moreover, key regulators of apoptosis themselves seem to have an active part in the proteolytic inactivation of death executors