Article abstract-Background: Rett syndrome (RTT) is a neurodevelopmental disorder caused by mutations in the X-linked methyl CpG binding protein 2 (MeCP2) gene. Methods: One hundred sixteen patients with classical and atypical RTT were studied for mutations of the MeCP2 gene by using DHPLC and direct sequencing. Results: Causative mutations in the MeCP2 gene were identified in 63% of patients, representing a total of 30 different mutations. Mutations were identified in 72% of patients with classical RTT and one third of atypical cases studied (8 of 25). The authors found 17 novel mutations, including a complex gene rearrangement found in one individual involving two deletions and a duplication. The duplication was identical to a region within the 3Ј untranslated region (UTR), and represents the first report of involvement of the 3Ј UTR in RTT. The authors also report the identification of MeCP2 mutations in two males; a Klinefelter's male with classic RTT (T158M) and a hemizygous male infant with a Xq27-28 inversion and a novel 32 bp frameshift deletion [1154(del32)]. Studies examining the relationship between mutation type, X-inactivation status, and severity of clinical presentation found significant differences in clinical presentation between different types of mutations. Mutations in the amino-terminus were significantly correlated with a more severe clinical presentation compared with mutations closer to the carboxyl-terminus of MeCP2. Skewed X-inactivation patterns were found in two asymptomatic carriers of MeCP2 mutations and six girls diagnosed with either atypical or classical RTT. Conclusion: This patient series confirms the high frequency of MeCP2 gene mutations causative of RTT in females and provides data concerning the molecular basis for clinical variability (mutation type and position and X-inactivation patterns).

Deficiencies in the activity of cytochrome c oxidase (COX), the terminal enzyme in the respiratory chain, are a frequent cause of autosomal recessive mitochondrial disease in infants. These patients are clinically and genetically heterogeneous, and all defects so far identified in this group have been found in genes coding for accessory proteins that play important roles in the assembly of the COX holoenzyme complex. Many patients, however, remain without a molecular diagnosis. We have used a panel of retroviral vectors expressing human COX assembly factors in these patients to identify the molecular basis for the COX deficiency by functional complementation. Here we show that overexpression of COX15, a protein involved in the synthesis of heme A, the heme prosthetic group for COX, can functionally complement the isolated COX deficiency in fibroblasts from a patient with fatal, infantile hypertrophic cardiomyopathy. Mutation analysis of COX15 in the patient identified a missense mutation (C700T) on one allele, changing a conserved arginine to tryptophan (R217W), and a splice-site mutation in intron 3 on the other allele (C447-3G), resulting in a deletion of exon 4. This splicing error introduces a frameshift and a premature stop codon, resulting in an unstable mRNA and, likely, a null allele. Mitochondrial heme A content was reduced in the patient's heart and fibroblast mitochondria, and levels of heme O were increased in the patient's heart. COX activity and the total amount of fully assembled enzyme were reduced by 50%-70% in patient fibroblasts. Expression of COX15 increased heme A content and rescued COX activity. These results suggest that reduced availability of heme A stalls the assembly of COX. This study establishes COX15 as an additional cause, along with SCO2, of fatal infantile, hypertrophic cardiomyopathy associated with isolated COX deficiency.

We have identified a 15-bp microdeletion in a highly conserved region of the mitochondrially encoded gene for cytochrome c oxidase (COX) subunit III in a patient with severe isolated COX deficiency and recurrent myoglobinuria. The mutant mitochondrial DNA (mtDNA) comprised 92% of the mtDNA in muscle and 0.7% in leukocytes. Immunoblots and immunocytochemistry suggested a lack of assembly or instability of the complex. Microdissected muscle fibres revealed significantly higher portions of mutant mtDNA in COX-negative than in COX-positive fibres. This represents the first case of isolated COX deficiency to be defined at the molecular level.

Rett syndrome is a neurodevelopmental disorder of early postnatal brain growth in girls. Patients show a normal neonatal period with subsequent developmental regression and a loss of acquired skills (communication and motor skills), deceleration of head growth, and development of typical hand stereotypies. Recent studies have shown that mutations in the X-linked methyl CpG binding protein 2 gene (MeCP2) cause most typical cases of Rett syndrome. The MeCP2 gene encodes a protein that binds methylated cytosine residues of CpG dinucleotides and mediates, with histone deacetylases and transcriptional repressors, the transcription "silencing" of other genes. Girls with Rett syndrome exhibit mosaic expression for the MeCP2 defect at the cellular level, with most patients showing random X-inactivation and approximately equal numbers of cells expressing the normal MeCP2 gene and the mutated MeCP2 gene. In rare cases, females with a MeCP2 mutation escape phenotypic expression of the disorder because of nonrandom X-inactivation and the preferential inactivation of the mutated MeCP2 allele. Nonrandom patterns of X-inactivation may also contribute to the clinical variability often seen in girls with Rett syndrome. The spectrum of clinical phenotype caused by MeCP2 mutations is wide, including milder "preserved speech" variants, the severe congenital Rett variant, and a subset of X-linked recessive mental retardation in boys. Studies have shown that atypical and classical Rett syndrome can caused by the same MeCP2 mutations, indicating clinical phenotype is variable even among girls with the same MeCP2 mutation. The relationship between type of MeCP2 mutation, X-inactivation status, and clinical phenotype of Rett syndrome is complex and likely involves other environmental and polygenic modifiers.

Rett's Syndrome (RTT) is a neurodevelopmental disorder resulting from mutation in the mecp2 gene that encodes methyl CpG binding protein 2, a transcriptional repressor. Because this disease primarily affects neurons, tissue is not available during active disease. We used the olfactory system as a model to investigate abnormalities in neuronal development in RTT, because olfactory receptor neurons (ORNs) are replaced throughout life by ongoing postnatal neurogenesis. Thus, even in the adult, the olfactory epithelium contains neurons at various developmental stages. We obtained biopsies of nasal epithelium containing ORNs from RTT patients and age-matched controls to study the status of the neuronal population using antibodies to stage-specific developmental markers. There were no postprocedure complications. Compared with age-matched controls, there were far fewer mature ORNs, as defined by olfactory marker protein expression, and significantly greater numbers of immature neuron-specific tubulin-positive ORNs present. In RTT biopsies, olfactory marker protein-positive neurons displayed abnormal structure. These results suggest that dysfunction of MeCP2 results in decreased survival of mature ORNs with a compensatory increase in neurogenesis, or a failure of immature neurons to mature. Our study indicates that olfactory biopsies provide a method to study neuronal developmental diseases in adults and children.