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Institut de génétique et de biologie moleculaire et cellulaire

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146 Projects, page 1 of 30
  • Funder: ANR Project Code: ANR-13-BSV8-0024
    Funder Contribution: 300,000 EUR
    Partners: Institut de génétique et de biologie moleculaire et cellulaire

    The bioactive form of vitamin D, calcitriol [1a,25-dihydroxyvitamin D3; VitD] is primarily a calcemic hormone. However, it not only stimulates intestinal calcium absorption, bone calcium resorption and renal calcium reabsorption, but also regulates growth and differentiation of many cell types, and displays immunoregulatory and anti-inflammatory activities. The latter properties provide a basis to treat many diseases (e.g. osteoporosis, autoimmune diseases, neurodegenerative diseases and various types of cancer). Unfortunately, as supra-physiological doses of VitD that are required to exhibit therapeutic activities induce hypercalcemia, causing mineralization of the kidneys, heart, blood vessels and cutaneous tissues that can lead to organ failure and death, this hormone cannot be used as such as a treatment. Calcitriol mediates pleiotropic effects through activation of the Vitamin D Receptor (VDR), a member of the nuclear receptor superfamily which heterodimerizes with retinoid X receptors (RXRs), to control target gene transcription. Rare genetic mutations in human, termed type II Hereditary Vitamin D Resistant Ricket (HVDRR), have confirmed the importance of VDR in the skeletal system. The discovery that VDR is expressed in many cell types/tissues highlights a central role of VitD signaling in many processes. Besides genomic and rapid non-genomic actions of calcitriol, VDR has also ligand-independent functions. Many companies and academic laboratories have synthesized VitD analogs to potentiate beneficial properties of VitD agonists. These analogs were designed before the crystal structure of liganded VDR was elucidated, by introducing chemical modifications of the calcitriol skeleton, and subsequent in vivo screening for specific activities, as their calcemic activity could not be predicted. However, up to now, all compounds identified with decreased calcemic activities have only a low therapeutic index, thus limiting their clinical use. Therefore, to facilitate the development of VitD analogs with wider therapeutic windows, it is crucial to unveil the molecular mechanisms that underlie cell and/or gene selectivity of VitD. To unravel these mechanisms, we propose a novel approach combining structure-function studies and mouse models, based on recent contributions made by the applicants that open up novel questions to be addressed. Taking advantage of VDR-null mice and VDRgem mice, expressing a mutant VDR the activity of which is selectively induced by Gemini VitD analogs, recently established in collaboration between the two partners involved in this project, combined with structural-functional studies, we intend to: - demonstrate that VDR mediates VitD-induced calcemic effects, - identify direct VDR target genes in various mouse tissues, - determine structural modifications of VitD analogs that promote cell and/or promoter specific actions via VDR, - and identify the key structural modifications of VDR responsible for such specificities. The results generated through this challenging project should uncover the cellular and molecular mechanisms underlying tissue/cell specific actions of VitD. The use of this novel knowledge will allow to design in silico VDR agonists and establish cell-based screens, to identify potent VitD analogs with reduced side effects, for the treatment of various diseases.

  • Funder: ANR Project Code: ANR-13-BSV6-0001
    Funder Contribution: 405,600 EUR
    Partners: Institut de génétique et de biologie moleculaire et cellulaire

    Histone proteins form the building blocks of eukaryotic chromatin. Covalent modifications of histones can regulate all DNA dependent processes. Multiple studies have highlighted histone N-terminal modifications as key regulators of chromatin function and the current model is that tail modifications act via the recruitment of specific binders that then regulate many different processes, such as transcription. However, fundamental questions are still unanswered: to date (i) the set of identified histone modifications is still incomplete and very little is known about modifications in the core of the nucleosome (ii) so far functional outcomes have not been directly attributable to a particular modification per se, but rather to the downstream recruitment of proteins with specialized binding domains (and their associated partner) and (iii) we still do not know whether histone modifications actually regulate transcription directly (i.e. have a causative role rather than being mere by-products of e.g. transcription). We want to address these questions. For this we are in particular interested in novel lysine acetylations. that map to regions in close proximity to the DNA, the so-called lateral surface of the histone octamer. Most of the modified residues on this lateral surface make direct or indirect contacts with the DNA. Our working model is that acetylation of these lysines neutralizes the charge of the negatively charged side chain and has therefore the potential to alter DNA-histone interaction. This could in turn regulate nucleosomal mobility/dynamics and hence gene expression directly. Our aim is to systematically and comprehensively study acetylation of lysines on the lateral surface of the histone octamer. Our preliminary mass spectrometry results identified several additional, previously undescribed acetylation sites on the lateral surface. These sites are: histone H3 lysine 64 (H3K64), H3 lysine 115 (H3K115) and H3 lysine 122 (H3K122). With the support of ANR we want to unravel the biological function of these acetylations at the lateral surface of the histone octamer. We will a) systematically study the distribution and dynamics of H3K64, H3K115, H3K122 acetylation in vivo b) investigate their regulation by the identification of the modifying enzymes, c) study the mechanism how these modifications regulate chromatin dynamics and impact on transcription and d) determine the in vivo relevance of these modification. Together these tasks will allow us to understand the function of lateral surface acetylations and thus, it will constitute a significant advance in our understanding of how histone modifications can (directly) impact on important biological processes. Furthermore we will link specific acetylation sites with a defined process(es) (i.e. transcription) and will demonstrate the underlying mechanism(s). This project combines the expertise of the Schneider lab in the functional characterisation of novel histone modifications and chromatin biochemistry with the expertise of the Tora lab in histone acetylation and characterization of the corresponding acetylating enzymes and complexes. Only the combined and complementary expertise will allow us to pursue this research program. Overall the aim of project is (i) to functionally characterise novel histone modifications, to (ii) significantly advance our understanding of how histone acetylations can regulate transcription, (iii) to try to demonstrate that histone modifications can regulate DNA-dependent processes beyond the recruitment of specific binders, as the current model for tail modifications dictates, and (iv) to try to show that acetylation of histone lysines can be causative, rather than merely consequential, with respect to regulating DNA dependent processes, such as transcription.

  • Funder: ANR Project Code: ANR-22-CE14-0017
    Funder Contribution: 485,773 EUR
    Partners: Institut de génétique et de biologie moleculaire et cellulaire

    In the mammalian testis, sustained spermatogenesis relies on spermatogonia (SG) stem cells. Their progeny either remain as stem SG (for self-renewal) or they proliferate as distinct populations of SG progenitors, and differentiate. Later, differentiated SG enter meiosis to achieve genetic recombination and to halve the number of chromosomes, producing thereby haploid gametes (spermatozoa). Fertility ultimately depends not only on SG maintenance and differentiation, but also on proper spermatozoa release. Genetic and pharmacological studies in mice have revealed that retinoic acid (ATRA, the active metabolite of vitamin A) and its nuclear receptors (RARA, RARB and RARG) are major players in male gametogenesis. Our ARDIGAM project aims to demonstrate that distinct SG populations are essential to perpetuate spermatogenesis and to elucidate how RARA in the somatic, supporting, Sertoli cells (SC) and RARG in SG control the fate of each population (aim #1); and that ATRA works in SC by alternately activating and repressing genes required for spermatozoa production and release (aim #2). To do this, we have a collection, unique worldwide by its diversity, of mutant mice in which one or several actors of the ATRA signaling pathway are knocked out. We will apply an innovative approach allowing to capture simultaneously both the transcriptomic and the epigenetic signatures in given single SG and SC, across thousands of cells. Beyond gametogenesis and the treatment of male infertility, our project will help to understand how ATRA can control the differentiation of stem cell populations.

  • Funder: ANR Project Code: ANR-17-CE15-0023
    Funder Contribution: 580,400 EUR
    Partners: Institut de génétique et de biologie moleculaire et cellulaire

    The Ikaros family of transcription factors includes 4 related zinc finger DNA-binding proteins (Ikaros, Aiolos, Helios, Eos) that are expressed mainly in hematopoietic cells, where they are essential for immune cell differentiation and function. Studies in mouse and man have demonstrated the absolute requirement for these proteins in coordinating gene expression during B and T cell development, and as tumor suppressors, whose loss-of-function mutations promote leukemia expansion. These results have revealed with surprising clarity that Ikaros proteins function in a non-redundant fashion in different cell types and stages of differentiation. Yet their underlying mechanisms of action remain unclear. Some studies have shown that Ikaros family proteins can act as transcriptional activators and repressors, through binding to a common GGGAA motif that is present in both proximal and distal regions of target genes. Other studies have indicated that Ikaros family proteins can function as homo- and heterodimers, each with potentially different mechanisms that depend at least in part on the overall chromatin environment and interacting partners, including the NuRD nucleosome remodeling and deacetylase complex, the SWI-SNF ATPase-dependent chromatin remodeling complex, and the PRC2 polycomb repressive complex. These complex scenarios, investigated in different systems, have been difficult to interpret, and they have often led to generalized conclusions about the Ikaros family as a whole. We hypothesize that Ikaros family proteins are unique in their binding specificity and protein interaction, and that these differences underlie their disparate biological activities. Understanding the differences is important because Ikaros family proteins are associated with diverse diseases, and it is critical to gain a clear view of their biological and molecular functions in order to design better ways to block or promote their activities. In this grant, we propose to evaluate the specificity of Ikaros family members at 3 levels: at the physiological level, by investigating a novel requirement for Ikaros family proteins in the DNA damage response of hematopoietic stem cells; at the molecular level, by investigating the regulation of gene expression, DNA binding specificities and protein interaction of Ikaros family homo- and heterodimers; and at the atomic level, by solving the 3-dimensional structure of human Ikaros. Specifically, we will: Aim 1 - Determine the physiological role of Ikaros family proteins in hematopoietic stem cells (HSC). We will study the DNA damage response of Helios-deficient HSCs and immediate progeny (Task 1). We will investigate the mechanism of Helios function (direct gene regulation vs. cooperation with p53 and NuRD complexes) (Task 2). We will determine how Helios differs from Ikaros and Eos in these cells (Task 3). Aim 2 - Define the molecular specificity of the Ikaros family. We will generate novel cellular systems to study homo- and heterodimer function (Task 4). We will investigate the ability of Ikaros family homodimers to modulate gene expression and differentiation in vitro (Task 5). We will determine the target gene repertoire of Ikaros family homo- and heterodimers (Task 6). We will identify their protein partners (Task 7). Aim 3 - Investigate the structure-function relationship of human Ikaros with DNA. We will solve the 3D structure of full-length Ikaros in the presence or absence of target DNA (Task 8). We will predict the consequences of pathological IKZF1 mutations on protein function (Task 9). Our proposed project will thus address fundamental questions of gene regulation by the Ikaros family. We expect that our results will have enormous biomedical potential for immunity and cancer.

  • Funder: ANR Project Code: ANR-12-BSV4-0028
    Funder Contribution: 300,000 EUR
    Partners: Institut de génétique et de biologie moleculaire et cellulaire

    Drug abuse is a chronic relapsing disorder with devastating consequences for individuals and their social environment. A major challenge in recovering from addiction is to maintain a drug-free state, also referred to as abstinent state. Prolonged abstinence is characterized by lowered mood and a negative affective state. These emotional dysfunctions are considered to contribute to relapse, and clinical studies show a marked co-morbidity between addiction and depressive disorders. However, the “abstinence syndrome” has received little attention in preclinical investigations, and the neurobiology of this particular brain state is poorly understood. We have successfully developed a novel mouse model of protracted abstinence to chronic morphine, which reflects some aspects of addiction-depression co-morbidity. In this model, animals previously exposed to a chronic morphine regimen develop despair-like behavior and social interaction deficits. These behavioral modifications are detectable only after prolonged abstinence, and are also observed after heroin or alcohol in pilot experiments. Importantly serotonin (5-HT) levels in the dorsal raphe nucleus (DRN) remain altered after protracted abstinence to morphine, suggesting enduring modifications in 5-HT neurons. Further, Selective Serotonin Uptake Inhibitor (SSRI) treatment during morphine abstinence prevents appearance of the emotional syndrome, indicating a causal implication of the 5-HT system. Serotonergic transmission is a well-established key mediator of emotional homeostasis, but has been neglected in addiction research. The present project will capitalize on this unique mouse model to identify molecular and circuit mechanisms underlying mood disruption in protracted abstinence from drugs of abuse, with a focus on serotonin-associated adaptations. The proposal has three objectives: Aim 1: we will extend our behavioral model using more sophisticated behavioral testing, as well as neurochemical and gene expression end-points. We will also establish that behavioral deficits generalize to heroin and alcohol, which are most disruptive illicit and licit drugs of abuse respectively. Aim 2: we will characterize molecular and signaling adaptations that occur in the DRN upon protracted abstinence to heroin, at both genome-wide transcriptome and miRNome levels. We will also identify genes and networks associated to SSRI-induced normalization of the emotional syndrome. We will further examine these adaptations in protracted abstinence to alcohol, to test generalization of the model at molecular level. Aim 3: we will test the hypothesis that mu and kappa opioid receptors, known to mediate euphoric and dysphoric states respectively, functionally contribute to emotional disruption via 5HT-mediated mechanisms at the level of DRN circuitry. Hence we will examine both heroin- and alcohol-induced abstinence syndromes in dorsal raphe-conditional knockout mice, both at behavioral and molecular levels. The latter approach will also be applicable to any other gene, signaling network or transmitter system, which will be identified upon genome-wide analyses of the DRN, in order to establish the functional relevance of novel candidate genes. Altogether this project will address a critical, yet poorly understood aspect of drug abuse that has not been modeled previously in animal research (1). In addition, the proposal will investigate dysfunction of serotonergic transmission in drug abuse, which remains a virtually unexplored field. The genome-wide analysis of molecular plasticity in the DRN (2) and the functional study of mu and kappa opioid receptors (3) provide both exploratory and hypothesis-driven approaches to molecular mechanisms of drug abstinence. Findings from this proposal will help understanding the negative affect characterizing abstinent individuals, and will open novel avenues in both addiction and depression research representing two major fields in molecular psychiatry.