Project description:Background MeCP2, methyl-CpG-binding protein 2, binds to methylated cytosines at CpG dinucleotides, as well as to unmethylated DNA, and affects chromatin condensation. MECP2 mutations in females lead to Rett syndrome, a neurological disorder characterized by developmental stagnation and regression, loss of purposeful hand use and speech, stereotypic hand movements, deceleration of brain growth, autonomic dysfunction and seizures. Most mutations occur de novo during spermatogenesis. Located at Xq28, MECP2 is subject to X inactivation, and affected females are mosaic. Rare hemizygous males suffer from a severe congenital encephalopathy. Methods To identify pathways mis-regulated by MeCP2 deficiency, microarray-based global gene expression studies were carried out on cerebellum of Mecp2 mutant mice. We compared transcript levels in mutant/wildtype male sibs of two different MeCP2-deficient mouse models at 2, 4 and 8 weeks of age. Increased transcript levels were evaluated by real-time quantitative RT-PCR. Chromatin immunoprecipitation assays were used to document in vivo MeCP2 binding to promoter regions of candidate target genes. Results In the mutants, several hundred genes showed altered expression levels. Twice as many were increased than decreased, and only 27 genes were differentially expressed at more than one time point. The number of misregulated genes was 30% lower in mice with an exon 3 deletion (Mecp2tm1.1Jae) than in mice with a larger deletion (Mecp2tm1.1Bird). Between the mutants, few misregulated genes overlapped at each time point. Real-time quantitative RT-PCR assays validated increased transcript levels for four genes: Irak1, interleukin-1 receptor-associated kinase 1; Fxyd1, phospholemman, associated with Na, K-ATPase; Reln, encoding reelin, an extracellular signaling molecule essential for neuronal lamination and synaptic plasticity; and Gtl2/Meg3, an imprinted maternally expressed non-translated RNA that serves as a host gene for C/D box snoRNAs and microRNAs. Chromatin immunoprecipitation assays documented in vivo MeCP2 binding to promoter regions of Fxyd1, Reln, and Gtl2. Conclusions Transcriptional profiling of cerebellum failed to detect significant global changes in Mecp2-mutant mice. Increased transcript levels of Irak1, Fxyd1, Reln, and Gtl2 may contribute to the neuronal dysfunction in MeCP2-deficient mice and individuals with Rett syndrome. Our data provide testable hypotheses for future studies of the regulatory or signaling pathways that these genes act on. A genetic modification design type is where an organism(s) has had genetic material removed, rearranged, mutagenized or added, such as knock out. Computed

Project description:Background MeCP2, methyl-CpG-binding protein 2, binds to methylated cytosines at CpG dinucleotides, as well as to unmethylated DNA, and affects chromatin condensation. MECP2 mutations in females lead to Rett syndrome, a neurological disorder characterized by developmental stagnation and regression, loss of purposeful hand use and speech, stereotypic hand movements, deceleration of brain growth, autonomic dysfunction and seizures. Most mutations occur de novo during spermatogenesis. Located at Xq28, MECP2 is subject to X inactivation, and affected females are mosaic. Rare hemizygous males suffer from a severe congenital encephalopathy. Methods To identify pathways mis-regulated by MeCP2 deficiency, microarray-based global gene expression studies were carried out on cerebellum of Mecp2 mutant mice. We compared transcript levels in mutant/wildtype male sibs of two different MeCP2-deficient mouse models at 2, 4 and 8 weeks of age. Increased transcript levels were evaluated by real-time quantitative RT-PCR. Chromatin immunoprecipitation assays were used to document in vivo MeCP2 binding to promoter regions of candidate target genes. Results In the mutants, several hundred genes showed altered expression levels. Twice as many were increased than decreased, and only 27 genes were differentially expressed at more than one time point. The number of misregulated genes was 30% lower in mice with an exon 3 deletion (Mecp2tm1.1Jae) than in mice with a larger deletion (Mecp2tm1.1Bird). Between the mutants, few misregulated genes overlapped at each time point. Real-time quantitative RT-PCR assays validated increased transcript levels for four genes: Irak1, interleukin-1 receptor-associated kinase 1; Fxyd1, phospholemman, associated with Na, K-ATPase; Reln, encoding reelin, an extracellular signaling molecule essential for neuronal lamination and synaptic plasticity; and Gtl2/Meg3, an imprinted maternally expressed non-translated RNA that serves as a host gene for C/D box snoRNAs and microRNAs. Chromatin immunoprecipitation assays documented in vivo MeCP2 binding to promoter regions of Fxyd1, Reln, and Gtl2. Conclusions Transcriptional profiling of cerebellum failed to detect significant global changes in Mecp2-mutant mice. Increased transcript levels of Irak1, Fxyd1, Reln, and Gtl2 may contribute to the neuronal dysfunction in MeCP2-deficient mice and individuals with Rett syndrome. Our data provide testable hypotheses for future studies of the regulatory or signaling pathways that these genes act on. A genetic modification design type is where an organism(s) has had genetic material removed, rearranged, mutagenized or added, such as knock out. Keywords: genetic_modification_design

Project description:Mutations in methyl-CpG-binding protein 2 (MeCP2), a major epigenetic regulator, are the predominant cause of Rett syndrome, an X-linked neurodevelopmental disorder. We previously found that Mecp2-null microglia are functionally impaired, and that engraftment of wild-type monocytes into the brain of Mecp2-deficient mice attenuates pathology. In this study we show that Mecp2 is expressed in macrophage and monocyte populations throughout the body, and is indispensable for their transcriptional regulation in multiple contexts. We demonstrate that Mecp2-null mice progressively lose or are chronically deficient in several macrophage populations and resident monocytes. Postnatal re-expression of Mecp2 driven by a tamoxifen-inducible CX3CR1 promoter significantly increased the lifespan of otherwise Mecp2-null mice, suggesting that epigenetic regulation of macrophage function by Mecp2 significantly contributes to pathology. RNA-Seq of acutely isolated microglia and peritoneal macrophages (to our knowledge, the first cell-specific RNA-Seq analysis comparing Mecp2-null and wild type cells of any kind) revealed significantly increased transcription of glucocorticoid- and hypoxia-signaling genes in Mecp2-null cells compared to that in their wild-type counterparts, suggesting that Mecp2 functions as a repressor of these pathways. Furthermore, in-vitro and in vivo validation studies demonstrated that the absence of Mecp2 is associated with cell-intrinsic dysfunction of signaling underlying inflammatory activation, suggesting that Mecp2 is important for regulation of specific macrophage gene-expression programs in response to stimuli and stressors. Our findings demonstrate a fundamental role for Mecp2 in the regulation of macrophage functions, which may provide a link to pathologies in Rett syndrome across multiple organs. Mecp2-null microglia and resident peritoneal macrophages from 10-12 week old mice were acutely isolated via AutoMACS, total RNA collected, and analyzed via RNA-Seq to compare for transcriptional differences in microglia and macrophages in the absence of Mecp2.

Dataset Information

Transcription profiling of cerebellum from Mecp2 mutant mice to identify pathways mis-regulated by MeCP2 deficiency in Rett Syndrome

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets