No description available.

No description available.

Understanding the transformation of a single-celled fertilized egg to a highlypatterned embryo with three di stinct axes has long been a goal of developmental biology. The Transforming Growth Factor ~ (TGF-~) family of morphogens have essential roles in many developmental patbways. Nodal, a member of this complex and tightly regulated signalling family is responsible for many of the patterning events taking place in the early vertebrate embryo, including germ layer specification and axis determination. Whi lst many components of this signalling pathway have been identified, the precise levels of control indicate other factors or targets still await identification. A Xenopus induction assay using active a member of the TGF-~ family led to up-regulation of Lrig3. Lrig3 is a single~pass transmembrane protein with a structured extracellular domain containing Leucine Rich Repeats and Immunoglobulin·like domains. The in vitro and in vivo expression of the Lrig gene family (Lrigs 1·3) in the developing mouse embryo and postnatal eNS are investigated. A Lrig3 knock-out mouse was created and analysed and bioinfonnatic data on protein structure and func tion was completed. The expression profiles for Lrig J-3 identify distinct differences in expression domains. Significant but not complete overlap exists between LrigJ and 3 which are expressed together in the primitive streak, node, somites and branchial arches. When combined with £he high degree of structural homology these data suggest that Lrigl 9 • I and 3 are able to compensate for each other. Differing expression is observed in me anterior neural crest, branchial arches, portions of the somite tissue and developing limbs. The Lrig3'/o mouse revealed no identifiable phenotype in the developing or adult mouse reinforcing the strong possibi lity of redundancy in the Lrig gene family. A novel low-complexity region was identified in the intracellular domain of the protein. Three-dimensional structural analysis indicated multiple binding sites in the extracellular portion of Lrig3. Expression of Lrig2 and 3 in the central nervous system indicated potential roles for Lrigs in neural plasticity. Future research should focus on identifying interacting partners for Lrig3 and further investigation of its funct ion by breeding LrigJ';o/Lrig3-1o mice . 10

Microscopy based phenotypic analysis is a powerful technique for investigating biological mechanisms and molecular signalling networks. For effective analysis high throughput approaches are required to estimate the inherent variability of biological systems. Automated image analysis is essential for processing the large volumes of data involved in the assays. Image segmentation is a critical part of the image analysis but obtaining accurate and reliable automated segmentation remains a challenge. hi my research 1 have worked to address the problem of estimating the foreground of a fluorescent nucleus marker image by identifying relationships between clusters of pixels. This technique is proven to be more effective at identifying the image foregrounc than the standard approach of classifying pixels based upon intensity thresholds.

No description available.

Pigment patterns of zebrafish are a beautiful example in which to study key processes of vertebrate development such as neural crest cell migration and patterning of neural crest-derived cell types. This can for example be achieved by characterizing mutants like parade in which pigment cell development is abnormal. Here, we present a chemico-genetic study of the pigment pattern mutant parade; uniquely, this mutant displays ectopic pigment cells in the ventral medial pathway of the trunk but the characteristic stripe pattern of zebrafish embryos is unaffected. Using a positional cloning approach, we have identified the parade gene as the cell surface receptor ednra2. This was further confirmed in transient knock-down assays. Combined sequencing data from three different parade alleles strongly indicates that the mutation disrupts ednra2 receptor function by deleting C-terminal regulatory and structural residues. To expand the available molecular tools in pigment cell research, notably to chemically dissect the parade phenotype, we have participated in small molecule screening for inhibitors of pigment cell development. From this, we have isolated 57 compounds which robustly alter the development of melanophores and iridophores in wild-type embryos; 26 of these compounds additionally affect the parade phenotype, primarily by rescuing the ectopic pigment cells. Notably, chemical rescue has shown that the MEK pathway is important for the development of the parade phenotype. Our study therefore adds to our understanding of pigment pattern formation in zebrafish embryos and reveals novel functions for ednra2 in dorso-ventral patterning and cell type specification of neural crest derivatives.

Une des caractéristiques les plus connues de la programmation métabolique est qu’un événement commun physiopathologique à l'âge adulte obtenu indépendamment du stress nutritionnel au début de la vie. Cela a conduit à penser que les altérations métaboliques dûes au stress nutritionnel précoce pouvaient résulter de la programmation seulement d’un petit nombre de gènes qui agissent comme gardiens d'un réseau de gènes ou d'une voie de signalisation. Ici nous avons l’intention de tester cette hypothèse par l'analyse combinée du transcriptome et méthylome avec des échantillons de foie des rats nés de mères nourries avec une alimentation restreinte en protéines (MPR) ou carencée en donneur de méthyles (MDD) pendant la gestation et la lactation et comparer entre les 2 modèls. Au moment du sevrage, la progéniture MDD a été sacrifiée, tandis que la progéniture du groupe MPR a reçu une nourriture standard jusqu'à l'âge de 6 mois. Les rats à jours 21 nés de mères nourries avec un régime MDD ont 3.269 gènes surexprimé (P <0,0009) et 2.841 gènes sous-exprimés (P <0,0004) par rapport aux témoins. Les modifications de méthylation de l'ADN ont été trouvées dans les régions promotrices de 1.032 gènes. Les analyses fonctionnelles ont révélé que ces gènes sont principalement impliqués dans le métabolisme des lipides et du glucose, du système nerveux, la coagulation, le stress du réticulum endoplasmique et la fonction mitochondriale. Les master genes présentant des changements à la fois dans l'expression et la méthylation d'ADN sont limités à 266 gènes et ils sont principalement impliqués dans le système rénine-angiotensine, le métabolisme de la mitochondrie et de l'homéostasie phospholipide. La plupart de ces master genes participent à la Non Alcoholic Fatty Liver Disease (NAFLD). La restriction protéique maternelle (MPR) a entraîné une augmentation de la masse grasse abnominale, de l'hypertriglycéridémie, de l'hypercholestérolémie et un taux élevé d’acides gras par rapport aux témoins. 3.020 gènes sont surexprimés (P<0,0003) et 3.601 sous-exprimés (P<0,002) au niveau du transcriptome et 3.968 gènes modifiés au niveau du méthylome par rapport aux témoins. L'analyse fonctionnelle a indiqué que les gènes surexprimés sont principalement impliqués dans les voies métaboliques et les gènes sous-exprimés et différemment méthylés sont principalement impliqués dans des processus du développement. 998 master genes ont été trouvés, et léanalyse fonctionnelle de ces gènes a indiqué un effet significatif sur le développement des tissus, la régulation de la transcription et le métabolisme, et beaucoup d'entre eux sont associés à des maladies chroniques comme l'hypertension, l'obésité centrale et le diabète. L'expression des gènes et la méthylation de l'ADN du génome obtenus en utilisant ces modèles ont été comparés aux données de méthylome et de transcriptome précédemment obtenus à partir de foie des rats restreints en protéines et sacrifiés à la naissance. Cette analyse a révélé un ensemble commun de 46 gènes qui sont sur-exprimés et 42 gènes sous-exprimés dans les trois modèles de programmation métabolique par rapport aux animaux témoins. La plupart des gènes surexprimés sont impliqués dans la régulation de la fonction mitochondriale alors que les gènes sous-exprimés sont principalement impliquées dans la régulation de la prolifération cellulaire et l'expression des gènes. Nous avons identifié également un ensemble de 122 gènes dont les niveaux de méthylation ont été modifiés à la fois par une carence en donneurs de méthyle et une restriction protéique. Ces observations soutiennent l’hypothèse qu’un petit nombre de gènes essentiels sont à la base de la programmation de troubles métaboliques, indépendamment du stress nutritionnel / One of the most striking features of metabolic programming is that a common physiopathological output at adulthood is obtained irrespective to the nutritional insult during early life. This has suggested that the metabolic alterations due to early nutritional stress might result from the programming of only a small number of genes which act as gatekeepers of a fundamental gene network or signalling pathway. Here we aimed to test this hypothesis through the combined analysis of the transcriptome and methylome in rat liver samples derived from animals born to dams fed either a protein-restricted diet (MPR) or a methyl donor deficient (MDD) diet through gestation and lactation. At weaning, the offspring born to MDD dams were sacrificed whereas the pups from the MPR group were fed standard chow until the age of 6 months. 21-day-old rats born to mothers fed a MDD diet during gestation and lactation have 3,269 over-expressed (P<0.0009) and 2,841 under-expressed (P<0.0004) genes compared to controls. Modifications of DNA methylation were found in the promoter regions of 1,032 genes. Functional analyses revealed that these genes are mainly involved in glucose and lipid metabolism, nervous system, coagulation, endoplasmic reticulum stress and mitochondrial function. Master genes exhibiting changes in both gene expression and DNA methylation are limited to 266 genes and are mainly involved in the renin-angiotensin system, mitochondrion metabolism and phospholipid homeostasis. Most of these master genes participate in Non Alcoholic Fatty Liver Disease (NAFLD). Maternal protein restriction (MPR) resulted in increased fat mass, hypertriglyceridemia, hypercholesterolemia and high fatty acids compared to control. 3,020 genes were up-regulated (p < 0.0003) and 3,601 (p ? 0.002) down-regulated by MPR compared to controls. Modifications of DNA methylation was found in 3,968 genes. The functional analysis indicated that the overexpressed genes were mainly involved in metabolic pathways and the under-expressed and differentially methylated genes were mainly involved in physiological process. 998 master genes were found, functional analysis of these genes indicated a significant effect on tissue development, regulation of transcription and metabolism, and many of them are associated with chronic diseases such as hypertension, central obesity and diabetes. The genome-wide expression and DNA metylation results obtained using these models, were compared to previous methylome and transcriptome data obtained using liver from MPR pups sacrificed at birth. This analysis revealed a common set of 46 genes that were up regulated and 42 genes down regulated in the three models of metabolic programming compared to control animals. Most of the up regulated genes are involved in the regulation of mitochondrial function whereas the down-regulated genes are mainly involved in the regulation of cell proliferation and gene expression. We identified also a set of 122 genes whose methylation levels were changed both by methyl donor deficiency and protein-restriction. These observations sustain the hypothesis that a small set of core genes underlies the programming of metabolic disorders irrespective of the nutritional insult