Ubiquitinylation et déubiquitinylation dans la régulation de l activation du facteur de transcription NF-kB

L activation de la signalisation NF- B par de nombreux immunorécepteurs met en jeu un large signalosome. Afin de propager cette signalisation en réponse à différents stimuli, l oligomérisation d adaptateurs pourvus de domaines d interaction protéine-protéine est nécessaire. Alors que certains adapteurs sont communs d autres sont spécifiques à un immunorécepteur donné. Une des principales caractéristiques partagées par toutes ses protéines est leur capacité à être poly-ubiquitinylé de façon non-dégradative afin d aboutir à une activation optimale de NF- B. Ce projet avait pour objectif d identifier de nouvelles déubiquitinylases impliquées dans la signalisation NF- B. C est ainsi que nous avons identifié USP34 comme étant un régulateur négatif de la signalisation NF- B induite par le TCR dans des cellules Jurkats, une lignée de lymphocytes T immortalisés. Nos données suggèrent un modèle dans lequel USP34 permet d éviter l activation excessive de NF- B, en agissant directement ou indirectement sur les dimères NF- B/I Ba, en aval d IKK, et en modulant l affinité du facteur de transcription pour l ADN. Parallèlement, l étude du microenvironnement des membranes endocellulaires responsables du recrutement des signalosomes formés en réponse à l activation du TCR, du TNFR et du CD40 a permis l identification d une protéine - clé de la signalisation NF- B, la MTDH. Cette protéine du RE s est révélée être un relais déterminant pour l activation d IKK et donc la propagation du signal NF- B.Large signalosome assembly is a prerequisite for NF- B signaling upon engagement of various immunoreceptors. Adaptor proteins containing protein-protein interaction domains oligomerise in response to such stimuli in order to propagate signaling. Each immunoreceptor uses distinct adaptors, as well as common ones, to achieve that. The main characteristic shared by these proteins is their ability to undergo poly-ubiquitinylation in a non-degradative manner, leading to optimal NF- B activation. In this work, we aimed to identify novel deubiquitinylating enzymes that control ubiquitinylation status. That is how USP34 came up to be a negative regulator of NF- B signaling in TCR-activated Jurkat cells, a T lymphocyte cell line. Our data suggest a model whereby USP34 prevents excessive NF- B activation by acting rather late, directly or indirectly on the NF- B:I Ba dimers, downstream of IKK, altering transcription factor DNA binding affinity. In parallel, studies of the endocellular membrane microenvironment that hosts mature signalosomes in response to TCR-, TNFR- and CD40 ligation led to the identification of an ER-residing protein, Metadherin (MTDH), which seems to globally integrate signaling before forwarding it to downstream pathway components able to activate IKK.PARIS11-SCD-Bib. électronique (914719901) / SudocSudocFranceF

MAVS ubiquitination by the E3 ligase TRIM25 and degradation by the proteasome is involved in type I interferon production after activation of the antiviral RIG-I-like receptors

<p>Abstract</p> <p>Background</p> <p>During a viral infection, the intracellular RIG-I-like receptors (RLRs) sense viral RNA and signal through the mitochondrial antiviral signaling adaptor MAVS (also known as IPS-1, Cardif and VISA) whose activation triggers a rapid production of type I interferons (IFN) and of pro-inflammatory cytokines through the transcription factors IRF3/IRF7 and NF-κB, respectively. While MAVS is essential for this signaling and known to operate through the scaffold protein NEMO and the protein kinase TBK1 that phosphorylates IRF3, its mechanism of action and regulation remain unclear.</p> <p>Results</p> <p>We report here that RLR activation triggers MAVS ubiquitination on lysine 7 and 10 by the E3 ubiquitin ligase TRIM25 and marks it for proteasomal degradation concomitantly with downstream signaling. Inhibition of this MAVS degradation with a proteasome inhibitor does not affect NF-κB signaling but it hampers IRF3 activation, and NEMO and TBK1, two essential mediators in type I IFN production, are retained at the mitochondria.</p> <p>Conclusions</p> <p>These results suggest that MAVS functions as a recruitment platform that assembles a signaling complex involving NEMO and TBK1, and that the proteasome-mediated MAVS degradation is required to release the signaling complex into the cytosol, allowing IRF3 phosphorylation by TBK1.</p

Differentiation Therapy: Targeting Human Renal Cancer Stem Cells with Interleukin 15

n/

Exploiting the potential of autophagy in cisplatin therapy: a new strategy to overcome resistance

Resistance to cisplatin is a major challenge in the current cancer therapy. In order to explore new therapeutic strategies to cisplatin resistance, we evaluated, in a model of lung cancer (H1299 and H460 cell lines), the nature of the pathways leading to cell death. We observed that H1299 displayed a natural resistance to cisplatin due to an inability to trigger an apoptotic response that correlates with the induction of autophagy. However, pharmacological and genetic approaches showed how autophagy was a mechanism associated to cell death rather than to resistance. Indeed, pro-autophagic stimuli such as mTOR or Akt inhibition mediate cell death in both cell lines to a similar extent. We next evaluated the response to a novel platinum compound, monoplatin, able to promote cell death in an exclusive autophagy-dependent manner. In this case, no differences were observed between both cell lines. Furthermore, in response to monoplatin, two molecular hallmarks of cisplatin response (p53 and MAPKs) were not implicated, indicating the ability of this pro-autophagic compound to overcome cisplatin resistance. In summary, our data highlight how induction of autophagy could be used in cisplatin resistant tumours and an alternative treatment for p53 mutated patient in a synthetic lethally approach

Participation of the Cell Polarity Protein PALS1 to T-Cell Receptor-Mediated NF-κB Activation

BACKGROUND: Beside their established function in shaping cell architecture, some cell polarity proteins were proposed to participate to lymphocyte migration, homing, scanning, as well as activation following antigen receptor stimulation. Although PALS1 is a central component of the cell polarity network, its expression and function in lymphocytes remains unknown. Here we investigated whether PALS1 is present in T cells and whether it contributes to T Cell-Receptor (TCR)-mediated activation. METHODOLOGY/PRINCIPAL FINDINGS: By combining RT-PCR and immunoblot assays, we found that PALS1 is constitutively expressed in human T lymphocytes as well as in Jurkat T cells. siRNA-based knockdown of PALS1 hampered TCR-induced activation and optimal proliferation of lymphocyte. We further provide evidence that PALS1 depletion selectively hindered TCR-driven activation of the transcription factor NF-κB. CONCLUSIONS: The cell polarity protein PALS1 is expressed in T lymphocytes and participates to the optimal activation of NF-κB following TCR stimulation

Rôle et mécanismes régulateurs des protéines pro-apoptotiques Bim et Puma dans l'apoptose des lymphocytes B normaux et tumoraux

LE KREMLIN-B.- PARIS 11-BU Méd (940432101) / SudocSudocFranceF

Mitochondrial dynamics regulate the RIG-I-like receptor antiviral pathway

The intracellular retinoic acid-inducible gene I-like receptors (RLRs) sense viral ribonucleic acid and signal through the mitochondrial protein mitochondrial antiviral signalling (MAVS) to trigger the production of type I interferons and proinflammatory cytokines. In this study, we report that RLR activation promotes elongation of the mitochondrial network. Mimicking this elongation enhances signalling downstream from MAVS and favours the binding of MAVS to stimulator of interferon genes, an endoplasmic reticulum (ER) protein involved in the RLR pathway. By contrast, enforced mitochondrial fragmentation dampens signalling and reduces the association between both proteins. Our finding that MAVS is associated with a pool of mitofusin 1, a protein of the mitochondrial fusion machinery, suggests that MAVS is capable of regulating mitochondrial dynamics to facilitate the mitochondria–ER association required for signal transduction. Importantly, we observed that viral mitochondria-localized inhibitor of apoptosis, a cytomegalovirus (CMV) antiapoptotic protein that promotes mitochondrial fragmentation, inhibits signalling downstream from MAVS, suggesting a possible new immune modulation strategy of the CMV