Reduced axonal diameter of peripheral nerve fibres in a mouse model of Rett syndrome

Rett syndrome (RTT) is a neurological disorder characterized by motor and cognitive impairment, autonomic dysfunction and a loss of purposeful hand skills. In the majority of cases, typical RTT is caused by de novo mutations in the X-linked gene, MECP2. Alterations in the structure and function of neurons within the central nervous system of RTT patients and Mecp2-null mouse models are well established. In contrast, few studies have investigated the effects of MeCP2-deficiency on peripheral nerves. In this study, we conducted detailed morphometric as well as functional analysis of the sciatic nerves of symptomatic adult female Mecp2+/- mice. We observed a significant reduction in the mean diameter of myelinated nerve fibers in Mecp2+/- mice. In myelinated fibers, mitochondrial densities per unit area of axoplasm were significantly altered in Mecp2+/- mice. However, conduction properties of the sciatic nerve of Mecp2 knockout mice were not different from control. These subtle changes in myelinated peripheral nerve fibers in heterozygous Mecp2 knockout mice could potentially explain some RTT phenotypes

Specific binding of the methyl binding domain protein 2 at the BRCA1-NBR2 locus

The methyl-CpG binding domain (MBD) proteins are key molecules in the interpretation of DNA methylation signals leading to gene silencing. We investigated their binding specificity at the constitutively methylated region of a CpG island containing the bidirectional promoter of the Breast cancer predisposition gene 1, BRCA1, and the Near BRCA1 2 (NBR2) gene. In HeLa cells, quantitative chromatin immunoprecipitation assays indicated that MBD2 is associated with the methylated region, while MeCP2 and MBD1 were not detected at this locus. MBD2 depletion (∼90%), mediated by a transgene expressing a small interfering RNA (siRNA), did not induce MeCP2 or MBD1 binding at the methylated area. Furthermore, the lack of MBD2 at the BRCA1-NBR2 CpG island is associated with an elevated level of NBR2 transcripts and with a significant reduction of induced-DNA-hypomethylation response. In MBD2 knockdown cells, transient expression of a Mbd2 cDNA, refractory to siRNA-mediated decay, shifted down the NBR2 mRNA level to that observed in unmodified HeLa cells. Variations in MBD2 levels did not affect BRCA1 expression despite its stimulation by DNA hypomethylation. Collectively, our data indicate that MBD2 has specific targets and its presence at these targets is indispensable for gene repression

A role for transcriptional repressor methyl-CpG-binding protein 2 and plasticity-related gene serum- and glucocorticoid-inducible kinase 1 in the induction of inflammatory pain states

Activity-dependent changes in neurons of the rat superficial dorsal horn are crucial for the induction and maintenance of neuropathic and inflammatory pain states. To identify the molecular mechanisms underlying this sensitization of superficial dorsal horn neurons, we undertook a genome-wide microarray profiling of dorsal horn gene transcripts at various times after induction of peripheral inflammation of the rat ankle joint. At early time points, upregulation of gene expression dominated, but by 7 d, downregulation was predominant. Two to 24 h after inflammation, we identified a small number of highly upregulated transcripts previously shown to be repressed by the Methyl-CpG-binding protein 2 (MeCP2), including serum-and glucocorticoid-inducible kinase (SGK1) and FK 506 binding protein 5, genes known to be important in experience-dependent plasticity. A decrease in expression of SIN3A, a corepressor in the MeCP2 silencing complex, was also found after inflammation. Phosphorylation of MeCP2 regulates activity-dependent gene transcription, and crucially we found that MeCP2 was phosphorylated in lamina I projection neurons 1 h after induction of peripheral inflammation. Lamina I projection neurons have been shown to be essential for the development of most pain states. SGK1 protein was also localized, in part, to lamina I projection neurons, and its expression in the superficial dorsal horn increased after inflammation. Furthermore, antisense knock-down of SGK1 delayed the onset of inflammatory hyperalgesia by 24 h at least. Our results uncover an unexpected complexity in the regulation of gene expression, including the modulation of transcriptional repression, that accompanies development and maintenance of an inflammatory pain state

Proteomic analyses reveal misregulation of LIN28 expression and delayed timing of glial differentiation in human iPS cells with MECP2 loss-of-function.

Rett syndrome (RTT) is a pervasive developmental disorder caused by mutations in MECP2. Complete loss of MECP2 function in males causes congenital encephalopathy, neurodevelopmental arrest, and early lethality. Induced pluripotent stem cell (iPSC) lines from male patients harboring mutations in MECP2, along with control lines from their unaffected fathers, give us an opportunity to identify some of the earliest cellular and molecular changes associated with MECP2 loss-of-function (LOF). We differentiated iPSC-derived neural progenitor cells (NPCs) using retinoic acid (RA) and found that astrocyte differentiation is perturbed in iPSC lines derived from two different patients. Using highly stringent quantitative proteomic analyses, we found that LIN28, a gene important for cell fate regulation and developmental timing, is upregulated in mutant NPCs compared to WT controls. Overexpression of LIN28 protein in control NPCs suppressed astrocyte differentiation and reduced neuronal synapse density, whereas downregulation of LIN28 expression in mutant NPCs partially rescued this synaptic deficiency. These results indicate that the pathophysiology of RTT may be caused in part by misregulation of developmental timing in neural progenitors, and the subsequent consequences of this disruption on neuronal and glial differentiation

CNV and nervous system diseases - what's new?

Several new genomic disorders caused by copy number variation (CNV) of genes whose dosage is critical for the physiological function of the nervous system have been recently identified. Dup(7)(q11.23) patients carry duplications of the genomic region deleted in Williams-Beuren syndrome, they are characterized by prominent speech delay. The phenotypes of Potocki-Lupski syndrome and MECP2 duplication syndrome were neuropsychologically examined in detail, which revealed autism as an endophenotype and a prominent behavioral feature of these disorders. Tandem duplication of LMNB1 was reported to cause adult-onset autosomal dominant leukodystrophy. PAFAH1B1/LIS1 and YWHAE, which were deleted in isolated lissencephaly (PAFAH1B1/LIS1 alone) and Miller-Dieker syndrome (both genes), were found to be duplicated in patients with developmental delay. Finally, two novel microdeletion syndromes affecting 17q21.31 and 15q13.3, as well as their reciprocal duplications, were also identified. In this review, we provide an overview of the phenotypic manifestation of these syndromes and the rearrangements causing them. Copyright (C) 2009 S. Karger AG, Base

Loss of MECP2 Leads to Activation of P53 and Neuronal Senescence.

To determine the role for mutations of MECP2 in Rett syndrome, we generated isogenic lines of human induced pluripotent stem cells, neural progenitor cells, and neurons from patient fibroblasts with and without MECP2 expression in an attempt to recapitulate disease phenotypes in vitro. Molecular profiling uncovered neuronal-specific gene expression changes, including induction of a senescence-associated secretory phenotype (SASP) program. Patient-derived neurons made without MECP2 showed signs of stress, including induction of P53, and senescence. The induction of P53 appeared to affect dendritic branching in Rett neurons, as P53 inhibition restored dendritic complexity. The induction of P53 targets was also detectable in analyses of human Rett patient brain, suggesting that this disease-in-a-dish model can provide relevant insights into the human disorder

An enhanced CRISPR repressor for targeted mammalian gene regulation.

The RNA-guided endonuclease Cas9 can be converted into a programmable transcriptional repressor, but inefficiencies in target-gene silencing have limited its utility. Here we describe an improved Cas9 repressor based on the C-terminal fusion of a rationally designed bipartite repressor domain, KRAB-MeCP2, to nuclease-dead Cas9. We demonstrate the system's superiority in silencing coding and noncoding genes, simultaneously repressing a series of target genes, improving the results of single and dual guide RNA library screens, and enabling new architectures of synthetic genetic circuits

Rett Syndrome: Revised diagnostic criteria and nomenclature

Objective: Rett syndrome (RTT) is a severe neurodevelopmental disease that affects approximately 1 in 10,000 live female births and is often caused by mutations in Methyl-CpG-binding protein 2 (MECP2). Despite distinct clinical features, the accumulation of clinical and molecular information in recent years has generated considerable confusion regarding the diagnosis of RTT. The purpose of this work was to revise and clarify 2002 consensus criteria for the diagnosis of RTT in anticipation of treatment trials. Method: RettSearch members, representing the majority of the international clinical RTT specialists, participated in an iterative process to come to a consensus on a revised and simplified clinical diagnostic criteria for RTT. Results: The clinical criteria required for the diagnosis of classic and atypical RTT were clarified and simplified. Guidelines for the diagnosis and molecular evaluation of specific variant forms of RTT were developed. Interpretation These revised criteria provide clarity regarding the key features required for the diagnosis of RTT and reinforce the concept that RTT is a clinical diagnosis based on distinct clinical criteria, independent of molecular findings. We recommend that these criteria and guidelines be utilized in any proposed clinical research