The balance between repressive and activating chromatin is maintained by the dynamic activity of specific proteins which mediate histone variant incorporation, histone modifications, nucleosome remodeling and DNA methylation. The heritability of the epigenetic profiles established by these proteins insures the maintenance of developmental and differentiation programs, and alterations in these profiles can result in profound and diverse effects on normal development. The study of the various chromatin remodeling factors in a developmental context is therefore of fundamental importance to understanding how epigenetic mechanisms influence metazoan development. - The aim of the proposed research project is to improve our understanding of epigenetic mechanisms of gene regulation in a developmental context using the nematode C. elegans. This nematode offers great potential for genetic analysis because of its rapid life cycle, small size, and ease of laboratory cultivation. The simple anatomy of C. elegans make it also an excellent model organism to study development at the systems level using functional genomics, and there are exceptionally good genomic resources available. The complete genome sequence of C. elegans has been determined, DNA microarrays and an RNAi library containing nearly every gene in the genome are readily available, and protein interactions have been studied on a large scale. - We have been studying the nematode homologues of the highly conserved HP1 (Heterochromatin Protein 1) family. HP1 proteins are key player in the dynamic organization of nuclear architecture, chromatin remodelling and transcriptional silencing, and directly contribute to a repressive chromatin structure by binding to histone H3 methylated on the lysine 9 residue (H3-K9Me). Evidence from mammals suggests than in additional to playing an essential role in heterochromatin formation, HP1 family proteins have a broader role in the epigenetic regulation of gene expression in euchromatic regions. The primordial importance of HP1 in the epigenetic regulation of gene expression is underlined by the fact that it is associated with diverse tumorigenic processes in mammals. Nonetheless, the function of these proteins in specific developmental pathways remains largely unknown. - In 2002, we identified for the first time two HP1 homologues in the C. elegans genome, which we named HPL-1 and HPL-2. We have shown that HPL-2 acts in a transcriptional repressor pathway which includes homologues of the human Rb complex. HPL-2 forms a complex with the LIN-13 Zinc finger protein, another member of the Rb related pathway which includes HPL-2. LIN-13 is required for the recruitment of HPL-2 to chromatin and we have identified potential targets for both genes. Given that both HPL-2 and LIN-13 contain Rb binding motifs, we believe that both proteins may act via Rb to bring about the repression of specific genes. Deletion of hpl-2 results in sterility and growth defects. In contrast, hpl-1 is dispensable for both germline and somatic development. However, HPL-1 and HPL-2 are redundantly required for post-embryonic development, as hpl-1;hpl-2 double mutants do not develop past the larval stage. Our data provides the first direct evidence for both redundant and unique functions of HP1 family proteins in metazoan development . In an RNAi based screen, we have isolated SET-2, the homolog of the human SET1 and mixed lineage leukemia (Mll) genes, as a suppressor of hpl-2 mutant phenotypes. Mammalian SET1/MLL proteins are found in a histone H3K4 specific methyltransferase complex and we have found that the nematode counterparts of other complex subunits also suppress hpl-2 mutant phenotypes. Furthermore, the H3K4 HMT activity of these proteins appears to be conserved in C. elegans. These results suggest that a C. elegans SET1/MLL complex may antagonize HPL-2 repressor function in development. - We plan to pursue the study of HPL-2 by using complementary approaches, i.

Protein phosphorylation-dephosphorylation plays a dominant role in signal transduction networks. However, for many years, protein phosphorylation-dephosphorylation was considered a cell's regulatory arsenal that was exclusive to eukaryotes and it was only after a long period of controversy that the existence of this modification was documented in bacteria. More precisely, concerning tyrosine-kinases, bacteria were thought to be devoid of these enzymes and this persistent tenet was only ruled out in 1997. Since then, a number of bacterial tyrosine-kinases have been identified and characterized in various bacteria, leading thus to the emerging picture that protein phosphorylation on tyrosine is an important regulatory arsenal of bacterial physiology. However, most of bacterial tyrosine-kinases known to date are unrelated to their eukaryotic counterparts and their catalytic mechanisms differ considerably, rendering thus these enzymes hardly predictable. Therefore, it seems that bacteria have developed their own idiosyncratic type of authentic tyrosine-kinases. By doing so, searching for new bacterial tyrosine-kinases and phosphorylation targets should contribute not only to understand better this type of enzyme, but also to access their importance in the life cycle of bacteria, and especially pathogens. One promising venue in this respect is the search for specific inhibitors of this type of enzymes. They present ideal targets for drug development to interfere, slow down and stop the growth of certain bacteria. Hence, it is interesting to develop an integrative approach combining biochemistry, molecular biology, bioinformatics and molecular modelling firstly, toward the characterization of bacterial tyrosine-kinases and their role in the bacterial cell and then, toward the design of drugs to modulate their activity. This is the central theme of this project that can be divided in two main work packages. The first one will consist in extending our initial studies to identify and characterize novel bacterial tyrosine-kinases and their substrates or partners in E. coli grown in several environmental conditions. For this, we propose to perform a systematic approach combining the use of radioactive phosphate, 2D-electophoresis, phosphoaminoacides analysis and mass spectrometry. In addition, bioinformatic analyses will be simultaneously performed to corroborate the experimental data. Then, once identified, and depending on the nature and the function of each phosphoprotein, specific tests will be carried out to measure the effect of phosphorylation on phosphorylated proteins activity. This will notably require the identification of the phosphorylation sites of the targeted substrates. Finally, genomic mutants and protein derivatives will be generated to check how identified tyrosine-kinases and their substrates affect bacterial life cycle. The second work package will concern processing of our biological data to bring new hypothesis with regard to the functioning of tyrosine-kinases and to design drugs. For this, we will develop bioinformatic expert tools to establish common features important for tyrosine-kinases activity. Then, we will perform tyrosine-kinases modelling to design drugs in order to target and affect their activity. This will be based on the work done for eukaryotic protein kinases in which inhibitors are modified peptides with a moiety that can mimic the ATP molecule, the peptide part having a great affinity and specificity for the protein kinase. Then, we will access the potential of the designed drugs to influence the development of E. coli and other pathogenic bacteria. Drugs will be first used to analyze in vitro their effect on tyrosine-kinases, and, depending on the metabolic function of the tyrosine-kinases and substrates, in vivo experiments will be carried out to check the drugs effect on the growth of certain bacteria.

This project proposes an interdisciplinary and original work focusing on modeling of erythropoiesis (production and regulation of red blood cells) and intracellular signaling pathways associated to it, and on the mathematical analysis of the resulting models. The first part of the project will consist in the description of regulation networks leading to immature cell death, differentiation and proliferation, considering the key proteins of these networks. To reach this goal, the main mathematical tools will be ordinary differential equations, with nonlinearities of Hill and/or Michaelis-Menten type. In the second part, we will concentrate on the dynamics of hematopoietic cells playing a role in erythropoiesis, from the less mature to the differentiated cells, by considering the main growth factors (Epo, glucocorticoids, etc.) acting on cell fate. Such a description will necessitate using nonlinear structured (age, maturity) partial differential equations. We will study asymptotic properties (stability, bifurcations, oscillating solutions) of nonlocal equations, nonlocalities being either in time or in the structure variable (cell maturity for instance). Systems of structured partial differential equations will also be analyzed in order to take into account different maturation levels of the considered cells. In the first two parts, theoretical and numerical results will be confronted to experimental data to validate the model. The third part will consist in coupling the two resulting models through a multi-scale system, to provide a more complete erythropoiesis model that will allow the study of regulation loops and their influences on cell fate and dysfunctions. In particular, we will focus our work on erythroleukemia, a blood cancer characterized by a rapid proliferation of immature cells mainly from the red cell lineage. We will search for causes of the disease in cellular regulation loops, using the previously obtained model. The last part of the project will be dedicated to modeling of other hematopoietic lineages, by using the know-how obtained during the erythropoiesis modeling part. A long term objective is to reach a complete modeling incorporating different hematopoiesis lineages. Biological aspects considered in this project will be of importance for biologists and clinicians. One of the objectives is to provide a predictive tool in order to bring explanations on cell fate during erythropoiesis, to better understand intracellular controls mediated by growth factors. Leukemia still represents the second type of cancer in France and we propose to focus our attention on origins and, possibly, treatment, of erythroleukemia, a particular type of acute leukemia. From a mathematical point of view, the project will contribute to the development of new techniques for multiscale model analysis, to the theory of nonlinear and nonlocal structured partial differential equations, by analyzing their stability and the existence of bifurcations. The determination of necessary and sufficient conditions for the existence of a Hopf bifurcation for differential equations with distributed delay will be particularly considered. The team mainly consists of young mathematician researchers, recently recruited by the UMR 5208 Institut Camille Jordan, with an important experience of interdisciplinary work with biologists and clinicians and necessary skills on partial differential equations analysis to achieve the realization of the project. Their current and past research works are related to biological issues. Two team members are biologists, specialists of cell differentiation and red cell lineage. Moreover, all members of the project team are gathered on the same university campus (University of Lyon 1) where numerous interdisciplinary works are performed, supporting the success of the project.

In bilaterian animals, Hox genes play important roles in conveying anteroposterior patterning information to the embryo. On the chromosome, Hox genes are usually linked in clusters and their order in these clusters correlates with their expression during embryogenesis. This so-called collinear expression is conserved in many animals, including most invertebrate chordates and vertebrates. Although in the vertebrate central nervous system (CNS) spatial collinearity of Hox genes is generally respected, there are some important exceptions to this rule. In jawed vertebrates, such as mice, Hox1 and Hox2 genes break collinearity. Whereas the anterior limit of Hox1 genes is limited to the border between rhombomeres 3 and 4, the Hox2 genes extend anteriorly to the limit of rhombomere 2. Importantly, there are even differences among the anterior limits of distinct Hox2 paralogs in jawed vertebrates. Whereas Hoxa2 has an expression limit at the border between rhombomeres 1 and 2 in all vertebrates analyzed including lampreys, Hoxb2 expression does not extend into rhombomere 2, but stops at the boundary between rhombomeres 2 and 3. In contrast, in the amphioxus CNS Hox1 and Hox2 obey collinearity, with the anterior border of Hox1 expression being more rostral than that of Hox2. In chordates, it has been shown that retinoic acid (RA), an endogenous vitamin A-derived morphogen, is directly involved in regulating collinear Hox expression along the anteroposterior body axis of the embryo. For example, in the vertebrate CNS RA treatments lead to an upregulation and anterior shift of Hox gene expression. These data suggest a link between the observed differences in collinear Hox expression and possible differences in the RA signaling pathway of invertebrate chordates and vertebrates. Thus, the aim of the proposed research is to understand the origins of the collinearity break between Hox1 and Hox2 and to elucidate the exact roles of RA signaling in this process. Using a comparative approach involving different animal systems that mark key positions of chordate phylogeny we will address the following questions: • Are there differences in RA signaling activity in the CNS of invertebrate chordates, jawless vertebrates and jawed vertebrates? • What roles does RA signaling have in establishing the anterior limits of Hox1 and Hox2 in invertebrate chordates, jawless vertebrates and jawed vertebrates? • Why is the anterior limit of Hox1 posterior to that of Hox2 in jawed vertebrates, but not in invertebrate chordates, and what is the expression of Hox1 (relative to Hox2) in jawless vertebrates? • Why do the different Hox2 genes in jawed vertebrates have distinct anterior expression limits in the CNS? The proposed work can be subdivided into three different sections: (a) RA signaling activity in the chordate CNS. In order to elucidate, in which regions of the chordate CNS RA signaling is active, we are going to study the functions of the nuclear receptors mediating the RA signal (RAR and RXR) as well as the genes implicated in synthesis (Raldh) and degradation (Cyp26) of endogenous RA. Moreover, we are going to assess the activity of a RA-sensitive construct in the developing chordate CNS. Since these topics have already extensively been studied in jawed vertebrates, we are going to focus our studies mainly on the invertebrate chordate amphioxus as well as on lampreys, which are jawless vertebrates. (b) Expression of anterior Hox genes in the chordate CNS. Here, we are going to study the expression of anterior Hox genes (Hox1 and Hox2) during development and in response to activation or inhibition of RA signaling. Again, we are going to focus mainly on amphioxus and lampreys. Together with the data already published in jawed vertebrates, our work on invertebrate chordates and jawless vertebrates will allow us to assemble a comprehensive gene expression data set, which will represent an important basis for in-depth comparative analyses. (c) Regulation of anterior Hox genes by RA in chordates. Finally, we will investigate the potential molecular link between the direct regulation of anterior Hox genes by RA signaling and the break of collinearity of Hox1 and Hox2 in the chordate CNS. Hence, we will be testing the activity of reporter constructs carrying regulatory sequences of anterior Hox genes in different chordate models (amphioxus, lampreys, chicken, mice). For selection of the regulatory regions, specific attention is going to be given to the location of RA-responsive elements (so-called RAREs). Ultimately, by addressing the four questions raised in this proposal with our comparative experimental studies, we aim to resolve why this alteration of collinear expression of Hox1 and Hox2 genes took place during chordate evolution. This work is thus going to allow a reconstruction of the evolutionary events leading to the elaboration of the anterior CNS early during vertebrate diversification.

Since the creation of the integrated circuit, more than 4 decades ago, a tenfold increase in computing performance is achieved every 5 years. The most significant impact of this trend is the continuing decrease of cost-per-chip which has led to significant improvement in economic productivity and overall quality of life though the proliferation of computers, communication and electronics in general. This outstanding improvement is essentially due to the scaling down of the CMOS technology (Complementary Metal Oxide Semiconductor). The International Technology Roadmap for Semiconductors anticipates that to continue this trend, innovative solutions must be introduced for post-CMOS technology within the next 10 years. To achieve this, a tremendous research effort on emerging devices or materials must be deployed to bring these technologies to an acceptable maturity level. These emerging research devices will need to be compatible with existing CMOS technologies in order to be heterogeneously integrated with CMOS circuits and capitalize on the established semiconductor industry. Also, these devices have to address the issue of power dissipation (<100 W/cm2) to be considered for a viable solution. Members of the Center of Excellence in Information Engineering (CEGI) at the Université de Sherbrooke, Canada have developed a radically new approach for the fabrication of single electron transistors (SETs) one of the emerging research devices proposed for post-CMOS technology. This engineering breakthrough has allowed to achieve SET with low impedance and extraordinarily small capacitances (paving the way for high-speed memories and logic devices), and concomitant reliable operation 100°C above room temperature. Research at INL (INSA Lyon, France) focuses on the modelling and the characterisation of emerging nanodevices (SET, SEM, RTD etc.) and also contributes to original technological process (e.g. AFM nanomanipulation). Since few years, these two labs have developed important collaborations within the International Laboratory for Nanotechnologies and Nanosystems (LIA LN2). The scientific objective of this project consists in stacking Single Electron Devices (transistors - SETs or memories - SEMs) on a standard CMOS technology and providing the tools to make use of such hybrid SET-CMOS technology (device / interconnect models etc.). This project is presented by INL (McF Francis Calmon, Professor Abdalkader Souifi, McF Nicolas Baboux, Professor Brice Gauthier, Engineer David Albertini) and CEGI (Professors Dominique Drouin, Jacques Beauvais and Serge Charlebois) with the support of STMicroelectronics, Crolles, France (CMOS wafer supply) and IBM Bromont, Canada (SED-CMOS die packaging). This research program will address the issue of energy consumption by capitalizing on the previously developed high power efficiency single electron device platform; 1) to heterogeneously integrated SET with CMOS technology from chip to package; 2) to investigate new electronics functionalities; 3) to increase performance of logic and memory devices. The impact of this research is ranging from more energy efficient portable electronic (Laptop, MP3 player, Cellular phone) to universal computer memory (single memory type regrouping DRAM, hard-disk and flash memory).