WW domain-containing oxidoreductase WWOX, a putative tumor suppressor, has been suggested to be involved in the hyperphosphorylation of Alzheimer¡¦s tau. Tau is a microtubule-associated protein which plays an important role in microtubule assembly and stability. GSK-3b regulates tau by phosphorylation as well as by regulating splice variance. Hyperphosphorylated tau has less affinity toward the microtubules and disrupting microtubule stability. By bioinformatics analysis, we found that WWOX has three GSK3£] binding motifs, FXXXLI/VXRLE. The results of immunoprecipitation demonstrated that WWOX interacted physically with GSK3£], which was supported by the colocalization of WWOX and GSK3£] in immunofluorescence study. Using in vitro GST pull down assay, we demonstrated that WWOX can bind directly to GSK3£] through its ADH/SDR domain. The site-direct mutagenesis was performed to locate the GSK3£] binding sequences within the ADH/SDR domain precisely. By in vitro kinase assay, we found that WWOX can inhibit Ser396 and Ser404 phosphorylation of Tau by GSK3£] in a dose- and time- dependent manner, indicating that WWOX may be involved in regulating GSK3£] activity in cells. It has been demonstrated that inhibition of GSK3£] plays an essential role during neuron differentiation. By in vitro MT formation assay, we showed that GSK3£] hyperphospohorylated Tau failed to promote tubulin polymerization. Together with the Tau hyperphosphorylation inhibited its function to promote microtubule formation and neurite outgrowth, we investigated the effect of WWOX in neuron differentiation. Interestingly, overexpression of WWOX enhanced the SH-SY5Y differentiation with or without retinoic acid treatment. SH-SY5Y cells increase expression of WWOX and decrease expression of pTau S396 as they differentiate in culture. RNAi-mediated knockdown of WWOX in RA-differentiated SH-SY5Y cells caused a decrease in neurite outgrowth. Furthermore, inhibition of GSK3 by Bio enhanced SH-SY5Y differentiation and overexpression of GSK3£] caused a decrease in neurite outgrowth. We concluded that WWOX may be involved in regulating GSK3£] activity to reduce the level of phosphorylated Tau that subsequently promoted the neurite outgrowth during neuron differentiation.

No description available.

Les ataxies cérébelleuses héréditaires sont un ensemble de pathologies neurodégénératives ou neuro-développementales rares responsables d’un handicap fonctionnel important. Nous décrivons la découverte dans deux familles consanguines avec une ataxie cérébelleuse, une épilepsie et un retard mental deux mutations homozygotes dans le gène WWOX à l’aide du séquençage de l’exome d’un des patients de chaque famille. Ce gène était connu comme un gène suppresseur de tumeur. Par une stratégie de capture ciblée de 57 gènes d’ataxies cérébelleuses sur une série de 155 patients, nous avons posé un diagnostic dans 20,6% des cas dont des mutations d’ANO10 et SYNE1. Des études multicentriques ont permis d’étendre les connaissances sur ces maladies et montrer l’existence de phénotypes sévères dans ARCA1.A partir de cette série, nous avons validé en aveugle la pertinence d’un algorithme diagnostique clinico-biologique proposé par l’article de Anheim dans le New England Journal of Medicine en 2012. / Hereditary cerebellar ataxias are a group of neurodegenerative or neurodevelopemental diseases responsible of major disability. We found thanks to exome sequencing mutations in the WWOX gene in two consaguineous families presenting with cerebellar ataxia, epilepsy and mental retardation. This gene was until recently only recognized to be a tumor suppressor.With a 57 ataxia genes targeted capture strategy, next generation sequencing in 155 patients found 20,6% of positive diagnosis, including several new mutations in ANO10 and SYNE1. Multi center studies allow to extend clinical knowledge with severes phenotypes especially in ARCA1.We validate a clinico-biological algorithm for recesssive ataxias diagnosis published by Anheim in the in New England Journal of Medicine, 2012 in a blinded manner.

No description available.

WWOX (WW domain-containing oxidoreductase) is a candidate tumor suppressor gene that has been shown to be involved in various cancers including breast, lung, prostate, gastric and hepatic. The Drosophila ortholog Wwox was identified and subjected to targeted ‘loss of function’ mutagenesis. The resulting mutants were found to be viable when homozygous with no obvious defects in the adult fly. As Wwox mutant flies were found to exhibit an increased sensitivity to ionising radiation (IR), a number of Wwox proteins specifically deleted or mutated at positions consisting of conserved functional protein motifs, or regions that are highly conserved among WWOX / Wwox homologs. The Wwox variants were tested for their ability to modify the IR sensitivity phenotype. In the course of this study, it was found that background mutations introduced during the generation of the mutant flies was responsible for the IR sensitivity phenotype. As a result, proteomic alterations resulting from changes in Wwox protein levels in Drosophila were investigated in order to ascertain the possible molecular functions of the Wwox protein. 2D-DIGE analysis was conducted on a number of different fly genotypes expressing differing levels of Wwox protein in both adult and embryonic flies. The proteomic changes resulting from lack of Wwox function as well as Wwox over-expression were detected with the proteins of interest identified by mass spectrometry (MS) using both MALDI-TOF/TOF-MS and LC-ESI-MS/MS. Label free quantitative MS analysis was also performed in order to determine the most abundant protein(s) in those spots found to contain multiple proteins. These proteomic studies identified changes in a wide variety of proteins with a significant number of metabolic proteins as well as proteins involved in oxidative stress response as a result of different levels of Wwox expression. Of particular interest, consistent changes in different isoforms of superoxide dismutase 1 (Sod1) were identified. Due to the known roles these proteins play in pro and anti-apoptotic pathways, it is possible that Sod1 and Wwox may work in concert to regulate the delicate balance of defence mechanisms in response to environmental stresses, particularly oxidative stress. The protein/gene targets identified in this work therefore offer some insights into normal Wwox function. / Thesis (Ph.D.) -- University of Adelaide, School of Molecular and Biomedical Science, 2008

No description available.

Afin de mieux connaître les mécanismes moléculaires impliqués dans le cancer du sein, nous avons entrepris l’étude de la fonction du gène suppresseur de tumeur WWOX comprenant le site fragile FRA16D. Nous avons mis en évidence l’association de WWOX avec des composants de la voie de signalisation intracellulaire Wnt/β-caténine : Dvl-2, BCL9 et BCL9-2, ainsi que, l’histone deacetylase 3 (HDAC3). Nous avons défini, pour la première fois, WWOX en tant que nouvel inhibiteur de la voie Wnt/β-caténine. Nos résultats suggèrent que WWOX agit sur cette voie en séquestrant Dvl-2 dans le cytoplasme et en inhibant les activités transcriptionnelles de BCL9 et BCL9-2. En outre, nous avons démontré que HDAC3 est également capable d’inhiber l’activité transcriptionelle de BCL9-2. HDAC3 agirait en recrutant WWOX sur BCL9-2 et cela indépendamment de son activité déacétylase. L’inhibition de la voie Wnt/β-caténine par WWOX suggère que l’inhibition de l’expression de WWOX, souvent observée dans le cancer du sein, pourrait conduire à la suractivation de cette voie et par conséquent à la stimulation de la progression tumorale. En parallèle de ce travail, nous avons étudié l’implication des nouveaux partenaires moléculaires de WWOX que nous avons trouvé dans la carcinogenèse mammaire. Nous avons mis en évidence une surexpression de BCL9, et non pas de BCL9-2, dans les tumeurs du sein, cette surexpression serait due, au moins en partie à des polyploïdies et des amplifications du gène, suggérant un rôle important de BCL9 dans cette pathologie. / In our attempt to better understand the molecular mecanisms in breast carcinogenesis, we studied the role of the tumor suppressor gene WWOX that encompasses the common fragile site FRD16D. We have identified actors of the Wnt/β-catenin pathway as new interactors of WWOX: Dvl-2, BCL9 and BCL9-2 just as HDAC3. We show, for the first time, that WWOX is an inhibitor of the Wnt/β-catenin pathway. Our results suggest that WWOX binds Dvl-2 in the cytoplasm and inhibits BCL9 and BCL9-2’s transcriptionnal activities. Moreover, we have shown that HDAC3 inhibits as well the transcriptional activity of BCL9-2 by recruiting it on WWOX. We then demonstrated that HDAC3 acts independently of its deacetylase activity. The inhibition of the Wnt/b-catenin pathway by WWOX suggests that the down expression of WWOX, frequently found in breast tumors, could trigger the over activation of the Wnt/β-catenin pathway and therefore a tumor progression. In parallel, we have studied the implication of the newly identified molecular partners of WWOX in the mammary carcinogenesis. We have identified an overexpression of BCL9, but not BCL9-2, in a large serie of invasive breast tumors, this overexpression is due, at least in part to polyploidy and gene amplification, suggesting BCL9 plays an important role in this pathology.

<p>Funding: Magnus Bergvall Cancer Foundation; Percy Falk foundation for research in breast and prostate cancer; Nyckelfonden; Örebro University Hospital; Lions cancer research foundation, Region Uppsala-Örebro</p>

WW domain-containing oxidoreductase (WWOX) is a novel tumor suppressor gene discovered few years ago. Many researches indicate that expression of WWOX is reduced in a variety of cancers including heptocellular carcinoma (HCC). A recent report suggests that WWOX is implicated in Wnt/£]-catenin pathway which is frequently affected in HCC. In this study, we used immunohistochemical (IHC) staining to analyze the expression of WWOX and Wnt/£]-catenin pathway components in HCC and adjacent non-tumor tissues. Our result showed that WWOX was significantly downregulated in poor differentiated HCC. In addition, downregulation of WWOX was significantly correlated with cytoplasmic £]-catenin expression. We also found that TCF4 was strongly expressed in HCC tissues and the expression was associated with tumor grade and stage. Consequently, our result implied that downregulation of WWOX in HCC might lead to accumulation of £]-catenin in the cytoplasm and the subsequent activation of Wnt/£]-catenin signaling pathway.

No description available.