43 research outputs found

    TRIP6 functions in brain ciliogenesis

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    TRIP6, a member of the ZYXIN-family of LIM domain proteins, is a focal adhesion compo- nent. Trip6 deletion in the mouse, reported here, reveals a function in the brain: ependymal and choroid plexus epithelial cells are carrying, unexpectedly, fewer and shorter cilia, are poorly differentiated, and the mice develop hydrocephalus. TRIP6 carries numerous protein interaction domains and its functions require homodimerization. Indeed, TRIP6 disruption in vitro (in a choroid plexus epithelial cell line), via RNAi or inhibition of its homodimerization, confirms its function in ciliogenesis. Using super-resolution microscopy, we demonstrate TRIP6 localization at the pericentriolar material and along the ciliary axoneme. The requirement for homodimerization which doubles its interaction sites, its punctate localiza- tion along the axoneme, and its co-localization with other cilia components suggest a scaf- fold/co-transporter function for TRIP6 in cilia. Thus, this work uncovers an essential role of a LIM-domain protein assembly factor in mammalian ciliogenesis

    RHAMM deficiency disrupts folliculogenesis resulting in female hypofertility

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    The postnatal mammalian ovary contains the primary follicles, each comprising an immature oocyte surrounded by a layer of somatic granulosa cells. Oocytes reach meiotic and developmental competence via folliculogenesis. During this process, the granulosa cells proliferate massively around the oocyte, form an extensive extracellular matrix (ECM) and differentiate into cumulus cells. As the ECM component hyaluronic acid (HA) is thought to form the backbone of the oocyte-granulosa cell complex, we deleted the relevant domain of the Receptor for HA Mediated Motility (RHAMM) gene in the mouse. This resulted in folliculogenesis defects and female hypofertility, although HA-induced signalling was not affected. We report that wild-type RHAMM localises at the mitotic spindle of granulosa cells, surrounding the oocyte. Deletion of the RHAMM C-terminus in vivo abolishes its spindle association, resulting in impaired spindle orientation in the dividing granulosa cells, folliculogenesis defects and subsequent female hypofertility. These data reveal the first identified physiological function for RHAMM, during oogenesis, and the importance of this spindle-associated function for female fertility

    RHAMM deficiency disrupts folliculogenesis resulting in female hypofertility

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    The postnatal mammalian ovary contains the primary follicles, each comprising an immature oocyte surrounded by a layer of somatic granulosa cells. Oocytes reach meiotic and developmental competence via folliculogenesis. During this process, the granulosa cells proliferate massively around the oocyte, form an extensive extracellular matrix (ECM) and differentiate into cumulus cells. As the ECM component hyaluronic acid (HA) is thought to form the backbone of the oocyte-granulosa cell complex, we deleted the relevant domain of the Receptor for HA Mediated Motility (RHAMM) gene in the mouse. This resulted in folliculogenesis defects and female hypofertility, although HA-induced signalling was not affected. We report that wild-type RHAMM localises at the mitotic spindle of granulosa cells, surrounding the oocyte. Deletion of the RHAMM C-terminus in vivo abolishes its spindle association, resulting in impaired spindle orientation in the dividing granulosa cells, folliculogenesis defects and subsequent female hypofertility. These data reveal the first identified physiological function for RHAMM, during oogenesis, and the importance of this spindle-associated function for female fertility

    Characterisation of prostate cancer lesions in heterozygous Men1 mutant mice

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    <p>Abstract</p> <p>Background</p> <p>Mutations of the <it>MEN1 </it>gene predispose to multiple endocrine neoplasia type 1 (MEN1) syndrome. Our group and others have shown that <it>Men1 </it>disruption in mice recapitulates MEN1 pathology. Intriguingly, rare lesions in hormone-dependent tissues, such as prostate and mammary glands, were also observed in the <it>Men1 </it>mutant mice.</p> <p>Methods</p> <p>To study the occurrence of prostate lesions, we followed a male mouse cohort of 47 <it>Men1</it><sup>+/- </sup>mice and 23 age-matched control littermates, starting at 18 months of age, and analysed the prostate glands from the cohort.</p> <p>Results</p> <p>Six <it>Men1</it><sup>+/- </sup>mice (12.8%) developed prostate cancer, including two adenocarcinomas and four <it>in situ </it>carcinomas, while none of the control mice developed cancerous lesions. The expression of menin encoded by the <it>Men1 </it>gene was found to be drastically reduced in all carcinomas, and partial LOH of the wild-type <it>Men1 </it>allele was detected in three of the five analysed lesions. Using immunostaining for the androgen receptor and p63, a basal epithelial cell marker, we demonstrated that the menin-negative prostate cancer cells did not display p63 expression and that the androgen receptor was expressed but more heterogeneous in these lesions. Furthermore, our data showed that the expression of the cyclin-dependent kinase inhibitor CDKN1B (p27), a <it>Men1 </it>target gene known to be inactivated during prostate cell tumorigenesis, was notably decreased in the prostate cancers that developed in the mutant mice.</p> <p>Conclusion</p> <p>Our work suggests the possible involvement of <it>Men1 </it>inactivation in the tumorigenesis of the prostate gland.</p

    Papillomavirus humains et lésions précancéreuses du col utérin (implication de p53, Rb, cycline D1, PCNA, R-EGF, ras)

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    Les pouvoirs oncogéniques des papillomavirus humains (HPV) sont en relation avec la capacité des protéines virales E6 et E7 d'inactiver les gènes suppresseurs de tumeur p53 et Rb. Pour évaluer le processus de cancérogénèse du col utérin, nous avons étudié dans des dysplasies cervicales, l'expression de: PCNA, p53, pRb, cycline D1, récepteurs à EGF (R-EGF), ras et ARNm de E6-E7. Dans l'épithélium normal adjacent aux lésions et dans les régions condylomateuses, l'expression de P53, pRb, cyline D1, R-EGF et ras s'accroit parallèlement à une activité prolifératrice accrue révélée par PCNA. Dans les dysplasies, on observe une disparition de l'expression de p53 et de cycline D1 et une diminution de pRb. L'ARNm codant E6-E7, n'a été mis en évidence que dans les cellules superficielles de régions condylomateuses.Il existe donc une prolifération accrue avec une altération de l'expression de p53, pRb et cycline D1 dans l'épithélium normal adjacent aux dysplasies. L'action des HPV s'exerce dés les étapes initiales de la cancérogénèse. Il apparait trés probable que c'est l'action des protéines E6 et E7 d'HPV sur les cellules épithéliales basales et surtout parabasales qui est à l'origine de la dérégulation de la prolifération épithéliale et de l'apparition des signes de dysplasie.LYON1-BU Santé (693882101) / SudocPARIS-BIUP (751062107) / SudocSudocFranceF

    Poly(ADP-ribose)polymérase-1 (PARP-1) et méthylation de l'ADN, nouveaux partenaires des récepteurs hormonaux dans la carcinogenèse de l'endomètre

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    Les récepteurs des œstrogènes (ER) et de la progestérone (PR) sont les facteurs pronostics et prédictifs les plus documentés dans le cancer endometrioïde. Des études biochimiques ont pu démontrer que la poly(ADP-ribose) polymérase (PARP-1), molécule sensible aux cassures de l ADN est associée avec le domaine de liaison à l ADN de PR. Nous avons étudié la méthylation globale de l ADN et l expression de PARP-1 dans les lésions prénéoplasiques et néoplasiques de l endomètre et leurs corrélations avec PR : les expressions de PARP-1 et de PR sont corrélées, dans chaque stade, positivement (p<0.0001), à l exception des carcinomes non-endometrioides. Une corrélation positive est également observée entre la méthylation globale de l ADN et l expression de PR (p <0.0001). La synergie de ces deux phénomènes : méthylation globale de l ADN et expression de PARP-1 joue un rôle crucial dans la régulation de l action de la progestérone dans le développement du cancer endométrioïde de l endomètreLYON1-BU.Sciences (692662101) / SudocSudocFranceF

    BRCA1 expression during prenatal development of the human mammary gland

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    International audienceGerm-line alterations of BRCA1 are associated with elevated risk of breast cancer. Evidence for the involvement of Brca1 in cellular differentiation and morphogenesis has been obtained in mouse models during embryogenesis. Although the presence of well-conserved functional domains might suggest a similar function for both human and mouse genes, very few data on BRCA1 expression in human fetal tissues are available. We have, therefore, investigated the expression of BRCA1 in the mammary gland from human female fetuses aged between 15 and 33 weeks. Quantification of BRCA1 transcripts, using a competitive reverse transcriptase PCR method, indicates a progressive decrease in BRCA1 expression with increasing fetal age between the 15th and 30th week of gestation. Subsequently, the amount of BRCA1 transcripts becomes similar to that found in adult mammary gland. Analysis of BRCA1 protein revealed, in fetal samples, a 220 kDa band corresponding to the 220 kDa BRCA1 protein described in human cell lines. These later experiments confirm that the relative level of the 220 kDa BRCA1 protein is highest in the early stages of mammary gland development. The temporal patterns of BRCA1 expression in human fetuses suggest a role for BRCA1 in the morphogenesis and differentiation of the human mammary gland

    MeCP2 and MBD2 expression during normal and pathological growth of the human mammary gland

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    International audienceDuring the last years, a direct link between DNA methylation and repressive chromatin structure has been established. This structural modi®cation is mediated by histone deacetylases targeted to the methylated sequences by Methyl Binding Proteins (MBD). Human cancer cells exhibit both a global hypomethylation and some localized hypermethylations suggesting that the deregulation of the methylation machinery is a central event in tumorigenesis. Therefore, we have investigated in human tissues the expression of two major MBDs, MeCP2 and MBD2, during the proliferation of normal breast and in benign and neoplasic breast tumors. Quantitation of the transcripts indicates that MBD2 mRNAs are 20 ± 30-fold more abundant than MeCP2 transcripts in the adult and fetal human mammary gland. In pathological tissues samples MBD2 mRNA levels are significantly higher (P=0.001) in benign tumors compared with normal breast tissues, whereas MeCP2 expression is not modified in these specimens. In neoplasic samples a deregulation of the expression of both genes was found. The amounts of MBD2 and MeCP2 transcripts vary greatly between samples in cancer cells compared to normal breast tissues or benign tumors, and in invasive ductal carcinomas the amount of MBD2 mRNA is signi®cantly (P=0.03) associated with the tumor size. Taken together these data suggest that upregulation of MBD2 might be associated with breast cell proliferation. In line with this hypothesis MBD2 is also upregulated during the prenatal development of the human mammary gland, but in contrast to that observed in tumor cells, MeCP2 is also coordinately upregulated in the fetal breast tissues, suggesting that deregulation of MeCP2 and MBD2 occurs in human breast cancers

    Down-regulation of BRCA1 in human sporadic breast cancer; analysis of DNA methylation patterns of the putative promoter region

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    International audienceGerm-line alterations of BRCA1 are responsible for about 50% of familial breast cancers. Although its biological function(s) has not yet been fully determined, it has been suggested that it may act as a tumor suppressor gene in breast and ovarian cancers. In sporadic breast cancers alterations of BRCA1 have not been detected and in vitro experiments have indicated that BRCA1 negatively regulates cellular proliferation. The present study was designed to identify and quantify, the BRCA1 mRNA levels, in normal and neoplasic human breast tissue. BRCA1 mRNA molecules were quanti®ed using competitive reverse transcriptase PCR assays. DNA methylation patterns of this gene have been analysed by Southern blot experiments using methylation sensitive restriction enzymes. We found that BRCA1 mRNA levels were signi®cantly lower in sporadic breast cancers (37 cases analysed, 24 cases of invasive ductal carcinomas not otherwise speci®ed (NOS), two lobular carcinomas in situ two medullary carcinomas, four invasive lobular carcinomas, two invasive mucinous carcinomas and three invasive ductal carcinomas with predominantly in situ component) compared with normal breast tissues (P=0.0003). This down-regulation of BRCA1 is observed in all histologic types analysed. In invasive ductal carcinomas NOS, this down-regulation does not correlate with any of the prognostic factors studied (tumor size, node status, histologic grade, hormone receptor status). In the samples analysed, alterations of DNA methylation patterns were not dectected in the vicinity of the major transcription start site. These data suggest the involvement of BRCA1 in the carcinogenesis of these histologic types

    ABUNDANCE OF BRCA1 TRANSCRIPTS IN HUMAN CANCER AND LYMPHOBLASTOID CELL LINES CARRYING BRCA1 GERM-LINE ALTERATIONS

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    International audienceA competitive polymerase chain reaction has been developed for quantitation of BRCA1 mRNA. In human cancer cell lines, the amount of BRCA1 mRNA is relatively low, ranging from 6 to 38 copies per cell. The decay rate of these transcripts in actinomycin-treated cells indicates that the half-life of these molecules is about 4 hr, suggesting that the low concentration of BRCA1 messages is not due to molecular unstability. In human lymphoblastoid cell lines derived from patients carrying germ-line alterations of BRCA1, the amount of BRCA1 mRNA per cell is lowered only in cell lines exhibiting alterations leading to specific loss of transcripts from the mutated allele. These data indicate that the amount of BRCA1 available in these cells can be related directly to the number of “active” allel
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