Evidence for a mitochondrial localization of the retinoblastoma protein

<p>Abstract</p> <p>Background</p> <p>The retinoblastoma protein (Rb) plays a central role in the regulation of cell cycle, differentiation and apoptosis. In cancer cells, ablation of Rb function or its pathway is a consequence of genetic inactivation, viral oncoprotein binding or deregulated hyperphosphorylation. Some recent data suggest that Rb relocation could also account for the regulation of its tumor suppressor activity, as is the case for other tumor suppressor proteins, such as p53.</p> <p>Results</p> <p>In this reported study, we present evidence that a fraction of the total amount of Rb protein can localize to the mitochondria in proliferative cells taken from both rodent and human cells. This result is also supported by the use of Rb siRNAs, which substantially reduced the amount of mitochondrial Rb, and by acellular assays, in which [³⁵S]-Methionine-labeled Rb proteins bind strongly to mitochondria isolated from rat liver. Moreover, endogenous Rb is found in an internal compartment of the mitochondria, within the inner-membrane. This is consistent with the protection of Rb from alkaline treatment, which destroys any interaction of proteins that are weakly bound to mitochondria.</p> <p>Conclusion</p> <p>Although a few data regarding an unspecific cytosolic localization of Rb protein have been reported for some tumor cells, our results are the first evidence of a mitochondrial localization of Rb. The mitochondrial localization of Rb is observed in parallel with its classic nuclear location and paves the way for the study of potential as-yet-unknown roles of Rb at this site.</p

Production et élimination des radicaux libres : les tests cellulaires

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ETUDE DE LA PROTEINE RB DANS L'APOPTOSE INDUITE PAR INACTIVATION DE L'ANTIGENE T DU VIRUS SV40

L'APOPTOSE ET LA SENESCENCE REPLICATIVE SONT DEUX FORMES D'ARRET IRREVERSIBLE DE LA PROLIFERATION CELLULAIRE. L'APOPTOSE EST UNE MORT CELLULAIRE PHYSIOLOGIQUE TANDIS QUE LA SENESCENCE REFLETE LA CAPACITE PROLIFERATIVE LIMITEE DES CELLULES. AFIN D'ETUDIER LE DETERMINISME MOLECULAIRE DE L'ENGAGEMENT DES CELLULES DANS L'APOPTOSE OU LA SENESCENCE, J'AI UTILISE DES FIBROBLASTES DE RAT IMMORTALISES PAR UN MUTANT THERMOSENSIBLE DE L'ANTIGENE T DU VIRUS SV40. A TEMPERATURE RESTRICTIVE, L'ANTIGENE T EST INACTIVE ET LIBERE LA PROTEINE P53 QUI PEUT INDUIRE L'ARRET DE PROLIFERATION ET L'APOPTOSE. LE PHENOTYPE ASSOCIE A LA PERTE DE L'ETAT IMMORTALISE VARIE EN FONCTION DE LA LIGNEE ETUDIEE : LES CELLULES DE LA LIGNEE RETSAF MEURENT PAR APOPTOSE ALORS QUE LES CELLULES DE LA LIGNEE RETSA15 DEVIENNENT SENESCENTES. OUTRE LA PROTEINE P53, L'ANTIGENE T INHIBE LA PROTEINE ONCOSUPPRESSIVE RB. CETTE PROTEINE EST UN REGULATEUR NEGATIF DU FACTEUR DE TRANSCRIPTION E2F-1 QUI FAVORISE L'ENTREE DES CELLULES EN PHASE S DU CYCLE CELLULAIRE AINSI QUE L'EFFET PRO-APOPTOTIQUE DE P53. L'ETUDE COMPARATIVE DES CELLULES RETSAF ET RETSA15 M'A PERMIS D'OBSERVER QUE LA PROTEINE RB EST REGULEE DE FACON DIFFERENTE DANS CES DEUX LIGNEES, A LA FOIS PAR DES PROCESSUS DE PHOSPHORYLATION ET DE CLIVAGE PROTEOLYTIQUE. A TEMPERATURE RESTRICTIVE, L'INACTIVATION DE LA PROTEINE RB PAR PHOSPHORYLATION DANS LES CELLULES RETSAF FAVORISERAIT L'ACTIVITE DU FACTEUR E2F-1, LA PROGRESSION DES CELLULES DANS LE CYCLE CELLULAIRE ET PEUT ETRE L'APOPTOSE. DE FACON SURPRENANTE, LE CLIVAGE DE LA PROTEINE RB PAR DES PROTEASES A CYSTEINE (CASPASES), TELLES QUE LES CASPASES-6 ET -7, FAVORISERAIT LA SURVIE DES CELLULES RETSA15 ET LEUR ARRET DANS LE CYCLE CELLULAIRE. EN ACCORD AVEC CETTE OBSERVATION, J'AI MIS EN EVIDENCE QUE LA PROTEINE BCL-2, UN INHIBITEUR UNIVERSEL DE L'APOPTOSE, FAVORISE LE CLIVAGE DE RB. CETTE PROPRIETE DE BCL-2 POURRAIT PARTICIPER A UNE VOIE D'INHIBITION DE L'APOPTOSE ENCORE JAMAIS DECRITE.PARIS7-Bibliothèque centrale (751132105) / SudocSudocFranceF

Tickets for p53 journey among organelles

International audienc

A corset of adhesions during development establishes individual neural stem cell niches and controls adult behaviour

International audienceNeural stem cells (NSCs) reside in a defined cellular microenvironment, the niche, which supports the generation and integration of neuronal lineages. The mechanisms building a sophisticated niche structure around NSCs, and their functional relevance for neurogenesis are yet to be understood. In the Drosophila larval brain, the cortex glia (CG) encase individual NSC lineages, organizing the stem cell population and newborn neurons into a stereotypic structure. We first found that lineage information is dominant over stem cell fate. We then discovered that, in addition to timing, the balance between multiple adhesion complexes supports the individual encasing of NSC lineages. An intra-lineage adhesion through homophilic Neuroglian interactions provides strong binding between cells of a same lineage, while a weaker interaction through Neurexin-IV exists between CG to NSC lineages. Their loss leads to random, aberrant grouping of several NSC lineages together, and to altered axonal projection of newborn neurons. Further, we link the loss of these two adhesion complexes during development to locomotor hyperactivity in the resulting adults. Altogether, our findings identify a corset of adhesions building a neurogenic niche at the scale of individual stem cell and provide the proof-of-principle that mechanisms supporting niche formation during development define adult behaviour

reaper and bax initiate two different apoptotic pathways affecting mitochondria and antagonized by bcl-2 in Drosophila

International audiencebcl-2 was the first regulator of apoptosis shown to be involved in oncogenesis. Subsequent studies in mammals, in the nematode and in Drosophila revealed wide evolutionary conservation of the regulation of apoptosis. Although dbok/debcl, a member of the bcl-2 gene family described in Drosophila, shows pro-apoptotic activities, no anti-apoptotic bcl-2 family gene has been studied in Drosophila. We have previously reported that the human anti-apoptotic gene bcl-2 is functional in Drosophila, suggesting that the fruit fly shares regulatory mechanisms with vertebrates and the nematode, involving anti-apoptotic members of the bcl-2 family. We now report that bcl-2 suppresses rpr-induced apoptosis in Drosophila. Additionally, we have compared features of bax-and rpr-induced apoptosis. Flow cytometry analysis of wing disc cells demonstrate that both killers trigger mitochondrial defects. Interestingly, bcl-2 suppresses both bax-and rpr-induced mitochondrial defects while the caspase-inhibitor p35 is specific to the rpr pathway. Finally, we show that the inhibition of apoptosis by bcl-2 is associated with the down-regulation of rpr expression

The Drosophila retinoblastoma protein, Rbf1, induces a Debcl-and Drp1-dependent mitochondrial apoptosis

International audienceIn accordance with its tumor suppressor role, the retinoblastoma protein pRb can ensure pro-apoptotic functions. Rbf1, the Drosophila homolog of Rb, also displays a pro-apoptotic activity in proliferative cells. We have previously shown that the Rbf1 pro-apoptotic activity depends on its ability to decrease the level of anti-apoptotic proteins such as the Bcl-2 family protein Buffy. Buffy often acts in an opposite manner to Debcl, the other Drosophila Bcl-2-family protein. Both proteins can localize at the mitochondrion, but the way they control apoptosis still remains unclear. Here, we demonstrate that Debcl and the pro-fission gene Drp1 are necessary downstream of Buffy to trigger a mitochondrial fragmentation during Rbf1-induced apoptosis. Interestingly, Rbf1-induced apoptosis leads to a Debcl-and Drp1-dependent reactive oxygen species production, which in turn activates the Jun Kinase pathway to trigger cell death. Moreover, we show that Debcl and Drp1 can interact and that Buffy inhibits this interaction. Notably, Debcl modulates Drp1 mitochondrial localization during apoptosis. These results provide a mechanism by which Drosophila Bcl-2 family proteins can control apoptosis, and shed light on a link between Rbf1 and mitochondrial dynamics in vivo

Differential adhesion during development establishes individual neural stem cell niches and shapes adult behaviour in Drosophila

International audienceNeural stem cells (NSCs) reside in a defined cellular microenvironment, the niche, which supports the generation and integration of newborn neurons. The mechanisms building a sophisticated niche structure around NSCs and their functional relevance for neurogenesis are yet to be understood. In the Drosophila larval brain, the cortex glia (CG) encase individual NSC lineages in membranous chambers, organising the stem cell population and newborn neurons into a stereotypic structure. We first found that CG wrap around lineage-related cells regardless of their identity, showing that lineage information builds CG architecture. We then discovered that a mechanism of temporally controlled differential adhesion using conserved complexes supports the individual encasing of NSC lineages. An intralineage adhesion through homophilic Neuroglian interactions provides strong binding between cells of a same lineage, while a weaker interaction through Neurexin-IV and Wrapper exists between NSC lineages and CG. Loss of Neuroglian results in NSC lineages clumped together and in an altered CG network, while loss of Neurexin-IV/Wrapper generates larger yet defined CG chamber grouping several lineages together. Axonal projections of newborn neurons are also altered in these conditions. Further, we link the loss of these 2 adhesion complexes specifically during development to locomotor hyperactivity in the resulting adults. Altogether, our findings identify a belt of adhesions building a neurogenic niche at the scale of individual stem cell and provide the proof of concept that niche properties during development shape adult behaviour