Evidence for population variation in TSC1 and TSC2 gene expression.

BACKGROUND: Tuberous sclerosis complex (TSC) is an autosomal dominant neurogenetic disorder caused by mutations in one of two genes, TSC1 or TSC2, which encode the proteins hamartin and tuberin, respectively 123. Common features of TSC include intractable epilepsy, mental retardation, and autistic features. TSC is associated with specific brain lesions, including cortical tubers, subependymal nodules and subependymal giant cell astrocytomas. In addition, this disease frequently produces characteristic tumors, termed hamartomas, in the kidneys, heart, skin, retina, and lungs. Disease severity in TSC can be quite variable and is not determined by the primary mutation alone. In fact, there is often considerable variability in phenotype within single families, where all affected individuals carry the same mutation. Factors suspected to influence phenotype in TSC include the specific primary mutation, random occurrence of second-hit somatic mutations, mosaicism, "modifying genes", and environmental factors. In addition to these factors, we hypothesize that differences in mRNA expression from the non-mutated TSC allele, or possibly from the mutated allele, play a part in modifying disease severity. Common genetic variants that regulate mRNA expression have previously been shown to play important roles in human phenotypic variability, including disease susceptibility. A prediction based on this idea is that common regulatory variants that influence disease severity in TSC should be detectable in non-affected individuals. METHODS: A PCR/primer extension assay was used to measure allele specific expression of TSC1 and TSC2 mRNAs in leukocytes isolated from normal volunteers. This assay can be used to measure "allelic expression imbalance" (AEI) in individuals by making use of heterozygous "marker" single nucleotide polymorphisms (SNPs) located within their mRNA. RESULTS: In this study we show for the first time that TSC1 and TSC2 genes exhibit allele-specific differences in mRNA expression in blood leukocytes isolated from normal individuals. CONCLUSIONS: These results support the possibility that allele-specific variation in TSC mRNA expression contributes to the variable severity of symptoms in TSC patients.

Abnormalities of cell packing density and dendritic complexity in the MeCP2 A140V mouse model of Rett syndrome/X-linked mental retardation.

BACKGROUND: Rett syndrome (RTT), a common cause of mental retardation in girls, is associated with mutations in the MECP2 gene. Most human cases of MECP2 mutation in girls result in classical or variant forms of RTT. When these same mutations occur in males, they often present as severe neonatal encephalopathy. However, some MECP2 mutations can also lead to diseases characterized as mental retardation syndromes, particularly in boys. One of these mutations, A140V, is a common, recurring missense mutation accounting for about 0.6% of all MeCP2 mutations and ranking 21st by frequency. It has been described in familial X-linked mental retardation (XLMR), PPM- X syndrome (Parkinsonism, Pyramidal signs, Macroorchidism, X-linked mental retardation) and in other neuropsychiatric syndromes. Interestingly, this mutation has been reported to preserve the methyl-CpG binding function of the MeCP2 protein while compromising its ability to bind to the mental retardation associated protein ATRX. RESULTS: We report the construction and initial characterization of a mouse model expressing the A140V MeCP2 mutation. These initial descriptive studies in male hemizygous mice have revealed brain abnormalities seen in both RTT and mental retardation. The abnormalities found include increases in cell packing density in the brain and a significant reduction in the complexity of neuronal dendritic branching. In contrast to some MeCP2 mutation mouse models, the A140V mouse has an apparently normal lifespan and normal weight gain patterns with no obvious seizures, tremors, breathing difficulties or kyphosis. CONCLUSION: We have identified various neurological abnormalities in this mouse model of Rett syndrome/X-linked mental retardation which may help to elucidate the manner in which MECP2 mutations cause neuronal changes resulting in mental retardation without the confounding effects of seizures, chronic hypoventilation, or other Rett syndrome associated symptoms.

Reduced neuronal size and mTOR pathway activity in the Mecp2 A140V Rett syndrome mouse model.

Rett syndrome (RTT) is a neurodevelopmental disorder caused by mutation in the X-linked MECP2 gene, encoding methyl-CpG-binding protein 2. We have created a mouse model ( Mecp2 A140V "knock-in" mutant) expressing the recurrent human MECP2 A140V mutation linked to an X-linked mental retardation/Rett syndrome phenotype. Morphological analyses focused on quantifying soma and nucleus size were performed on primary hippocampus and cerebellum granule neuron (CGN) cultures from mutant ( Mecp2A140V/y) and wild type ( Mecp2+/y) male mice. Cultured hippocampus and cerebellar granule neurons from mutant animals were significantly smaller than neurons from wild type animals. We also examined soma size in hippocampus neurons from individual female transgenic mice that express both a mutant  (maternal allele) and a wild type Mecp2 gene linked to an eGFP transgene (paternal allele). In cultures from such doubly heterozygous female mice, the size of neurons expressing the mutant (A140V) allele also showed a significant reduction compared to neurons expressing wild type MeCP2, supporting a cell-autonomous role for MeCP2 in neuronal development. IGF-1 (insulin growth factor-1) treatment of neuronal cells from Mecp2 mutant mice rescued the soma size phenotype. We also found that Mecp2  mutation leads to down-regulation of the mTOR signaling pathway, known to be involved in neuronal size regulation. Our results suggest that i) reduced neuronal size is an important in vitro cellular phenotype of Mecp2 mutation in mice, and ii) MeCP2 might play a critical role in the maintenance of neuronal structure by modulation of the mTOR pathway. The definition of a quantifiable cellular phenotype supports using neuronal size as a biomarker in the development of a high-throughput, in vitro assay to screen for compounds that rescue small neuronal phenotype ("phenotypic assay").

Skewed allele-specific expression of the NF1 gene in normal subjects: a possible mechanism for phenotypic variability in neurofibromatosis type 1.

Neurofibromatosis type 1 is an autosomal dominant disorder characterized by neurocutaneous abnormalities, learning disabilities, and attention-deficit disorder. Neurofibromatosis type 1 symptom severity can be highly variable even within families where all affected members carry the same mutation. We hypothesized that variation in the expression of the normal NF1 allele may be a mechanism that participates in producing variable phenotypes. We performed allelic expression imbalance assays on healthy control individuals to estimate the prevalence of skewed allelic expression of the NF1 gene. Approximately 30% of individuals in our sample population showed significant skewing of allelic expression away from the expected 50:50 ratio, indicating that differential regulation of the NF1 alleles occurs in a high proportion of individuals. Differences of up to 25% in allele-specific expression of the NF1 alleles were identified. In individuals with Neurofibromatosis type 1, who carry a mutant allele (haploinsufficient), this degree of expression skewing may be sufficient to modulate the phenotype.

Autosomal recessive spastic ataxia of Charlevoix-Saguenay: compound heterozygotes for nonsense mutations of the SACS gene.

Mutations of the SACS gene have been reported in patients with autosomal recessive spastic ataxia of Charlevoix-Saguenay from Canada (Quebec), Tunisia, Japan, Turkey, Belgium, Italy, Spain, the Netherlands, and Germany. Features that distinguish autosomal recessive spastic ataxia of Charlevoix-Saguenay from other recessive ataxias include sensory motor polyneuropathy and hypermyelinated retinal nerve fibers. We describe the clinical, electrophysiological, and radiological features in 2 white American siblings diagnosed with autosomal recessive spastic ataxia of Charlevoix-Saguenay. The 2 affected children are compound heterozygotes for nonsense mutations of the SACS gene (c. 3484 G>T, p. E 1162 X; and c. 11,707 C>T, p. R 3903 X). We have measured allele-specific SACS mRNA abundance in peripheral blood and show that these specific mutant mRNAs are not degraded. We suggest that in children with early onset cerebellar ataxia and spasticity, ophthalmological examination and nerve conduction testing may guide genetic testing.