Régulation of b3-tubulin expression

Bien que le niveau d expression de la tubuline b3 ait été corrélé à une diminution de la sensibilité aux agents antimicrotubulaires, son mécanisme de régulation était inconnu. Ce travail s est concentré sur la régulation de l expression génique par l exploration successive de deux systèmes dans un modèle de cancer du sein : la lignée cellulaire MCF-7. Tout d abord, la tubuline b3 s est révélée être un gène estrogeno-régulé par une voie dépendante des récepteurs aux estrogènes. Le caractère inductible de l expression de la tubuline b3 a ensuite été confirmé par la vinorelbine. En effet, ce vinca alcaloïde déclenche l expression du gène de la tubuline 3 par l activation d un site AP-1 situé en position 757-767. Enfin le développent d une lignée cellulaire dérivée de MCF-7 et sur-exprimant la tubuline b3 a permis de disposer d un modèle cellulaire en vue de l étude de l implication du niveau de tubuline b3 dans la réponse aux agents antimicrotubulairesAlthough b3-tubulin expression level was correlated with decreased response to antimicrotubule agents, its regulation remained unclear. This work has focused on gene expression regulation with the exploration of two different systems in the breast cancer cell line MCF-7. First b3-tubulin appears to be an estrogen regulated gene via an ER dependant pathway. The inducible character of b3-tubulin expression was then confirmed by vinorelbine. Indeed this vinca alkaloid triggers b3-tubulin gene expression mediated by AP-1 site located in position 757-767. Finally, development of b3-tubulin over-expressing cell line derived from MCF-7 cells permits to dispose of a cellular model to investigate the involvement of b3-tubulin level in antimicrotubule agents responseLYON1-BU.Sciences (692662101) / SudocSudocFranceF

The microtubule poison vinorelbine kills cells independently of mitotic arrest and targets cells lacking the APC tumour suppressor more effectively

Colorectal cancers commonly carry truncation mutations in the adenomatous polyposis coli (APC) gene. The APC protein contributes to the stabilization of microtubules. Consistently, microtubules in cells lacking APC depolymerize more readily in response to microtubule-destabilizing drugs. This raises the possibility that such agents are suitable for treatment of APC-deficient cancers. However, APC-deficient cells have a compromised spindle assembly checkpoint, which renders them less sensitive to killing by microtubule poisons whose toxicity relies on the induction of prolonged mitotic arrest. Here, we describe the novel discovery that the clinically used microtubule-depolymerizing drug vinorelbine (Navelbine) kills APC-deficient cells in culture and in intestinal tissue more effectively than it kills wild-type cells. This is due to the ability of vinorelbine to kill cells in interphase independently of mitotic arrest. Consistent with a role for p53 in cell death in interphase, depletion of p53 renders cells less sensitive to vinorelbine, but only in the presence of wild-type APC. The pro-apoptotic protein BIM (also known as BCL2L11) is recruited to mitochondria in response to vinorelbine, where it can inhibit the anti-apoptotic protein BCL2, suggesting that BIM mediates vinorelbine-induced cell death. This recruitment of BIM is enhanced in cells lacking APC. Consistently, BIM depletion dampens the selective effect of vinorelbine on these cells. Our findings reveal that vinorelbine is a potential therapeutic agent for colorectal cancer, but they also illustrate the importance of the APC tumour suppressor status when predicting therapeutic efficacy

The sequential activation of the mitotic microtubule assembly pathways favors bipolar spindle formation

Centrosome maturation is the process by which the duplicated centrosomes recruit pericentriolar components and increase their microtubule nucleation activity before mitosis. The role of this process in cells entering mitosis has been mostly related to the separation of the duplicated centrosomes and thereby to the assembly of a bipolar spindle. However, spindles can form without centrosomes. In fact, all cells, whether they have centrosomes or not, rely on chromatin-driven microtubule assembly to form a spindle. To test whether the sequential activation of these microtubule assembly pathways, defined by centrosome maturation and nuclear envelope breakdown, plays any role in spindle assembly, we combined experiments in tissue culture cells and Xenopus laevis egg extracts with a mathematical model. We found that interfering with the sequential activation of the microtubule assembly pathways compromises bipolar spindle assembly in tissue culture cells but not in X. laevis egg extracts. Our data suggest a novel function for centrosome maturation that determines the contribution of the chromosomal microtubule assembly pathway and favors bipolar spindle formation in most animal cells in which tubulin is in limiting amounts.T.C. was supported by Formación de Personal Investigador (FPI) Fellowship BES-2010-031355. This work was supported by Spanish ministry grants BFU2009-10202 and BFU2012-37163. We acknowledge support of the Spanish Ministry of Economy and Competitiveness, Centro de Excelencia Severo Ochoa 2013-2017, SEV-2012-0208