Depletion of the oncoprotein Bcl-3 induces centrosome amplification and aneuploidy in cancer cells

Bcl-3 is an atypical member of the inhibitor of NF-kappa B family of proteins since it can function as a coactivator of transcription. Although this oncogene was described in leukemia, it is overexpressed in a number of solid tumors as well. The oncogenic potential of Bcl-3 has been associated with its capacity to increase proliferation by means of activating the cyclin D1 promoter and to its antiapoptotic role mediated by the inhibiton of p53 activity. In the course of dissecting these properties, we found that depleting Bcl-3 protein using shRNAs induce a decrease of proliferation and clonogenic survival associated with the induction of multinucleation and increased ploidy. These effects were associated with a DNA damage response, a delay in G2/M checkpoint and the induction of centrosome amplificatio

Role of Smac/DIABLO in cancer progression

Second mitochondria-derived activator of caspase/direct inhibitor of apoptosis-binding protein with low pI (Smac/DIABLO) is a proapoptogenic mitochondrial protein that is released to the cytosol in response to diverse apoptotic stimuli, including commonly used chemotherapeutic drugs. In the cytosol, Smac/DIABLO interacts and antagonizes inhibitors of apoptosis proteins (IAPs), thus allowing the activation of caspases and apoptosis. This activity has prompted the synthesis of peptidomimetics that could potentially be used in cancer therapy. For these reasons, several authors have analyzed the expression levels of Smac/DIABLO in samples of patients from different tumors. Although dissimilar results have been found, a tissue-specific role of this protein emerges from the data. The objective of this review is to present the current knowledge of the Smac/DIABLO role in cancer and its possible use as a marker or therapeutic target for drug design

Survivin isoform Delta Ex3 regulates tumor spheroid formation

Survivin is an important member of the Inhibitor of Apoptosis Proteins (IAPs) family and has essential roles in apoptosis and cell cycle progression. This gene is commonly upregulated in human cancer and provides an exciting diagnostic and therapeutic target. Survivin is expressed as several isoforms that are generated by alternative splicing, and some of these present antagonistic activities. Currently, information regarding the regulation of these isoforms is lacking. In this study, we sought to analyze survivin Delta Ex3 expression in a three-dimensional model of avascular tumors and its overexpression effects in processes such as proliferation, clonogenicity and apoptosis. We found a positive correlation between spheroid growth and survivin Delta Ex3 expression during the exponential phase. We demonstrated that this isoform not only decreased apoptosis but also inhibited tumor spheroid formation by decreasing proliferation and clonogenic survival. These results point toward a dual and antagonistic effect of this spliced survivin isoform in cancer development

Tissue Inhibitor of Metalloproteinase-4 Triggers Apoptosis in Cervical Cancer Cells

<div><p>Tissue inhibitor of metalloproteinase-4 (TIMP-4) is a member of extracellular matrix (ECM) metalloproteinases inhibitors that has pleiotropic functions. However, TIMP-4 roles in carcinogenesis are not well understood. Cell viability and flow cytometer assays were employed to evaluate cell death differences between H-Vector and H-TIMP-4 cell lines. Immunobloting and semi-quantitative RT-PCR were used to evaluate the expression of apoptosis regulators. We showed that TIMP-4 has apoptosis-sensitizing effects towards several death stimuli. Consistent with these findings, regulators of apoptosis from Inhibitors of Apoptosis Proteins (IAP), FLICE-like inhibitor proteins (FLIP) and Bcl-2 family members were modulated by TIMP-4. In addition, TIMP-4 knockdown resulted in cell survival increase after serum deprivation, as assessed by clonogenic cell analyses. This report shows that TIMP-4 regulates carcinogenesis through apoptosis activation in cervical cancer cells. Understanding TIMP-4 effects in tumorigenesis may provide clues for future therapies.</p></div

Analysis of apoptosis genes in HeLa cells.

<p>A, Accumulation of FLIP isoforms’ mRNA was analyzed by RT-PCR in HeLa cells treated with hrTIMP-4 (10 nM, upper panels) during the indicated time. FLIP protein expression was determined by immunoblotting in H-Vector and H-TIMP-4 cell lines (lower panels). B, cIAP-1, cIAP-2, and survivin mRNA presence were examined by semi-quantitative RT-PCR in H-Vector and H-TIMP-4 cell lines.</p

TIMP-4 overexpression decreases cell viability.

<p>A, TIMP-4 expression analysis by RT-PCR in H-Vector and H-TIMP-4 cells. B, The graph shows the H-Vector and H-TIMP-4 cell viability percentages following growth with FBS (8%) or without FBS (0%) for 5 days. C, The graph displays the H-Vector and H-TIMP-4 cell viability percentages after TNF-α (20 ng/mL, 48 h) and TRAIL (20 ng/mL, 7 h) treatment. D, The graph displays HeLa cell viability analyzed after hrTIMP-4 (10 nM, 48 h) and TRAIL (20 ng/mL, 7 h) exposure. E, The graph shows the HeLa cells viability after a 48 h preincubation period with hrTIMP-4 (10 nM) followed by TNF-α (20 ng/mL, 48 h) stimulation. Asterisks indicate significant differences with values of p = 0.03 in A, p = 0.006 in C and p = 0.003 in D and E.</p

TIMP-4 up-regulation potentiates Annexin-V translocation.

<p>H-Vector and H-TIMP-4 cells were FBS starved (5 days) or treated with TNF-α (20 ng/mL, 48 h,) and analyzed for outer plasma membrane phosphatidylserine presence by flow cytometry. Early, late and total proportion of apoptotic cells is presented. Results represent the average of three independent experiments. H-Vector apoptotic cells were considered as “1”.</p><p>TIMP-4 up-regulation potentiates Annexin-V translocation.</p

Effects of TIMP-4 on cell growth and cell viability.

<p>Effects of TIMP-4 on cell growth and cell viability.</p

TIMP-4 induces cell death in vitro.

<p>A, TIMP-4 modulates TNFRI, TNFRII and DISC components TRAF2 and TRADD. Proteins were detected by immunoblot in H-Vector and H-TIMP-4 cells protein extracts.</p