50 research outputs found
The Naturally Processed CD95L Elicits a c-Yes/Calcium/PI3K-Driven Cell Migration Pathway
Patients affected by chronic inflammatory disorders display high amounts of soluble CD95L. This homotrimeric ligand arises from the cleavage by metalloproteases of its membrane-bound counterpart, a strong apoptotic inducer. In contrast, the naturally processed CD95L is viewed as an apoptotic antagonist competing with its membrane counterpart for binding to CD95. Recent reports pinpointed that activation of CD95 may attract myeloid and tumoral cells, which display resistance to the CD95-mediated apoptotic signal. However, all these studies were performed using chimeric CD95Ls (oligomerized forms), which behave as the membrane-bound ligand and not as the naturally processed CD95L. Herein, we examine the biological effects of the metalloprotease-cleaved CD95L on CD95-sensitive activated T-lymphocytes. We demonstrate that cleaved CD95L (cl-CD95L), found increased in sera of systemic lupus erythematosus (SLE) patients as compared to that of healthy individuals, promotes the formation of migrating pseudopods at the leading edge of which the death receptor CD95 is capped (confocal microscopy). Using different migration assays (wound healing/Boyden Chamber/endothelial transmigration), we uncover that cl-CD95L promotes cell migration through a c-yes/Ca2+/PI3K-driven signaling pathway, which relies on the formation of a CD95-containing complex designated the MISC for Motility-Inducing Signaling Complex. These findings revisit the role of the metalloprotease-cleaved CD95L and emphasize that the increase in cl-CD95L observed in patients affected by chronic inflammatory disorders may fuel the local or systemic tissue damage by promoting tissue-filtration of immune cells
The Necrotic Signal Induced by Mycophenolic Acid Overcomes Apoptosis-Resistance in Tumor Cells
The amount of inosine monophosphate dehydrogenase (IMPDH), a pivotal enzyme for the biosynthesis of the guanosine tri-phosphate (GTP), is frequently increased in tumor cells. The anti-viral agent ribavirin and the immunosuppressant mycophenolic acid (MPA) are potent inhibitors of IMPDH. We recently showed that IMPDH inhibition led to a necrotic signal requiring the activation of Cdc42.Herein, we strengthened the essential role played by this small GTPase in the necrotic signal by silencing Cdc42 and by the ectopic expression of a constitutive active mutant of Cdc42. Since resistance to apoptosis is an essential step for the tumorigenesis process, we next examined the effect of the MPA–mediated necrotic signal on different tumor cells demonstrating various mechanisms of resistance to apoptosis (Bcl2-, HSP70-, Lyn-, BCR-ABL–overexpressing cells). All tested cells remained sensitive to MPA–mediated necrotic signal. Furthermore, inhibition of IMPDH activity in Chronic Lymphocytic Leukemia cells was significantly more efficient at eliminating malignant cells than apoptotic inducers.These findings indicate that necrosis and apoptosis are split signals that share few if any common hub of signaling. In addition, the necrotic signaling pathway induced by depletion of the cellular amount of GTP/GDP would be of great interest to eliminate apoptotic-resistant tumor cells
The role of c-FLIP splice variants in urothelial tumours
Deregulation of apoptosis is common in cancer and is often caused by overexpression of anti-apoptotic proteins in tumour cells. One important regulator of apoptosis is the cellular FLICE-inhibitory protein (c-FLIP), which is overexpressed, for example, in melanoma and Hodgkin's lymphoma cells. Here, we addressed the question whether deregulated c-FLIP expression in urothelial carcinoma impinges on the ability of death ligands to induce apoptosis. In particular, we investigated the role of the c-FLIP splice variants c-FLIPlong (c-FLIPL) and c-FLIPshort (c-FLIPS), which can have opposing functions. We observed diminished expression of the c-FLIPL isoform in urothelial carcinoma tissues as well as in established carcinoma cell lines compared with normal urothelial tissues and cells, whereas c-FLIPS was unchanged. Overexpression and RNA interference studies in urothelial cell lines nevertheless demonstrated that c-FLIP remained a crucial factor conferring resistance towards induction of apoptosis by death ligands CD95L and TRAIL. Isoform-specific RNA interference showed c-FLIPL to be of particular importance. Thus, urothelial carcinoma cells appear to fine-tune c-FLIP expression to a level sufficient for protection against activation of apoptosis by the extrinsic pathway. Therefore, targeting c-FLIP, and especially the c-FLIPL isoform, may facilitate apoptosis-based therapies of bladder cancer in otherwise resistant tumours
Measurement of the Bottom-Strange Meson Mixing Phase in the Full CDF Data Set
We report a measurement of the bottom-strange meson mixing phase \beta_s
using the time evolution of B0_s -> J/\psi (->\mu+\mu-) \phi (-> K+ K-) decays
in which the quark-flavor content of the bottom-strange meson is identified at
production. This measurement uses the full data set of proton-antiproton
collisions at sqrt(s)= 1.96 TeV collected by the Collider Detector experiment
at the Fermilab Tevatron, corresponding to 9.6 fb-1 of integrated luminosity.
We report confidence regions in the two-dimensional space of \beta_s and the
B0_s decay-width difference \Delta\Gamma_s, and measure \beta_s in [-\pi/2,
-1.51] U [-0.06, 0.30] U [1.26, \pi/2] at the 68% confidence level, in
agreement with the standard model expectation. Assuming the standard model
value of \beta_s, we also determine \Delta\Gamma_s = 0.068 +- 0.026 (stat) +-
0.009 (syst) ps-1 and the mean B0_s lifetime, \tau_s = 1.528 +- 0.019 (stat) +-
0.009 (syst) ps, which are consistent and competitive with determinations by
other experiments.Comment: 8 pages, 2 figures, Phys. Rev. Lett 109, 171802 (2012
Comparison of Gene Expression Profiles in Chromate Transformed BEAS-2B Cells
Hexavalent chromium [Cr(VI)] is a potent human carcinogen.
Occupational exposure has been associated with increased risk of respiratory
cancer. Multiple mechanisms have been shown to contribute to Cr(VI) induced
carcinogenesis, including DNA damage, genomic instability, and epigenetic
modulation, however, the molecular mechanism and downstream genes mediating
chromium's carcinogenicity remain to be elucidated.We established chromate transformed cell lines by chronic exposure of normal
human bronchial epithelial BEAS-2B cells to low doses of Cr(VI) followed by
anchorage-independent growth. These transformed cell lines not only
exhibited consistent morphological changes but also acquired altered and
distinct gene expression patterns compared with normal BEAS-2B cells and
control cell lines (untreated) that arose spontaneously in soft agar.
Interestingly, the gene expression profiles of six Cr(VI) transformed cell
lines were remarkably similar to each other yet differed significantly from
that of either control cell lines or normal BEAS-2B cells. A total of 409
differentially expressed genes were identified in Cr(VI) transformed cells
compared to control cells. Genes related to cell-to-cell junction were
upregulated in all Cr(VI) transformed cells, while genes associated with the
interaction between cells and their extracellular matrices were
down-regulated. Additionally, expression of genes involved in cell
proliferation and apoptosis were also changed.This study is the first to report gene expression profiling of Cr(VI)
transformed cells. The gene expression changes across individual chromate
exposed clones were remarkably similar to each other but differed
significantly from the gene expression found in anchorage-independent clones
that arose spontaneously. Our analysis identified many novel gene expression
changes that may contribute to chromate induced cell transformation, and
collectively this type of information will provide a better understanding of
the mechanism underlying chromate carcinogenicity
Comparison of Expression Profiles in Ovarian Epithelium In Vivo and Ovarian Cancer Identifies Novel Candidate Genes Involved in Disease Pathogenesis
Molecular events leading to epithelial ovarian cancer are poorly understood but
ovulatory hormones and a high number of life-time ovulations with concomitant
proliferation, apoptosis, and inflammation, increases risk. We identified genes
that are regulated during the estrous cycle in murine ovarian surface epithelium
and analysed these profiles to identify genes dysregulated in human ovarian
cancer, using publically available datasets. We identified 338 genes that are
regulated in murine ovarian surface epithelium during the estrous cycle and
dysregulated in ovarian cancer. Six of seven candidates selected for
immunohistochemical validation were expressed in serous ovarian cancer,
inclusion cysts, ovarian surface epithelium and in fallopian tube epithelium.
Most were overexpressed in ovarian cancer compared with ovarian surface
epithelium and/or inclusion cysts (EpCAM, EZH2, BIRC5) although BIRC5 and EZH2
were expressed as highly in fallopian tube epithelium as in ovarian cancer. We
prioritised the 338 genes for those likely to be important for ovarian cancer
development by in silico analyses of copy number aberration and
mutation using publically available datasets and identified genes with
established roles in ovarian cancer as well as novel genes for which we have
evidence for involvement in ovarian cancer. Chromosome segregation emerged as an
important process in which genes from our list of 338 were over-represented
including two (BUB1, NCAPD2) for which there
is evidence of amplification and mutation. NUAK2, upregulated in ovarian surface
epithelium in proestrus and predicted to have a driver mutation in ovarian
cancer, was examined in a larger cohort of serous ovarian cancer where patients
with lower NUAK2 expression had shorter overall survival. In conclusion,
defining genes that are activated in normal epithelium in the course of
ovulation that are also dysregulated in cancer has identified a number of
pathways and novel candidate genes that may contribute to the development of
ovarian cancer
Proximity Ligation Assay (PLA) to Evaluate DISC and MISC Composition
International audienceDistribution of the death receptor CD95 into lipid rafts (aggregation) and/or its internalization may contribute to the implementation of the apoptotic signal at the detriment of the non-apoptotic signaling pathway [1-6]. Also CD95 can form different protein complexes via dynamic protein-protein interactions (PPIs) according to its interaction with soluble or transmembrane CD95L. Therefore, spatiotemporal identification of these PPIs is pivotal to anticipate the signaling pathway implemented in cells stimulated with different forms of CD95L. Also, many disorders result from dysfunctions in terms of PPI subcellular distribution and/or their intensity, rendering evaluation of these features crucial to better understand pathogenesis.In situ proximity ligation assay (PLA) is a methodology that offers the possibility to identify PPIs and to determine where these PPIs occur in subcellular location (Fig. 1). Moreover, based on imaging, this method allows a quantification of PPIs at the cellular level and with a higher specificity than classical immunofluorescence assays. We here describe PLA used to confirm CD95/FADD interaction, a protocol that may serve to highlight other CD95 partners
Review of PI3K/mTOR inhibitors entering clinical trials to treat triple negative breast cancers
International audienceBackground: Constitutive activation of the PI3K/mTOR signaling pathway is observed in most, if not all, breast cancers. Accordingly, many PI3K and/or mTOR inhibitors have entered clinical trials, and completed studies should soon reveal the efficacy of these new drug families in the treatment of cancer patients. Objective: We present the PI3K/Akt/mTOR signaling pathway and the structure and the anti-tumor efficiency of some mTOR inhibitors such as rapalogues and competitive inhibitors, which have entered clinical trials. We also discuss some of the clinical trial results associated with these molecules mainly focusing on studies performed on relapsing breast cancer patients-but not only. Results: Most of the clinical trials with PI3K/mTOR inhibitors alone or in combination with chemotherapies were performed in heavily pre-treated patients and revealed non-negligible amounts of partial responses and long-term stable disease for these patients. Therefore, these compounds seem to prevent tumor growth and survival of cancer cells in Human, representing a new range of anti-tumor drugs that can be utilized not only as first-line treatments but as second-and third-line agents for patients who relapse. Conclusion: Drugs inhibiting the PI3K/mTOR signaling pathway may represent tailored anti-tumor agents, paving the way for their clinical application in different tumor types. © 2016 Bentham Science Publishers
CD95 Stimulation with CD95L and DISC Analysis
International audienceCD95 and its ligand CD95L play a major role in immune surveillance and homeostasis. CD95L is expressed by activated T lymphocytes and NK cells to induce apoptosis in cancer and virus-infected cells. The goal of this chapter is to describe a method used to immunoprecipitate CD95 and analyze its associated protein complex in cells stimulated with a cytotoxic CD95L (i.e., Ig-CD95L)
T cell landscape in triple negative breast cancer patients
International audience15th St.Gallen International Breast Cancer Conference - P22