GTL : Une interface User-Friendly pour l'analyse de données biologique pour le Centre Jean Perrin

International audienceGTL : Une interface User-Friendly pour l'analyse de données biologique pour le Centre Jean Perri

Removing noise from pyrosequenced amplicons

Background In many environmental genomics applications a homologous region of DNA from a diverse sample is first amplified by PCR and then sequenced. The next generation sequencing technology, 454 pyrosequencing, has allowed much larger read numbers from PCR amplicons than ever before. This has revolutionised the study of microbial diversity as it is now possible to sequence a substantial fraction of the 16S rRNA genes in a community. However, there is a growing realisation that because of the large read numbers and the lack of consensus sequences it is vital to distinguish noise from true sequence diversity in this data. Otherwise this leads to inflated estimates of the number of types or operational taxonomic units (OTUs) present. Three sources of error are important: sequencing error, PCR single base substitutions and PCR chimeras. We present AmpliconNoise, a development of the PyroNoise algorithm that is capable of separately removing 454 sequencing errors and PCR single base errors. We also introduce a novel chimera removal program, Perseus, that exploits the sequence abundances associated with pyrosequencing data. We use data sets where samples of known diversity have been amplified and sequenced to quantify the effect of each of the sources of error on OTU inflation and to validate these algorithms

Antiprion drugs 6-aminophenanthridine and guanabenz reduce PABPN1 toxicity and aggregation in oculopharyngeal muscular dystrophy

Oculopharyngeal muscular dystrophy (OPMD) is an adult-onset syndrome characterized by progressive degeneration of specific muscles. OPMD is caused by extension of a polyalanine tract in poly(A) binding protein nuclear 1 (PABPN1). Insoluble nuclear inclusions form in diseased muscles. We have generated a Drosophila model of OPMD that recapitulates the features of the disorder. Here, we show that the antiprion drugs 6-aminophenanthridine (6AP) and guanabenz acetate (GA), which prevent formation of amyloid fibers by prion proteins in cell models, alleviate OPMD phenotypes in Drosophila, including muscle degeneration and nuclear inclusion formation. The large ribosomal RNA and its activity in protein folding were recently identified as a specific cellular target of 6AP and GA. We show that deletions of the ribosomal DNA locus reduce OPMD phenotypes and act synergistically with sub-effective doses of 6AP. In a complementary approach, we demonstrate that ribosomal RNA accelerates in vitro fibril formation of PABPN1 N-terminal domain. These results reveal the conserved role of ribosomal RNA in different protein aggregation disorders and identify 6AP and GA as general anti-aggregation molecules

Inhibition of DNA methylation promotes breast tumor sensitivity to netrin-1 interference

In a number of human cancers, NTN1 upregulation inhibits apoptosis induced by its so-called dependence receptors DCC and UNC5H, thus promoting tumor progression. In other cancers however, the selective inhibition of this dependence receptor death pathway relies on the silencing of pro-apoptotic effector proteins. We show here that a substantial fraction of human breast tumors exhibits simultaneous DNA methylation-dependent loss of expression of NTN1 and of DAPK1, a serine threonine kinase known to transduce the netrin-1 dependence receptor pro-apoptotic pathway. The inhibition of DNA methylation by drugs such as decitabine restores the expression of both NTN1 and DAPK1 in netrin-1-low cancer cells. Furthermore, a combination of decitabine with NTN1 silencing strategies or with an anti-netrin-1 neutralizing antibody potentiates tumor cell death and efficiently blocks tumor growth in different animal models. Thus, combining DNA methylation inhibitors with netrin-1 neutralizing agents may be a valuable strategy for combating cancer

Dynamics of MBD2 deposition across methylated DNA regions during malignant transformation of human mammary epithelial cells

DNA methylation is thought to induce transcriptional silencing through the combination of two mechanisms: the repulsion of transcriptional activators unable to bind their target sites when methylated, and the recruitment of transcriptional repressors with specific affinity for methylated DNA. The Methyl CpG Binding Domain proteins MeCP2, MBD1 and MBD2 belong to the latter category. Here, we present MBD2 ChIPseq data obtained from the endogenous MBD2 in an isogenic cellular model of oncogenic transformation of human mammary cells. In immortalized (HMEC-hTERT) or transformed (HMLER) cells, MBD2 was found in a large proportion of methylated regions and associated with transcriptional silencing. A redistribution of MBD2 on methylated DNA occurred during oncogenic transformation, frequently independently of local DNA methylation changes. Genes downregulated during HMEC-hTERT transformation preferentially gained MBD2 on their promoter. Furthermore, depletion of MBD2 induced an upregulation of MBD2-bound genes methylated at their promoter regions, in HMLER cells. Among the 3,160 genes downregulated in transformed cells, 380 genes were methylated at their promoter regions in both cell lines, specifically associated by MBD2 in HMLER cells, and upregulated upon MBD2 depletion in HMLER. The transcriptional MBD2-dependent downregulation occurring during oncogenic transformation was also observed in two additional models of mammary cell transformation. Thus, the dynamics of MBD2 deposition across methylated DNA regions was associated with the oncogenic transformation of human mammary cells

“Biological Geometry Perception”: Visual Discrimination of Eccentricity Is Related to Individual Motor Preferences

In the continuum between a stroke and a circle including all possible ellipses, some eccentricities seem more “biologically preferred” than others by the motor system, probably because they imply less demanding coordination patterns. Based on the idea that biological motion perception relies on knowledge of the laws that govern the motor system, we investigated whether motorically preferential and non-preferential eccentricities are visually discriminated differently. In contrast with previous studies that were interested in the effect of kinematic/time features of movements on their visual perception, we focused on geometric/spatial features, and therefore used a static visual display.In a dual-task paradigm, participants visually discriminated 13 static ellipses of various eccentricities while performing a finger-thumb opposition sequence with either the dominant or the non-dominant hand. Our assumption was that because the movements used to trace ellipses are strongly lateralized, a motor task performed with the dominant hand should affect the simultaneous visual discrimination more strongly. We found that visual discrimination was not affected when the motor task was performed by the non-dominant hand. Conversely, it was impaired when the motor task was performed with the dominant hand, but only for the ellipses that we defined as preferred by the motor system, based on an assessment of individual preferences during an independent graphomotor task.Visual discrimination of ellipses depends on the state of the motor neural networks controlling the dominant hand, but only when their eccentricity is “biologically preferred”. Importantly, this effect emerges on the basis of a static display, suggesting that what we call “biological geometry”, i.e., geometric features resulting from preferential movements is relevant information for the visual processing of bidimensional shapes

Une odyssée en génétique du développement : de la drosophile aux tumeurs mammaires triple négatives !

Certains ne jurent que par leur Destinée et se voient emprunter une voie tracée pour eux. Au mieux d’autres tracent cette ligne eux-mêmes et s'y accrochent obstinément. Je crois au hasard qui provoque des rencontres.Ce sont ces rencontres qui ont construit mon parcours jusqu’ici : à l’IUT (pour ne pas m’engager dans des études trop longues !), au milieu des analyses visant à caractériser les maladies humaines, Arlette Darfeuille-Michaud m’a donné goût à la recherche. Sans pour autant me faire aimer la bactério. La discipline qui étincelait pour moi à la même époque, c’était la Biologie Moléculaire de Serge Alziari. Au moment de choisir un stage de DEA, c’est donc vers lui que je me suis tourné et j’ai découvert le modèle drosophile et sa formule magique : Biologie Moléculaire + Drosophile = Génétique Moléculaire (la contrepartie de cette formule étant un sortilège tout aussi puissant, condensé dans une maxime de Jean-Louis Couderc : « Drosophiliste Un Jours, Drosophiliste Toujours© »). En thèse l’année d’après, Krzysztof Jagla a introduit un nouveau paramètre dans l’équation : la mouche permet d’étudier, de comprendre, voire de soigner les pathologies développementales humaines, que je pensais avoir quittées en même tant que l’IUT. Son réseau (celui du directeur de thèse, pas de l’insecte) m’a permis de creuser ce sillon à travers 2 post-docs. Mon recrutement dans l’équipe d’Yves-Jean Bignon, pionnier de l’oncogénétique en France, a bouclé la boucle : quitte à rechercher pendant 10 ans les causes génétiques de pathologies humaines, autant étudier directement la génétique des êtres humains.Ce document est l'occasion de revenir sur ces travaux de recherche, en expliquant brièvement les résultats principaux obtenus dans les différents laboratoires qui m'ont accueilli. Mon activité actuelle fait l'objet d'une description un peu plus précise. Elle se répartie entre le développement de nouvelles approches à visée de diagnostic, l'établissement de collaborations technologiques et des travaux de recherche au sein de l'équipe d'accueil universitaire ERTICa. Le volet Recherche de ce manuscrit se termine par l'exposé de 2 projets de recherche sur la caractérisation moléculaire du cancer du sein triple négatif. Le premier porte sur l'hétérogénéité tumorale comme moteur de l'échappement métastatique. Il est financé et fera l'objet d'une thèse dont le master 2 est en cours. Le second se développera à plus long terme et s'inscrit dans l'un des axes fondateurs de la nouvelle Unité Inserm IMoST crée en Janvier 2017. Il consiste en un projet de recherche clinique avec analyse précoce des changements moléculaires des tumeurs mammaires triple négatives au cours de leur traitement néoadjuvant, afin de déterminer une signature prédictive de réponse thérapeutique.Ce manuscrit présente également mes activités d'enseignement et les étudiants que j'ai eu l'occasion d'accompagner, et il liste les communications auxquelles j'ai participé, qu'elles soient écrites, orales ou affichées

New Insights into the Implication of Epigenetic Alterations in the EMT of Triple Negative Breast Cancer

Breast cancer is the most common cancer and leading cause of cancer death among women worldwide, encompassing a wide heterogeneity of subtypes with different clinical features. During the last two decades, the use of targeted therapies has emerged in clinical research in order to increase treatment efficiency, improve prognosis and reduce recurrence. However, the triple negative breast cancer (TNBC) subtype remains a clinical challenge, with poor prognosis since no therapeutic targets have been identified. This aggressive breast cancer entity lacks expression of oestrogen receptor (ER) and progesterone receptor (PR), and it does not overexpress human epidermal growth factor receptor 2 (HER2). The major reason for TNBC poor prognosis is early therapeutic escape from conventional treatments, leading to aggressive metastatic relapse. Metastases occur after an epithelial-mesenchymal transition EMT of epithelial cells, allowing them to break free from the primary tumour site and to colonize distant organs. Cancer-associated EMT consists not only of acquired migration and invasion ability, but involves complex and comprehensive reprogramming, including changes in metabolism, expression levels and epigenetic. Recently, many studies have considered epigenetic alterations as the primary initiator of cancer development and metastasis. This review builds a picture of the epigenetic modifications implicated in the EMT of breast cancer. It focuses on TNBC and allows comparisons with other subtypes. It emphasizes the role of the main epigenetic modifications lncRNAs, miRNAs, histone and DNA- modifications in tumour invasion and appearance of metastases. These epigenetic alterations can be considered biomarkers representing potential diagnostic and prognostic factors in order to define a global metastatic signature for TNBC

La cinématique humaine, une caractéristique essentielle du lien action-langage

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