Dataset Information

Altered microtubule dynamics and vesicular transport in mouse and human MeCP2-deficient astrocytes.

ABSTRACT: Rett syndrome (RTT) is a rare X-linked neurodevelopmental disorder, characterized by normal post-natal development followed by a sudden deceleration in brain growth with progressive loss of acquired motor and language skills, stereotypic hand movements and severe cognitive impairment. Mutations in the methyl-CpG-binding protein 2 (MECP2) cause more than 95% of classic cases. Recently, it has been shown that the loss of Mecp2 from glia negatively influences neurons in a non-cell-autonomous fashion, and that in Mecp2-null mice, re-expression of Mecp2 preferentially in astrocytes significantly improved locomotion and anxiety levels, restored respiratory abnormalities to a normal pattern and greatly prolonged lifespan compared with globally null mice. We now report that microtubule (MT)-dependent vesicle transport is altered in Mecp2-deficient astrocytes from newborn Mecp2-deficient mice compared with control wild-type littermates. Similar observation has been made in human MECP2 p.Arg294* iPSC-derived astrocytes. Importantly, administration of Epothilone D, a brain-penetrant MT-stabilizing natural product, was found to restore MT dynamics in Mecp2-deficient astrocytes and in MECP2 p.Arg294* iPSC-derived astrocytes in vitro. Finally, we report that relatively low weekly doses of Epothilone D also partially reversed the impaired exploratory behavior in Mecp2(308/y) male mice. These findings represent a first step toward the validation of an innovative treatment for RTT.

SUBMITTER: Delepine C

PROVIDER: S-EPMC4690499 | biostudies-literature | 2016 Jan

REPOSITORIES: biostudies-literature

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Altered microtubule dynamics and vesicular transport in mouse and human MeCP2-deficient astrocytes.

Delépine Chloé C Meziane Hamid H Nectoux Juliette J Opitz Matthieu M Smith Amos B AB Ballatore Carlo C Saillour Yoann Y Bennaceur-Griscelli Annelise A Chang Qiang Q Williams Emily Cunningham EC Dahan Maxime M Duboin Aurélien A Billuart Pierre P Herault Yann Y Bienvenu Thierry T

Human molecular genetics 20151124 1

Rett syndrome (RTT) is a rare X-linked neurodevelopmental disorder, characterized by normal post-natal development followed by a sudden deceleration in brain growth with progressive loss of acquired motor and language skills, stereotypic hand movements and severe cognitive impairment. Mutations in the methyl-CpG-binding protein 2 (MECP2) cause more than 95% of classic cases. Recently, it has been shown that the loss of Mecp2 from glia negatively influences neurons in a non-cell-autonomous fashio ...[more]

PMID: 26604147

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Project description:Schizophrenia (SCZ) is a common, disabling mental illness with high heritability but complex, poorly understood genetic etiology. As the first phase of a genomic convergence analysis of SCZ, we generated 16.7 billion nucleotides of short read, shotgun sequences of cDNA from post-mortem cerebellar cortices of 14 patients and six matched controls. A rigorous analysis pipeline was developed for analysis of digital gene expression studies. Sequences aligned to approximately 33,200 transcripts in each sample, with average coverage of 450 reads per gene. Following adjustments for confounding clinical, sample and experimental sources of variation, 215 genes differed significantly in expression between cases and controls. Golgi apparatus, vesicular transport, membrane association, Zinc binding and regulation of transcription were over-represented among differentially expressed genes. Twenty three genes with altered expression and involvement in presynaptic vesicular transport, Golgi function and GABAergic neurotransmission define a unifying molecular hypothesis for dysfunction in cerebellar cortex in SCZ. Experiment Overall Design: 16.7 billion nucleotides of shotgun, full length cDNA sequence data were generated using Illumina Genome Analyzer platforms with sequencing-by-synthesis (SBS) chemistry from 20 mRNA samples (Lister et al., 2008; Morin et al., 2008)(Mortazvi et al., 2008). mRNA samples were isolated post-mortem from the lateral hemispheres of the cerebellar cortices of 14 patients with SCZ and 6 control individuals (Paz et al., 2006; Bullock et al., In Press)(Table 1). Unrelated subjects were chosen to facilitate sampling of genetic heterogeneity (Freimer and Sabatti, 2004; McClellan et al., 2007). Cases and controls were approximately matched for age (cases, 45.2 + 11.8 years; controls 41.3 + 9.2 years), sex (all male), race, post-mortem interval (cases, 12.2 + 5.0 hours; controls 17.7 + 3.3 hours), cause of death, autopsy brain pH (cases, 6.54 + 0.19; controls 6.46 + 0.10) and RNA integrity number (cases, 8.06 + 0.53; controls, 7.87 + 0.41) (Table 1). 12.5 – 38.7 million, high quality sequences of length 32 – 36 bp were generated per sample (Table 2). Sequences were aligned to the human genome and RefSeq transcript databases using the algorithm GMAP, which allowed < 2 (< 6%) mismatches (Wu and Watanabe, 2005). There was little intra-sample variability in the number of sequences aligned to each locus from run-to-run or instrument-to-instrument (Fig. 1A; all source coefficient of variation 3.4%). 43.5 + 6.7% of sequences aligned to a transcript and 69.4 + 9.6% to the genome, evidence that annotation of mRNA isoforms in Homo sapiens is incomplete (Birney et al., 2007) (Table 2). 91% of alignments were unique (Table 2). Reads aligning to more than one location contained repetitive, paralogous, polymorphic or low complexity sequences and primarily mapped to untranslated regions or highly polymorphic gene families, such as major histocompatibility genes (Sugarbaker et al., 2008). Unmapped sequences did not align to mitochondrial or 1879 viral genomes, offering negative evidence of chronic viral etiology for SCZ in these patients.

Project description:Rett syndrome (RTT) is a neurodevelopmental disorder that represents the most common genetic cause of severe intellectual disability in females. Most patients carry mutations in the X-linked MECP2 gene, coding for the methyl-CpG-binding protein 2 (MeCP2), originally isolated as an epigenetic transcriptional factor able to bind methylated DNA and repress transcription. Recent data implicated a role for glia in RTT, showing that astrocytes express Mecp2 and that its deficiency affects their ability to support neuronal maturation by non-cell autonomous mechanisms. To date, some molecular, structural and functional alterations have been attributed to Mecp2 null astrocytes, but how they evolve over time and whether they follow a spatial heterogeneity are two aspects which deserve further investigations. In this study, we assessed cytoskeletal features of astrocytes in Mecp2 deficient brains by analyzing their arbor complexity and processes in reconstructed GFAP+ cells at different ages, corresponding to peculiar stages of the disorder, and in different cerebral regions (motor and somatosensory cortices and CA1 layer of hippocampus). Our findings demonstrate the presence of defects in Mecp2 null astrocytes that worsen along disease progression and strictly depend on the brain area, highlighting motor and somatosensory cortices as the most affected regions. Of relevance, astrocyte cytoskeleton is impaired also in the somatosensory cortex of symptomatic heterozygous animals, with Mecp2 + astrocytes showing slightly more pronounced defects with respect to the Mecp2 null cells, emphasizing the importance of non-cell autonomous effects. We reported a temporal correlation between the progressive thinning of layer I and the atrophy of astrocytes, suggesting that their cytoskeletal dysfunctions might contribute to cortical defects. Considering the reciprocal link between morphology and function in astrocytes, we analyzed the effect of Mecp2 deficiency on the expression of selected astrocyte-enriched genes, which describe typical astrocytic features. qRT-PCR data corroborated our results, reporting an overall decrement of gene expression, which is area and age-dependent. In conclusion, our data show that Mecp2 deficiency causes structural and molecular alterations in astrocytes, which progress along with the severity of symptoms and diversely occur in the different cerebral regions, highlighting the importance of considering heterogeneity when studying astrocytes in RTT.

Dataset Information

Altered microtubule dynamics and vesicular transport in mouse and human MeCP2-deficient astrocytes.

Publications

Altered microtubule dynamics and vesicular transport in mouse and human MeCP2-deficient astrocytes.

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