Mutations in MeCP2, a methyI-CpG-binding protein that functions as a global transcriptional repressor, are a major cause of Rett Syndrome (RTT), an X-linked progressive neurological disorder. While the selective inactivation of MeCP2 in neurons is sufficient to confer a Rett-like phenotype in mice, the specific functions of MeCP2 in post-mitotic neurons are not known. We have found that MeCP2 binds to a site in BDNF promoter III just 3' to the site of transcriptional initiation and functions to repress expression of the BDNF gene. Membrane depolarization triggers the calcium-dependent phosphorylation and release of MeCP2 from the BDNF promoter, thereby facilitating BDNF promoter Ill-dependent transcription. These findings indicate that MeCP2 plays a key role in the control of activity-dependent gene expression and suggest that the deregulation of this process may underlie the pathology of RTT. To begin to test this hypothesis, we propose the following specific aims: 1) To characterize the sites of membrane depolarization/calcium-dependent MeCP2 phosphorylation. We will utilize a variety of methods to identify sites of activity-regulated phosphorylation on MeCP2 and develop phosphorylation site-specific antibodies to investigate the regulation of these modifications in cultured neurons and brain sections in response to a variety of stimulation protocols. 2) To assess the effect of phosphorylation on MeCP2 activity. Non-phosphorylatable mutant forms of MeCP2 will be generated and expressed in neuronal cultures to test the hypothesis that activity-induced MeCP2 phosphorylation is required for proper regulation of BDNF promoter activity as well as for neuronal processes such as synaptic development and maintenance. 3) To identify additional activity-regulated neuronal targets of MeCP2. Our findings raise the possibility that MeCP2 may be a general regulator of activity-dependent gene expression. We will employ a variety of techniques including gene expression profiling, RT-PCR, and chromatin immunoprecipitation to identify other activity-regulated targets of MeCP2 in post mitotic neurons. It is our hope that the proposed experiments will provide a better understanding of MeCP2 function, give insight into the mechanisms of activity-dependent gene expression, and provide new opportunities for the development of therapeutic strategies to alleviate RTT pathology.

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Research Project (R01)
Project #
5R01NS048276-03
Application #
7022217
Study Section
Neurogenesis and Cell Fate Study Section (NCF)
Program Officer
Mamounas, Laura
Project Start
2004-03-01
Project End
2009-02-28
Budget Start
2006-03-01
Budget End
2007-02-28
Support Year
3
Fiscal Year
2006
Total Cost
$365,822
Indirect Cost
Name
Children's Hospital Boston
Department
Type
DUNS #
076593722
City
Boston
State
MA
Country
United States
Zip Code
02115
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