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Genome-wide activity-dependent MeCP2 phosphorylation regulates nervous system development and function [cultured cortical neurons]

ABSTRACT: To examine potential differences in activity-dependent gene expression, we analyzed mRNA expression in cultured neurons isolated from Wild-type vs MeCP2 S421A mice at 0 (unstimulated), 1 or 6 hours after membrane depolarization by exposure to high extracellular KCl (55mM) All mice were male littermates from one of three litters. We isolated RNA from dissociated cortical cultures (E16+7DIV) isolated from Wild-Type or MeCP2 S421A knock-in mice littermates. Cells were either left unstimulated or depolarized for 1 or 6 hours by addition of 55mM KCl to the media. mRNA expression was analyzed using the Affymetrix GeneChip Mouse Expression Set 430 2.0 microarray platform.

ORGANISM(S): Mus musculus

SUBMITTER: Harrison Gabel

PROVIDER: E-GEOD-31849 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Genome-wide activity-dependent MeCP2 phosphorylation regulates nervous system development and function.

Cohen Sonia S Gabel Harrison W HW Hemberg Martin M Hutchinson Ashley N AN Sadacca L Amanda LA Ebert Daniel H DH Harmin David A DA Greenberg Rachel S RS Verdine Vanessa K VK Zhou Zhaolan Z Wetsel William C WC West Anne E AE Greenberg Michael E ME

Neuron 20111001 1

Autism spectrum disorders such as Rett syndrome (RTT) have been hypothesized to arise from defects in experience-dependent synapse maturation. RTT is caused by mutations in MECP2, a nuclear protein that becomes phosphorylated at S421 in response to neuronal activation. We show here that disruption of MeCP2 S421 phosphorylation in vivo results in defects in synapse development and behavior, implicating activity-dependent regulation of MeCP2 in brain development and RTT. We investigated the mechan ...[more]

PMID: 21982370

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Genome-wide activity-dependent MeCP2 phosphorylation regulates nervous system development and function [ChIP-Seq]

Project description:Autism spectrum disorders such as Rett syndrome (RTT) have been hypothesized to arise from defects in experience-dependent synapse maturation. RTT is caused by mutations in MECP2, a nuclear protein that becomes phosphorylated at S421 in response to neuronal activation. We show here that disruption of MeCP2 S421 phosphorylation in vivo results in defects in synapse development and behavior, implicating activity-dependent regulation of MeCP2 in brain development and RTT. We investigated the mechanism by which S421 phosphorylation regulates MeCP2 function and show by chromatin immunoprecipitation-sequencing that this modification occurs on MeCP2 bound across the genome. The phosphorylation of MeCP2 S421 appears not to regulate the expression of specific genes; rather, MeCP2 functions as a histone-like factor whose phosphorylation may facilitateM-CM-^BM-BM- a genome-wide response of chromatin to neuronal activity during nervous system development. We propose that RTT results in part from a loss of this experience-dependent chromatin remodeling. To examine MeCP2 binding across the neuronal genome and where on the genome MeCP2 is phosphorylated at Serine 421 in response to neuronal activity we performed anti-total MeCP2 and anti-phospho-Serine 421 specific Chromatin immunoprecipitation from cultured cortical neurons that were either left unstimulated or membrane depolarized for 2 hours by addition of 55mM KCl to the media. ChIP DNA was verified for successful IP by qPCR then cloned and sequenced using ABI SOLiD system 4. ChIP was performed from E16 +7DIV disociated cortical cultures from one or two independent dissections using an anti-c-terminal antiserum recognizing MeCP2 independent of its phosphorylation state or with an anti-pS421 antiserum that specifically immunoprecipitates MeCP2 phosphorylated at serine 421. Samples were qPCR validated and sequenced using ABI SOLiD system 4 library preparation and sequencing.

2011-09-08 | E-GEOD-31951 | biostudies-arrayexpress

Genome-wide activity-dependent MeCP2 phosphorylation regulates nervous system development and function [cultured cortical neurons]

Project description:To examine potential differences in activity-dependent gene expression, we analyzed mRNA expression in cultured neurons isolated from Wild-type vs MeCP2 S421A mice at 0 (unstimulated), 1 or 6 hours after membrane depolarization by exposure to high extracellular KCl (55mM) All mice were male littermates from one of three litters.

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Genome-wide activity-dependent MeCP2 phosphorylation regulates nervous system development and function [ChIP-Seq]

Project description:Autism spectrum disorders such as Rett syndrome (RTT) have been hypothesized to arise from defects in experience-dependent synapse maturation. RTT is caused by mutations in MECP2, a nuclear protein that becomes phosphorylated at S421 in response to neuronal activation. We show here that disruption of MeCP2 S421 phosphorylation in vivo results in defects in synapse development and behavior, implicating activity-dependent regulation of MeCP2 in brain development and RTT. We investigated the mechanism by which S421 phosphorylation regulates MeCP2 function and show by chromatin immunoprecipitation-sequencing that this modification occurs on MeCP2 bound across the genome. The phosphorylation of MeCP2 S421 appears not to regulate the expression of specific genes; rather, MeCP2 functions as a histone-like factor whose phosphorylation may facilitateÂ a genome-wide response of chromatin to neuronal activity during nervous system development. We propose that RTT results in part from a loss of this experience-dependent chromatin remodeling. To examine MeCP2 binding across the neuronal genome and where on the genome MeCP2 is phosphorylated at Serine 421 in response to neuronal activity we performed anti-total MeCP2 and anti-phospho-Serine 421 specific Chromatin immunoprecipitation from cultured cortical neurons that were either left unstimulated or membrane depolarized for 2 hours by addition of 55mM KCl to the media. ChIP DNA was verified for successful IP by qPCR then cloned and sequenced using ABI SOLiD system 4.

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RNA bisulfite sequencing of mRNAs from E16.5 mouse cortical neurons depolarized with 55mM KCl for 0h, 2h and 6h.

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