Induction in myeloid leukemic cells of genes that are expressed in different normal tissues

Using DNA microarray and cluster analysis of expressed genes in a cloned line (M1-t-p53) of myeloid leukemic cells, we have analyzed the expression of genes that are preferentially expressed in different normal tissues. Clustering of 547 highly expressed genes in these leukemic cells showed 38 genes preferentially expressed in normal hematopoietic tissues and 122 other genes preferentially expressed in different normal non-hematopoietic tissues including neuronal tissues, muscle, liver and testis. We have also analyzed the genes whose expression in the leukemic cells changed after activation of wild-type p53 and treatment with the cytokine interleukin 6 (IL-6) or the calcium mobilizer thapsigargin (TG). Out of 620 such genes in the leukemic cells that were differentially expressed in normal tissues, clustering showed 80 genes that were preferentially expressed in hematopoietic tissues and 132 genes in different normal non-hematopietic tissues that also included neuronal tissues, muscle, liver and testis. Activation of p53 and treatment with IL-6 or TG induced different changes in the genes preferentially expressed in these normal tissues. These myeloid leukemic cells thus express genes that are expressed in normal non-hematopoietic tissues, and various treatments can reprogram these cells to induce other such non-hematopoietic genes. The results indicate that these leukemic cells share with normal hematopoietic stem cells the plasticity of differentiation to different cell types. It is suggested that this reprogramming to induce in malignant cells genes that are expressed in different normal tissues may be of clinical value in therapy

Proteolytic cleavage of p53 mutants in response to mismatched DNA

Interaction of p53 with mismatched DNA induces proteolytic cleavage with release of a 35-kDa protein fragment from the p53–DNA complexes. The 35-kDa cleavage product is activated for specific biochemical function(s) and may play a role in the cellular response to DNA damage (Molinari et al (1996) Oncogene13: 2077–2086; Okorokov et al (1997) EMBO J16: 6008–6017). In the present study we have asked if mutants of p53 retain the ability to undergo similar proteolytic cleavage, and compared sequence-specific ‘DNA contact’ with ‘structural’ mutants commonly found in human cancer. In addition, a series of phosphorylation site mutants were generated to investigate the possible effects of phosphorylation/dephosphorylation on the proteolytic cleavage of p53. All mutants tested bound to a mismatched DNA target in vitro. Moreover, studies in vitro and in vivo indicate that p53 mutants with intact conformational structure (as determined by immunoreactivity with PAb246 and PAb1620) retain the ability to undergo proteolytic cleavage similar, if not identical, to the wild-type p53 protein. Our results suggest that the capacity for p53 to bind mismatched DNA is independent of structural conformation of the central core domain. Proteolytic cleavage, however, is crucially dependent upon a wild-type conformation of the protein. © 1999 Cancer Research Campaig

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

Lysosomes in iron metabolism, ageing and apoptosis

The lysosomal compartment is essential for a variety of cellular functions, including the normal turnover of most long-lived proteins and all organelles. The compartment consists of numerous acidic vesicles (pH ∼4 to 5) that constantly fuse and divide. It receives a large number of hydrolases (∼50) from the trans-Golgi network, and substrates from both the cells’ outside (heterophagy) and inside (autophagy). Many macromolecules contain iron that gives rise to an iron-rich environment in lysosomes that recently have degraded such macromolecules. Iron-rich lysosomes are sensitive to oxidative stress, while ‘resting’ lysosomes, which have not recently participated in autophagic events, are not. The magnitude of oxidative stress determines the degree of lysosomal destabilization and, consequently, whether arrested growth, reparative autophagy, apoptosis, or necrosis will follow. Heterophagy is the first step in the process by which immunocompetent cells modify antigens and produce antibodies, while exocytosis of lysosomal enzymes may promote tumor invasion, angiogenesis, and metastasis. Apart from being an essential turnover process, autophagy is also a mechanism by which cells will be able to sustain temporary starvation and rid themselves of intracellular organisms that have invaded, although some pathogens have evolved mechanisms to prevent their destruction. Mutated lysosomal enzymes are the underlying cause of a number of lysosomal storage diseases involving the accumulation of materials that would be the substrate for the corresponding hydrolases, were they not defective. The normal, low-level diffusion of hydrogen peroxide into iron-rich lysosomes causes the slow formation of lipofuscin in long-lived postmitotic cells, where it occupies a substantial part of the lysosomal compartment at the end of the life span. This seems to result in the diversion of newly produced lysosomal enzymes away from autophagosomes, leading to the accumulation of malfunctioning mitochondria and proteins with consequent cellular dysfunction. If autophagy were a perfect turnover process, postmitotic ageing and several age-related neurodegenerative diseases would, perhaps, not take place