70 research outputs found
CTL Escape Mediated by Proteasomal Destruction of an HIV-1 Cryptic Epitope
Cytotoxic CD8+ T cells (CTLs) play a critical role in controlling viral
infections. HIV-infected individuals develop CTL responses against epitopes
derived from viral proteins, but also against cryptic epitopes encoded by viral
alternative reading frames (ARF). We studied here the mechanisms of HIV-1 escape
from CTLs targeting one such cryptic epitope, Q9VF, encoded by an
HIVgag ARF and presented by HLA-B*07. Using PBMCs of
HIV-infected patients, we first cloned and sequenced proviral DNA encoding for
Q9VF. We identified several polymorphisms with a minority of proviruses encoding
at position 5 an aspartic acid (Q9VF/5D) and a majority encoding an asparagine
(Q9VF/5N). We compared the prevalence of each variant in PBMCs of
HLA-B*07+ and HLA-B*07- patients. Proviruses encoding Q9VF/5D were
significantly less represented in HLA-B*07+ than in HLA-B*07-
patients, suggesting that Q9FV/5D encoding viruses might be under selective
pressure in HLA-B*07+ individuals. We thus analyzed ex
vivo CTL responses directed against Q9VF/5D and Q9VF/5N. Around
16% of HLA-B*07+ patients exhibited CTL responses targeting Q9VF
epitopes. The frequency and the magnitude of CTL responses induced with Q9VF/5D
or Q9VF/5N peptides were almost equal indicating a possible cross-reactivity of
the same CTLs on the two peptides. We then dissected the cellular mechanisms
involved in the presentation of Q9VF variants. As expected, cells infected with
HIV strains encoding for Q9VF/5D were recognized by Q9VF/5D-specific CTLs. In
contrast, Q9VF/5N-encoding strains were neither recognized by Q9VF/5N- nor by
Q9VF/5D-specific CTLs. Using in vitro proteasomal digestions
and MS/MS analysis, we demonstrate that the 5N variation introduces a strong
proteasomal cleavage site within the epitope, leading to a dramatic reduction of
Q9VF epitope production. Our results strongly suggest that HIV-1 escapes CTL
surveillance by introducing mutations leading to HIV ARF-epitope destruction by
proteasomes
Identification of Keratinocyte Growth Factor as a Target of microRNA-155 in Lung Fibroblasts: Implication in Epithelial-Mesenchymal Interactions
International audienceBACKGROUND: Epithelial-mesenchymal interactions are critical in regulating many aspects of vertebrate embryo development, and for the maintenance of homeostatic equilibrium in adult tissues. The interactions between epithelium and mesenchyme are believed to be mediated by paracrine signals such as cytokines and extracellular matrix components secreted from fibroblasts that affect adjacent epithelia. In this study, we sought to identify the repertoire of microRNAs (miRNAs) in normal lung human fibroblasts and their potential regulation by the cytokines TNF-alpha, IL-1beta and TGF-beta. METHODOLOGY/PRINCIPAL FINDINGS: MiR-155 was significantly induced by inflammatory cytokines TNF-alpha and IL-1beta while it was down-regulated by TGF-beta. Ectopic expression of miR-155 in human fibroblasts induced modulation of a large set of genes related to "cell to cell signalling", "cell morphology" and "cellular movement". This was consistent with an induction of caspase-3 activity and with an increase in cell migration in fibroblasts tranfected with miR-155. Using different miRNA bioinformatic target prediction tools, we found a specific enrichment for miR-155 predicted targets among the population of down-regulated transcripts. Among fibroblast-selective targets, one interesting hit was keratinocyte growth factor (KGF, FGF-7), a member of the fibroblast growth factor (FGF) family, which owns two potential binding sites for miR-155 in its 3'-UTR. Luciferase assays experimentally validated that miR-155 can efficiently target KGF 3'-UTR. Site-directed mutagenesis revealed that only one out of the 2 potential sites was truly functional. Functional in vitro assays experimentally validated that miR-155 can efficiently target KGF 3'-UTR. Furthermore, in vivo experiments using a mouse model of lung fibrosis showed that miR-155 expression level was correlated with the degree of lung fibrosis. CONCLUSIONS/SIGNIFICANCE: Our results strongly suggest a physiological function of miR-155 in lung fibroblasts. Altogether, this study implicates this miRNA in the regulation by mesenchymal cells of surrounding lung epithelium, making it a potential key player during tissue injury
Morphological and Chemical Effects of Plasma Treatment with Oxygen (O2) and Sulfur Hexafluoride (SF6) on Cellulose Surface
Activité rémanente de la moxidectine dans le traitement de la dictyocaulose et de l'ostertagiose des bovins
*INRA, centre de Tours Diffusion du document : INRA, centre de ToursNational audienc
Analyse XPS des surfaces de Si et SiO2 exposĂ©es aux plasmas de CHF3 et CHF3âC2F6. PolymĂ©risation et gravure
Des surfaces de Si et SiO2 exposĂ©es Ă des plasmas de CHF 3 et CHF3âC2F6 sont caractĂ©risĂ©es par spectroscopie de photoĂ©lectrons (XPS). Un dĂ©pĂŽt fluorocarbonĂ©, d'Ă©paisseur variable suivant la composition du gaz et le temps d'exposition, est observĂ© Ă la surface des matĂ©riaux ; les groupements CF3, CF2, CF, CâCFx x = 1 Ă 3 sont mis en Ă©vidence. La composition et la structure du polymĂšre sont discutĂ©es. L'interface polymĂšre-substrat est analysĂ©e en dĂ©tail ainsi que le rĂŽle des ions du plasma sur le mĂ©canisme d'initiation du polymĂšre. La compĂ©tition entre les processus de gravure et de polymĂ©risation est Ă©tudiĂ©e dans les mĂ©langes CHF3âC 2F6. Une corrĂ©lation est faite entre les rĂ©sultats d'analyse de surface et ceux d'analyse du plasma par spectromĂ©trie de masse. La gravure du polymĂšre en plasma d'oxygĂšne est enfin Ă©tudiĂ©e : la vitesse de gravure et la pression partielle du principal effluent de gravure COF2 sont Ă©tudiĂ©es en fonction du dĂ©bit ; les surfaces nettoyĂ©es sont caractĂ©risĂ©es
Contamination of Silicon Surfaces Exposed to âCHFâ3 Plasmas: An XPS Study of the Film and the FilmâSurface Interface
Investigation of lanthanum and hafnium-based dielectric films by X-ray reflectivity, spectroscopic ellipsometry and X-ray photoelectron spectroscopy
International audienc
MiR-210 promotes a hypoxic phenotype and increases radioresistance in human lung cancer cell lines.
The resistance of hypoxic cells to radiotherapy and chemotherapy is a major problem in the treatment of cancer. Recently, an additional mode of hypoxia-inducible factor (HIF)-dependent transcriptional regulation, involving modulation of a specific set of micro RNAs (miRNAs), including miR-210, has emerged. We have recently shown that HIF-1 induction of miR-210 also stabilizes HIF-1 through a positive regulatory loop. Therefore, we hypothesized that by stabilizing HIF-1 in normoxia, miR-210 may protect cancer cells from radiation. We developed a non-small cell lung carcinoma (NSCLC)-derived cell line (A549) stably expressing miR-210 (pmiR-210) or a control miRNA (pmiR-Ctl). The miR-210-expressing cells showed a significant stabilization of HIF-1 associated with mitochondrial defects and a glycolytic phenotype. Cells were subjected to radiation levels ranging from 0 to 10 Gy in normoxia and hypoxia. Cells expressing miR-210 in normoxia had the same level of radioresistance as control cells in hypoxia. Under hypoxia, pmiR-210 cells showed a low mortality rate owing to a decrease in apoptosis, with an ability to grow even at 10 Gy. This miR-210 phenotype was reproduced in another NSCLC cell line (H1975) and in HeLa cells. We have established that radioresistance was independent of p53 and cell cycle status. In addition, we have shown that genomic double-strand breaks (DSBs) foci disappear faster in pmiR-210 than in pmiR-Ctl cells, suggesting that miR-210 expression promotes a more efficient DSB repair. Finally, HIF-1 invalidation in pmiR-210 cells removed the radioresistant phenotype, showing that this mechanism is dependent on HIF-1. In conclusion, miR-210 appears to be a component of the radioresistance of hypoxic cancer cells. Given the high stability of most miRNAs, this advantage could be used by tumor cells in conditions where reoxygenation has occurred and suggests that strategies targeting miR-210 could enhance tumor radiosensitization
Structural analysis of RF sputtered Ge-Sb-Se thin films by Raman and X-ray photoelectron spectroscopies
Chalcogenide thin films (GeSe2)100 â x(Sb2Se3)x (with x = 10 and 50) were deposited by Radio-frequency (RF) magnetron sputtering. In order to study the impact of Ar pressure on the structure and the composition of selenide thin films structural properties of thin films and targets were investigated by means of Raman scattering spectroscopy and X-ray photoelectron spectroscopy (XPS). Under low pressure (5 · 10â 3 mbar), the increase of wrong bonds like Ge(Sb)-Ge(Sb) was confirmed by Raman and also XPS for both composition. The observed structural changes with Ar pressure are linked with modification of the composition of the selenide films analyzed by EDS and XPS. Furthermore for higher Ar pressure (5 · 10â 2 mbar), RF sputtered thin film and target structure present a great similarity. These differences driven by Ar pressure modification are probably related to distinctive sputtering rate and mean free path of the particles ejected from target for the different Ar pressures
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