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Who is involved in EuroRETT ?
You will find below a presentation of the research carried out in the different laboratories participating to EuroRETT. The following laboratories are involved (by alphabetical order) :
- Thierry BIENVENU – Inserm U567 – Paris, FRANCE.
- Vania BROCCOLI – San Raffaele Scientific Institute – Milano, ITALY.
- Cristina CARDOSO – Technische Universität – Darmstadt, GERMANY.
- Maurizio D’ESPOSITO - CNR - Naples, ITALY.
- Manel ESTELLER – National Cancer Center – Madrid, SPAIN.
- Eva GAK – Sheba Medical Center – Tel Hashomer, ISRAEL.
- Maurizio GIUSTETTO - Torino University - Torino, ITALY.
- Peter HUPPKE – Faculty of Medicine – Göttingen, GERMANY.
- Etienne JOLY - IPBS - Toulouse, FRANCE.
- Nicoletta LANDSBERGER - University of Insubria - Busto Arsizio, ITALY.
- Giovanni LAVIOLA – Istituto Superiore di Sanità – Rome, ITALY.
- Giovanni LÉVI – CNRS 5166 – Paris, FRANCE.
- Giorgio PINI - UONPIA - Viareggio, ITALY.
- Tommaso PIZZORUSSO - CNR - Pisa, ITALY.
- Alessandra RENIERI - University of Siena - Siena, ITALY.
- Silvia RUSSO – Istituto Auxologico Italiano – Milano, ITALY.
- Laurent VILLARD – Inserm U910 – Marseille, FRANCE (Coordination).
21/10/2008
Thierry BIENVENU - Institut Cochin, Inserm U567, Paris.
1- [Thierry Bienvenu, Nadia Bahi-Buisson].
Following the implication of the MECP2 gene in Rett syndrome (RTT), we initially determined the frequency and the distribution of the sequence variations in the MECP2 and CDKL5 genes in the French population (Bienvenu et al. Hum Mol Genet 2000 ; Bienvenu et al. Hum Mutat 2001 ; Bienvenu et al. Genet Test 2002). Since 2002, we obtained an important collection a. of genomic DNA from MECP2 or CDKL5 mutated patients (more than 115 and 20 patients, respectively) b. of lymphoblastoid cells from patients with MECP2 mutations (more than 20) c. of fibroblasts from patients with MECP2 or CDKL5 mutations (more than 15) This large collection of biological material should be relevant for other teams of the EuroRett consortium - to establish genotype – phenotype correlations on a large European cohort - to identify other genetic factors which could modulate the phenotype Twin studies could be an interesting alternative to identify the influence of genetic and non-genetic factors (such as nutrition, therapeutics, ..)
2- [Nadia Bahi-Buisson, T.Bienvenu].
We also developed an interactive French clinico-biological database ([SYRENE->http://afsr.in2p3.fr/RETT/ ]) devoted to RTT , supplemented continuously via Internet by many French clinicians and geneticists. Basically, this database contains more than 100 files of patients with typical or atypical Rett syndrome. This database should enable us to better define the natural history of the disease according to the mutations, to carry out correlations between the clinical phenotype and the genotype on a significant sample and to be a tool for clinical trials development. The first meeting organized in Sienna with the help of Alessandra Renieri had to define the criteria to combine the different databases in only one European database.
3- [Juliette Nectoux].
We have a collection of genomic DNA from patients without MECP2 and CDKL5 mutations - to screen new genes involved in typical or atypical RTT - to validate new candidate genes in atypical or typical RTT Recently, we send to the team of Alessandra Renieri 56 DNA samples from patients with severe neonatal epileptic encephalopathy without CDKL5 mutation to screen the novel gene involved in atypical RTT, FOXG1.
4- [Juliette Nectoux and Yann Fichou].
Ongoing projects in our laboratory consist in studying the disturbances of genes expression at the transcript and protein levels in girls with MECP2 or CDKL5 mutations. We have obtained from different patients with MECP2 or CDKL5 mutations different clones of human fibroblasts expressing either the wild type or the mutated alleles. Comparative studies have been performed and new molecular targets of MeCP2 and CDKL5 have been identified. It could be interesting to test these different human targets in the mouse models available in the different teams of the EuroRett, and especially in the new CDKL5 mouse model, if available.
5- [Yann Fichou].
We have started to develop cell models with MECP2 (wild-type and mutated) in frame with GFP. These cells should be used to test drugs which could restore MeCP2 expression (such as PTC124 in patients with non-sense mutations).
20/10/2008
Cristina CARDOSO - “Molecular and Cell Biology of the Epigenome”, TU Darmstadt, Germany.
We have found that the MBD domain of MeCP2 is necessary and sufficient to induce large-scale chromatin reorganization during differentiation (Brero et al. 2005). Hence, we are particularly interested in the consequences of MBD RTT missense mutations on :
a) the ability of MeCP2 to interact with methylated DNA ;
b) the ability of MeCP2 to cluster chromatin.
In parallel, we are investigating using 3D fluorescent in situ hybridization (3D-FISH) whether genes whose expression level is correlated with the level of MeCP2 concomitantly change their nuclear position and move in or out of heterochromatin compartments. For subsequent data analysis, we are developing and validating a semi-automated image analysis tool.
The mode(s) of MeCP2 interaction with DNA and its role on chromatin organization and subsequent changes of gene expression profile, are paramount to understand the function of this protein in normal and pathological conditions and will hopefully provide new grounds for the design of therapeutic targets and strategies.
14/10/2009
Maurizio D’ESPOSITO - Institute of Genetics and Biophysics "A.Buzzati Traverso", Naples.
Rett syndrome (RTT) is a devastating neurological disorder affecting 1:10000 females. After an apparently normal developmental period, stereotypic hand movements, progressive loss of motor and communication skills, autistic features begin to appear. Unfortunately, there are no specific treatments to halt or reverse the progression of the disease and current therapies merely manage some symptoms. Almost 90-95% of classical RTT have pathogenic mutations in the X-linked gene encoding the methyl cytosine-binding protein 2 (MeCP2).
Mecp2 mutant mice have been used as a model system to study the disease mechanisms that however remain unclear. Recent studies demonstrate disease reversibility in these models, suggesting that the neurological defects in MECP2 disorders are not permanent. Among many functions, MECP2 may act as global transcriptional repressor, and mediate the expression of a subset of specific targets in selected neurons. Findings suggest that MeCP2 is a multifunctional protein, with roles in RNA splicing and chromatin remodelling. The latter can be altered by DNA methylation (and thus by MeCP2), leading to subtle but profound consequences on gene expression.
The goal of our project is to study the yet unknown role of MeCP2 in the genomic architectural organization, its relationship with gene expression, and the effects of selected drugs on chromatin remodelling in the brain. To this aim we will use embryonic and neural stem cells, and neuronal primary cultures at various steps of differentiation, obtained from wild type and Mecp2 deficient mice. These cellular models will be preliminarly treated with desipramine, a drug known to ameliorate respiratory abnormalities (often associated with RTT) and to improve survival in RTT mice. Finally, we will exploit a high throughput screening of chemical compounds effective in RTT, on Mecp2 null embryonic stem cells. by utilizing a robotic station available in our Institute.
05/12/2008
Maurizio GIUSTETTO – University of Torino, Torino.
Our lab is interested in the study of the neuronal and synaptic organization of the brain of MeCP2-KO mice, a murine model of Rett’s syndrome that recapitulates the progression of the disease symptoms. We are currently combining different experimental strategies to understand whether structural and-or molecular abnormalities may occur at the level of excitatory connections in the brain of the mutant mice.
MeCP2 regulates epigenetics mechanisms (i.e. histones deacetylation) leading to the remodeling of chromatin structure and modulation of gene expression. Importantly, MeCP2 is involved in activity-dependant regulation of genes important for neural development and function (e.g. BDNF), thus, absence of functional MeCP2 may tamper with correct gene expression and cause neuronal and synaptic dysfunction that leads to disease manifestation. However, what is the role of MeCP2 regulation of gene expression in neuronal and synaptic function is still unclear.
Basic questions we are currently challenging in our laboratory :
1- What synaptic abnormalities are associated with MeCP2 deletion in the brain ?
2- Do structural defects extend to the entire brain or may they be area- or neuron-specific ?
3- Do synaptic defects correlate with the progression of the symptoms or are they present early in development, when KO-animals are asymptomatic ?
4- Is activity-dependant structural plasticity preserved in the absence of MeCP2 ?
To tackle these questions we use several different high-resolution morphological techniques, from diolistic labelling and confocal laser microscopy to quantitative electron microscopy. We are analysing MeCP2 hemizygous male mice at 2 and 4 weeks of age, when these animals are asymptomatic, as well as at 8 weeks of age, when mutants show severe motor defects.
Our initial observations suggest that MeCP2 mutation produces distinctive structural effects on excitatory synapses in different brain areas and indicate that defects in the formation and maintenance of excitatory synaptic connectivity may be important for the pathological signs associated with RTT.
31/08/2009
Etienne JOLY - Institute of Pharmacology and Structural Biology, Toulouse.
Our initial interest in Rett syndrome came from the hypothesis that MeCP2 may play a role in the regulation of the immune system, and more specifically in the expression of MHC molecules in the central nervous system (Miralvès et al., 2007).
For the last couple of years, we have also initiated a program to develop various reagents for the specific detection of MeCP2.
1- A polyclonal rabbit antibody which reacts with the C-terminal portion of MeCP2, which we have shared with several laboratories of the EuroRETT network ;
2- A recombinant form of MeCP2, tagged with a small fragment of the GFP protein, which allows for its extremely sensitive detection with the split-GFP system (manuscript in preparation) ;
3- We are currently developing rabbit polyclonal sera specific for either of the two isoforms of MeCP2, E1 and E2.
24/09/2009
Nicoletta LANDSBERGER – University of Insubria, Busto Arsizio.
PhD POSITION AVAILABLE IN THIS LABORATORY - DOWNLOAD THE PDF FILE WITH DETAILS
Mutations in the X-linked cyclin dependent kinase-like 5 (CDKL5) gene have during the last few years been identified in RTT patients with the Hanefeld variant characterized by the onset of seizures in the first months of life and in patients with early infantile epileptic encephalopathy. Even if this indicates an important role of CDKL5 for nervous system functions, CDKL5 remains almost completely uncharacterized.
Our laboratory is interested in characterizing the role of CDKL5 for neuronal functions. In accordance with the fact that mutations in CDKL5 show a phenotype resembling that caused by mutations in MECP2 we have previously shown that the two proteins work in a common molecular pathway and that, in vitro, MeCP2 is phosphorylated in the presence of CDKL5. Moreover, in collaboration with Vania Broccoli (San Raffaele Scientific Institute, Milan) we have shown that CDKL5 and MeCP2 are coexpressed in most brain regions of the developing mouse brain. The expression profile and subcellular localization of CDKL5 appear more dynamic, though, than that of MeCP2. A major objective within EuroRETT is to understand the role of the MeCP2-CDKL5 interaction for neuronal functions and reveal the role of CDKL5 for proper brain functions. Specifically, our aims are to
- map the MeCP2 residue(s) that are modified by CDKL5 and study the role of this modification in vivo.
- analyze pathological CDKL5 mutations to reveal the molecular mechanisms involved in regulating the activities of the kinase and to start addressing if it is possible to generate a genotype-phenotype correlation.
- generate (in collaboration with Vania Broccoli) a conditional Cdkl5-null mouse model, which will be important for studying the role of CDKL5 for neuronal functions. We believe that this mouse model also will represent a valuable model for the future development of therapeutic approaches for mental retardation associated with infantile spasms.
Charlotte Kilstrup-Nielsen is coordinating the laboratory together with Nicoletta Landsberger.
More information about the group can be obtained logging into our website busto.dipbsf.uninsubria.it/laboratori/epigenetica/
23/06/2009
Laurent VILLARD - Inserm U910, Marseilles.
Our major objective for the EuroRETT project will be to identify the cellular and molecular mechanisms causing catechelaminergic deficits in the Mecp2-deficient mouse and to try to correct them. Specifically we will :
Jean-Christophe Roux, PhD is supervising the work on Rett syndrome in the lab. More information can be obtained by logging into our website www.germaco.net.