Prolactin activates tyrosyl phosphorylation of insulin receptor substrate 1 and phosphatidylinositol-3-OH kinase

Prolactin (PRL) has been demonstrated to induce tyrosine phosphorylation and activation of the cytoplasmic tyrosine kinase JAK2. The present study represents an initial effort to identify the phosphorylation repertoire of the PRL receptor (PRLR). For this purpose we have modified the rat PRLR cDNA to encode an additional N-terminal epitope specifically designed to allow the rapid purification of the PRLR and associated proteins from transfected cells. The Flag-tagged PRLR was stably expressed in the human 293 cell line. PRL induced tyrosine phosphorylation of proteins of 85, 95, and 185 kDa from 10 to 30 min after PRL stimulation. Immunoblot analysis of immunoprecipitation indicates that p85 corresponds to the 85-kDa regulatory subunit of phosphatidylinositol (PI)-3' kinase, p95 to PRLR, and p185 to insulin receptor substrate 1 (IRS-1). Both PI-3' kinase and IRS-1 appear to associate with PRLR in a PRL-dependent manner. These results thus indicate that kinases other than JAK2, namely PI-3' kinase, are activated by PRL

Subcellular localization of microcystin in the liver and the gonads of medaka fish acutely exposed to microcystin-LR

International audienceAmong the diverse toxic components produced by cyanobacteria, microcystins (MCs) are one of the most toxic and notorious cyanotoxin groups. Besides their potent hepatotoxicity, MCs have been revealed to induce potential reproductive toxicity in various animal studies. However, little is still known regarding the distribution of MCs in the reproductive organ, which could directly affect reproductive cells. In order to respond to this question, an acute study was conducted in adult medaka fish (model animal) gavaged with 10 μg.g −1 body weight of pure MC-LR. The histological and immunohistochemical examinations reveal an intense distribution of MC-LR within hepatocytes along with a severe liver lesion in the toxin-treated female and male fish. Besides being accumulated in the hepatocytes, MC-LR was also found in the connective tissue of the ovary and the testis, as well as in oocytes and degenerative spermatocyte-like structures but not spermatocytes. Both liver and gonad play important roles in the reproductive process of oviparous vertebrates. This observation constitutes the first observation of the presence of MC-LR in reproductive cells (female, oocytes) of a vertebrate model with in vivo study. Our results, which provide intracellular localization of MC-LR in the gonad, advance our understanding of the potential reproductive toxicity of MC-LR in fish

Global Metabolomic Characterizations of Microcystis spp. Highlights Clonal Diversity in Natural Bloom-Forming Populations and Expands Metabolite Structural Diversity

Cyanobacteria are photosynthetic prokaryotes capable of synthesizing a large variety of secondary metabolites that exhibit significant bioactivity or toxicity. Microcystis constitutes one of the most common cyanobacterial genera, forming the intensive blooms that nowadays arise in freshwater ecosystems worldwide. Species in this genus can produce numerous cyanotoxins (i.e., toxic cyanobacterial metabolites), which can be harmful to human health and aquatic organisms. To better understand variations in cyanotoxin production between clones of Microcystis species, we investigated the diversity of 24 strains isolated from the same blooms or from different populations in various geographical areas. Strains were compared by genotyping with 16S-ITS fragment sequencing and metabolite chemotyping using LC ESI-qTOF mass spectrometry. While genotyping can help to discriminate among different species, the global metabolome analysis revealed clearly discriminating molecular profiles among strains. These profiles could be clustered primarily according to their global metabolite content, then according to their genotype, and finally according to their sampling location. A global molecular network of all metabolites produced by Microcystis species highlights the production of a wide set of chemically diverse metabolites, including a few microcystins, many aeruginosins, microginins, cyanopeptolins, and anabaenopeptins, together with a large set of unknown molecules. These components, which constitute the molecular biodiversity of Microcystis species, still need to be investigated in terms of their structure and potential bioactivites (e.g., toxicity)

Meta-analysis of SHANK Mutations in Autism Spectrum Disorders: A Gradient of Severity in Cognitive Impairments.

International audienceSHANK genes code for scaffold proteins located at the post-synaptic density of glutamatergic synapses. In neurons, SHANK2 and SHANK3 have a positive effect on the induction and maturation of dendritic spines, whereas SHANK1 induces the enlargement of spine heads. Mutations in SHANK genes have been associated with autism spectrum disorders (ASD), but their prevalence and clinical relevance remain to be determined. Here, we performed a new screen and a meta-analysis of SHANK copy-number and coding-sequence variants in ASD. Copy-number variants were analyzed in 5,657 patients and 19,163 controls, coding-sequence variants were ascertained in 760 to 2,147 patients and 492 to 1,090 controls (depending on the gene), and, individuals carrying de novo or truncating SHANK mutations underwent an extensive clinical investigation. Copy-number variants and truncating mutations in SHANK genes were present in ∼1% of patients with ASD: mutations in SHANK1 were rare (0.04%) and present in males with normal IQ and autism; mutations in SHANK2 were present in 0.17% of patients with ASD and mild intellectual disability; mutations in SHANK3 were present in 0.69% of patients with ASD and up to 2.12% of the cases with moderate to profound intellectual disability. In summary, mutations of the SHANK genes were detected in the whole spectrum of autism with a gradient of severity in cognitive impairment. Given the rare frequency of SHANK1 and SHANK2 deleterious mutations, the clinical relevance of these genes remains to be ascertained. In contrast, the frequency and the penetrance of SHANK3 mutations in individuals with ASD and intellectual disability-more than 1 in 50-warrant its consideration for mutation screening in clinical practice

Etude par transgenèse ciblée de l'effet de la forme dominant-négatif du récepteur de la prolactine sur le développement mammaire normal et tumoral

PARIS7-Bibliothèque centrale (751132105) / SudocSudocFranceF

Etude protéomique des effets toxiques de la microcystine-LR sur le foie de Médaka (Oryzias latipes)

Les microcystines (MCs) sont des hépatotoxines inhibitrices de protéines phosphatases PP1 et PP2A. Ces peptides non-ribosomaux retiennent de plus en plus l attention en raison des risques toxiques qu ils peuvent induire et de leur potentiel tumoral. Ces toxines sont produites par des cyanobactéries. Une étude toxicologique a été conduite sur un modèle aquatique : le poisson médaka. Les détails du mécanisme d action des microcystines sont inconnus. Le variant des microcystines le plus toxique et le plus répandu est la microcystine-LR (MC-Leucine-Arginine). La MC-LR a e té utilisée pour mener des expériences centrées sur le foie de médaka. Deux modes d expositions ont été utilisés, la balnéation et le gavage. L analyse différentielle du protéome ainsi que du phosphoprotéome de la fraction cytosolique du foie a été conduit pour obtenir de nouvelles informations sur les réponses à la toxine. Quatre familles de protéines ont été identifiées comme principalement perturbées par la toxine après analyse des profils sur gel 2D des protéines phosphorylées et des protéines totales. Elles paraissent impliquées dans l assemblage du cytosquelette, la transduction des signaux, le stress oxydatif et l apoptose. Ces résultats montrent que l approche protéomique et phosphoprotéomique est prometteuse et doit permettre d identifier des évènements pouvant aboutir à l identification de biomarqueurs spécifiques à la MC-LRMicrocystins (MCs) are hepatotoxins with potent inhibitor activity to protein phosphatases PP1 and PP2A. These non-ribosomal peptides are getting more and more attention due to their acute toxicity and potent tumor-promoting activity. These toxins are produced by freshwater cyanobacteria. A toxicological study has been conducted on aquatic animal models such as the medaka fish. So far, the detailed mechanisms underlying toxicity of microcystins are unknown. The most toxic and most commonly encountered variant in aquatic environment is MC-LR (MC-Leucine-Arginine). It has been used for toxicological investigations on the liver of intoxicated medaka. Two kinds of exposure way were employed, balneation and gavage essays. Differential proteome as well as differential phosphoproteome analyses have been performed for providing new informations on early responses to the toxin. The experiments are also aiming at selecting biomarkers of MC-LR exposure. In the 2-DE gel protein maps from cytosol of liver cells of animals exposed or non-exposed to the cyanotoxin. Four major protein families were identified that modulated in phosphospecific and total protein stain. They appear to be involved in cytoskeleton assembly, cell signalling, oxidative stress and apoptosis. Such results confirm that proteomics and phosphoproteomics approaches may become valuable tools to identify signalling pathways involved in MC-LR effects. From accumulated data, specific pools of biomarkers should possibly be selected as specific for toxin exposurePARIS-Museum Hist.Naturelle (751052304) / SudocSudocFranceF

La toxico-protéomique : prochaine étape dans l'évolution des biomarqueurs environnementaux ?

La toxico-protéomique : prochaine étape dans l'évolution des biomarqueurs environnementaux

Les contaminations chroniques de poissons par les cyanobactéries

Les écosystèmes aquatiques subissent des altérations croissantes, dont l’apparition d’efflorescences à cyanobactéries produisant des cyanotoxines. Ces efflorescences mettent en péril le fonctionnement des milieux aquatiques. Le but de ce projet était l’étude des contaminations chroniques des poissons par les cyanobactéries

Potentiation of growth hormone-induced liver suppressors of cytokine signaling messenger ribonucleic acid by cytokines.

Endotoxin and proinflammatory cytokines such as interleukin-1beta (IL-1beta) and tumor necrosis factor-alpha (TNFalpha) induce a state of GH resistance. A new family of suppressors of cytokine signaling (SOCS), induced by cytokines activating the Janus kinase (JAK)-signal transducer and activator of transcription (STAT) pathway, has been recently identified as a negative feedback loop of intracellular signaling. Overexpression of some SOCS (SOCS-3, CIS, and SOCS-2) has been reported to inhibit the JAK-STAT pathway stimulated by GH. To assess the possible role of these three SOCS proteins in the GH resistance induced by endotoxin and cytokines, we investigated the regulation of their gene expression by endotoxin and GH in rat liver and by proinflammatory cytokines and GH in primary culture hepatocytes. Both GH and lipopolysaccharide induced the three SOCS messenger RNAs (mRNAs) in vivo. In vitro, GH also increased the liver mRNAs encoding SOCS-2, SOCS-3, and CIS. Although IL-1/beta and TNFalpha alone induced only weakly the expression of SOCS-3 and CIS, these cytokines strongly potentiated the induction of these two SOCS by GH. In contrast, IL-6 alone markedly induced SOCS-3 mRNA, but did not potentiate the GH action on SOCS-3 and CIS mRNAs. The GH induction of SOCS-2 was not potentiated by any of these cytokines. Considering the ability of these SOCS to inhibit the JAK-STAT pathway induced by GH, these results suggest that the overexpression of SOCS-3 and CIS mRNAs induced by IL-1beta and TNFalpha or by endotoxin in vivo may play a role in the GH resistance induced by sepsis