Autism and other neuropsychiatric symptoms are prevalent in individuals with MeCP2 duplication syndrome

Abstract

Objective: There have been no objective assessments to determine whether boys with MECP2 duplication have autism or whether female carriers manifest phenotypes. This study characterizes the clinical and neuropsychiatric phenotypes of affected boys and carrier females.

Methods: Eight families (9 males and 9 females) with MECP2 duplication participated. A detailed history, physical examination, electroencephalogram, developmental evaluation, Autism Diagnostic Observation Schedule, and Autism Diagnostic Interview-Revised were performed for each boy. Carrier females completed the Symptom Checklist-90-R, Wechsler Abbreviated Scale of Intelligence, Broad Autism Phenotype Questionnaire, and detailed medical and mental health histories. Size and gene content of each duplication were determined by array comparative genome hybridization. X-chromosome inactivation patterns were analyzed using leukocyte DNA. MECP2 and IRAK1 RNA levels were quantified from lymphoblast cell lines, and western blots were performed to assess MeCP2 protein levels.

Results: All of the boys demonstrated mental retardation and autism. Poor expressive language, gaze avoidance, repetitive behaviors, anxiety, and atypical socialization were prevalent. Female carriers had psychiatric symptoms, including generalized anxiety, depression, and compulsions that preceded the birth of their children. The majority exhibited features of the broad autism phenotype and had higher nonverbal compared to verbal reasoning skills.

Interpretation: Autism is a defining feature of the MECP2 duplication syndrome in boys. Females manifest phenotypes despite 100% skewing of X-inactivation and normal MECP2 RNA levels in peripheral blood. Analysis of the duplication size, MECP2 and IRAK1 RNA levels, and MeCP2 protein levels revealed that most of the traits in affected boys are likely due to the genomic region spanning of MECP2 and IRAK1. The phenotypes observed in carrier females may be secondary to tissue-specific dosage alterations and require further study. Ann Neurol 2009;66:771-782.

Figure 1

Pedigrees for eight of the families included in this study. The pedigree for subject one is not included since he was adopted. Maternal subject numbers are purposefully excluded to preserve anonymity. Standard pedigree symbols are used.

Figure 2

Genomic region duplicated in subjects with MECP2 duplications determined by oligonucleotide array CGH. The smallest region of overlap (SRO) includes the MECP2 and IRAK1 genes and covers 231 kb. Solid red bars represent oligonucleotide probes for which the mean normalized log₂ (Cy5/Cy3) ratio of the CGH signal, amid a 5 kb window, indicated a gain consistent with duplication. Subject 9 presented log₂ values suggestive of both triplicated (in blue) and duplicated regions; the triplication does not involve the MECP2 gene. The graphical normalized data for each patient was obtained by inputting the most distal and proximal oligonucleotide genomic probe coordinates into the custom track at UCSC site http://genome.ucsc.edu/cgi-bin/hgGateway. Positions are given relative to NCBI build 35 for the X chromosome.

Figure 3

Quantification of MECP2 and IRAK1 mRNA levels as measured by quantitative real-time PCR and normalized to GAPDH. A. MECP2 mRNA levels are 2 to 6 fold higher and IRAK1 mRNA levels are 2 to 7 fold higher in boys with Xq28 duplication compared to control boys. B. MECP2 and IRAK1 mRNA levels are not altered in carrier vs. control females.

Figure 4

Analysis of MeCP2 protein levels as measured by Western blot and normalized to histone H3. A. Western blot demonstrating MeCP2 protein (upper half of blot, MW ∼64 kDa) and histone H3 protein (lower half of blot, MW ∼17 kDa) levels in boys with MECP2 duplication compared with control boys. B. MeCP2 protein levels are ∼1.5 to 1.7-fold greater in the lymphoblast cell lines of boys with MECP2 duplication compared with control boys. MeCP2 protein levels are normalized to total histone H3 protein levels.

Figure 5

X-chromosome inactivation (XCI) patterns in females with MECP2 duplication. Panel A demonstrates control DNA with random XCI; Panel B demonstrates control DNA with 100% skewed XCI; The remaining panels demonstrate DNA from MECP2 duplication carriers with 100% skewed XCI. In each pair, the top panel represents treatment with HpaII enzyme and the bottom panel is without enyme. R = number of repeats. Panel C is subject 10, D is subject 11, E is subject 12, F is subject 13, G is subject 13's mother, H is subject 14, I is subject 15, and J is subject 18.