Agenesis of corpus callosum,ocular, and skeletal anomalies (X-linked dominant aicardi's syndrome) in a girl with balanced X/3 translocation

Summary Aicardi's syndrome, which is characterized by agenesis of the corpus callosum, specific chorioretinal abnormalities, and defects of vertebrae and ribs, is considered a probable X-linked dominant trait with male lethality. All features of this syndrome were seen in a girl with a de novo balanced X/3 translocation (46,X,t(X;3)(p22;q12)). It is hypothesized that the clinical picture is the consequence of chromosome breakage within the Aicardi locus. Then, unusual X-inactivation patterns in blood and fibroblasts of this patient can be explained by somatic selection against cells with the Aicardi phenotype.     

Non-random X chromosome inactivation in Aicardi syndrome

Most females have random X-chromosome inactivation (XCI), defined as an equal likelihood for inactivation of the maternally- or paternally-derived X chromosome in each cell. Several X-linked disorders have been associated with a higher prevalence of non-random XCI patterns, but previous studies on XCI patterns in Aicardi syndrome were limited by small numbers and older methodologies, and have yielded conflicting results. We studied XCI patterns in DNA extracted from peripheral blood leukocytes of 35 girls with typical Aicardi syndrome (AIC) from 0.25 to 16.42 years of age, using the human androgen receptor assay. Data on 33 informative samples showed non-random XCI in 11 (33%), defined as a >80:20% skewed ratio of one versus the other X chromosome being active. In six (18%) of these, there was a >95:5% extremely skewed ratio of one versus the other X chromosome being active. XCI patterns on maternal samples were not excessively skewed. The prevalence of non-random XCI in Aicardi syndrome is significantly different from that in the general population (p < 0.0001) and provides additional support for the hypothesis that Aicardi syndrome is an X-linked disorder. We also investigated the correlation between X-inactivation patterns and clinical severity and found that non-random XCI is associated with a high neurological composite severity score. Conversely, a statistically significant association was found between random XCI and the skeletal composite score. Correlations between X-inactivation patterns and individual features were made and we found a significant association between vertebral anomalies and random XCI.     

Studies of X inactivation and isodisomy in twins provide further evidence that the X chromosome is not involved in Rett syndrome.

Rett syndrome (RS), a progressive encephalopathy with onset in infancy, has been attributed to an X-linked mutation, mainly on the basis of its occurrence almost exclusively in females and its concordance in female MZ twins. The underlying mechanisms proposed are an X-linked dominant mutation with male lethality, uniparental disomy of the X chromosome, and/or some disturbance in the process of X inactivation leading to unequal distributions of cells expressing maternal or paternal alleles (referred to as a "nonrandom" or "skewed" pattern of X inactivation). To determine if the X chromosome is in fact involved in RS, we studied a group of affected females including three pairs of MZ twins, two concordant for RS and one uniquely discordant for RS. Analysis of X-inactivation patterns confirms the frequent nonrandom X inactivation previously observed in MZ twins but indicates that this is independent of RS. Analysis of 29 RS females reveals not one instance of uniparental X disomy, extending the observations previously reported. Therefore, our findings contribute no support for the hypothesis that RS is an X-linked disorder. Furthermore, the concordant phenotype in most MZ female twins with RS, which has not been observed in female twins with known X-linked mutations, argues against an X mutation.     

De novo interstitial triplication of MECP2 in a girl with neurodevelopmental disorder and random X chromosome inactivation

Loss-of-function mutations of the MECP2 gene are the cause of most cases of Rett syndrome in females, a progressive neurodevelopmental disorder characterized by severe mental retardation, global regression, hand stereotypies, and microcephaly. On the other hand, gain of dosage of this gene causes the MECP2 duplication syndrome in males characterized by severe mental retardation, absence of speech development, infantile hypotonia, progressive spasticity, recurrent infections, and facial dysmorphism. Female carriers of a heterozygous duplication show a skewed X-inactivation pattern which is the most probable cause of the lack of clinical symptoms. In this paper, we describe a girl with a complex de novo copy number gain at Xq28 and non-skewed X-inactivation pattern that causes mental retardation and motor and language delay. This rearrangement implies triplication of the MECP2 and IRAK1 genes, but it does not span other proximal genes located in the common minimal region of patients affected by the MECP2 duplication syndrome. We conclude that the triplication leads to a severe phenotype due to random X-inactivation, while the preferential X chromosome inactivation in healthy carriers may be caused by a negative selection effect of the duplication on some proximal genes like ARD1A or HCFC1.     

Mapping of a further locus for X-linked craniofrontonasal syndrome

Craniofrontonasal syndrome is a rare dysostosis syndrome with an unusual pattern of X-linked inheritance, because males are usually not or less severely affected than females. Previously, a CFNS locus has been localised in Xp22. We report on a haplotype analysis in a German CFNS family, mapping the CFNS locus to the pericentromeric region of the X chromosome. This discrepancy can be explained by locus heterogeneity. Furthermore, random X inactivation could be demonstrated in affected females. The most plausible interpretation for this unusual pattern of X-linked inheritance is metabolic interference. Consequently, we propose that the CFNS gene escapes X inactivation.     

Complex Segmental Duplications Mediate a Recurrent dup(X)(p11.22-p11.23) Associated with Mental Retardation, Speech Delay, and EEG Anomalies in Males and Females

Submicroscopic copy-number variations make a considerable contribution to the genetic etiology of human disease. We have analyzed subjects with idiopathic mental retardation (MR) by using whole-genome oligonucleotide-based array comparative genomic hybridization (aCGH) and identified familial and de novo recurrent Xp11.22-p11.23 duplications in males and females with MR, speech delay, and a peculiar electroencephalographic (EEG) pattern in childhood. The size of the duplications ranges from 0.8–9.2 Mb. Most affected females show preferential activation of the duplicated X chromosome. Carriers of the smallest duplication show X-linked recessive inheritance. All other affected individuals present dominant expression and comparable clinical phenotypes irrespective of sex, duplication size, and X-inactivation pattern. The majority of the rearrangements are mediated by recombination between flanking complex segmental duplications. The identification of common clinical features, including the typical EEG pattern, predisposing genomic structure, and peculiar X-inactivation pattern, suggests that duplication of Xp11.22-p11.23 constitutes a previously undescribed syndrome.     

X-chromosomale Entwicklungsstörungen im weiblichen Geschlecht

In recent years, an increasing number of X?chromosomal genes were found to be mutated in girls with neurodevelopmental disorders (NDDs). This has blurred the traditional line between X?recessive and X?dominant inheritance. Many X?chromosomal NDDs are now characterized by a phenotypic spectrum that encompasses both males and females. To date, the mechanisms which result in variable disease manifestations between genders but also among females are only poorly understood. Various factors such as the nature, localisation and “severity” of the respective underlying mutation, as well as X?inactivation in particular, are assumed to contribute. This article provides an overview of the current knowledge on X?chromosomal NDDs in females. Additionally, several exemplary new X?chromosomal syndromes in females caused by de novo mutations will be described and discussed in more detail.     

High male:female ratio of germ-line mutations: an alternative explanation for postulated gestational lethality in males in X-linked dominant disorders.

In this paper I suggest that a vastly higher rate of de novo mutations in males than in females would explain some, if not most, X-linked dominant disorders associated with a low incidence of affected males. It is the inclusion of the impact of a high ratio of male:female de novo germ-line mutations that makes this model new and unique. Specifically, it is concluded that, if an X-linked disorder results in a dominant phenotype with a significant reproductive disadvantage (genetic lethality), affected females will, in virtually all cases, arise from de novo germ-line mutations inherited from their fathers rather than from their mothers. Under this hypothesis, the absence of affected males is explained by the simple fact that sons do not inherit their X chromosome (normal or abnormal) from their fathers. Because females who are heterozygous for a dominant disorder will be clinically affected and will, in most cases, either be infertile or lack reproductive opportunities, the mutant gene will not be transmitted by them to the next generation (i.e., it will be a genetic lethal). This, not gestational lethality in males, may explain the absence of affected males in most, if not all, of the 13 known X-linked dominant diseases characterized by high ratios of affected female to male individuals. Evidence suggesting that this mechanism could explain the findings in the Rett syndrome is reviewed in detail.     

De novo MECP2 duplications in two females with intellectual disability and unfavorable complete skewed X-inactivation

Xq28 microduplications of MECP2 are a prominent cause of a severe syndromic form of intellectual disability (ID) in males. Females are usually unaffected through near to complete X-inactivation of the aberrant X chromosome (skewing). In rare cases, affected females have been described due to random X-inactivation. Here, we report on two female patients carrying de novo MECP2 microduplications on their fully active X chromosomes. Both patients present with ID and additional clinical features. Mono-allelic expression confirmed complete skewing of X-inactivation. Consequently, significantly enhanced MECP2 mRNA levels were observed. We hypothesize that the cause for the complete skewing is due to a more harmful mutation on the other X chromosome, thereby forcing the MECP2 duplication to become active. However, we could not unequivocally identify such a second mutation by array-CGH or exome sequencing. Our data underline that, like in males, increased MECP2 dosage in females can contribute to ID too, which should be taken into account in diagnostics.     

X-inactivation and human disease: X-linked dominant male-lethal disorders

X chromosome inactivation (XCI) is the process by which the dosage imbalance of X-linked genes between XX females and XY males is functionally equalized. XCI modulates the phenotype of females carrying mutations in X-linked genes, as observed in X-linked dominant male-lethal disorders such as oral-facial-digital type I (OFDI) and microphthalmia with linear skin-defects syndromes. The remarkable degree of heterogeneity in the XCI pattern among female individuals, as revealed by the recently reported XCI profile of the human X chromosome, could account for the phenotypic variability observed in these diseases. Furthermore, the recent characterization of a murine model for OFDI shows how interspecies differences in the XCI pattern between Homo sapiens and Mus musculus result in discrepancies between the phenotypes observed in patients and mice.     

X-linked hydrocephalus: another two families with an L1 mutation

X-linked hydrocephalus is a variable condition caused by mutations in the gene encoding for L1CAM. This gene is located at Xq28. Clinically the spectrum ranges from males with lethal congenital hydrocephalus to mild/moderate mental retardation and spastic paraplegia. Few carrier females show minimal signs of the syndrome. Although most cases are familial, de novo situations have been reported. We report two new families with the syndrome and a L1 mutation. Family 1 has two patients and family 2 a single patient. Clinical diagnosis in all three affected boys was beyond doubt. Prenatal testing through chorionic villus biopsy is possible only with a demonstrated L1 mutation. In lethal sporadic cases neuropathology is very important in order to evaluate for features of the syndrome. We stress the importance of further clinical reports including data on neuropathology and DNA analysis in order to further understand the mechanisms involved in this disorder.     

Skewed X-Chromosome Inactivation Is Common in Fetuses or Newborns Associated with Confined Placental Mosaicism

The inactivation of one X chromosome in females is normally random with regard to which X is inactivated. However, exclusive or almost-exclusive inactivation of one X may be observed in association with some X-autosomal rearrangements, mutations of the XIST gene, certain X-linked diseases, and MZ twinning. In the present study, a methylation difference near a polymorphism in the X-linked androgen-receptor gene was used to investigate the possibility that nonrandom X inactivation is increases in fetuses and newborns that are associated with confined placental mosaicism (CPM) involving an autosomal trisomy. Extreme skewing was observed in 7 (58%) of 12 cases with a meiotic origin of the trisomy, but in none of 10 cases examined with a somatic origin of the trisomy, and in only 1 (4%) of 27 control adult females. In addition, an extremely skewed X-inactivation pattern was observed in 3 of 10 informative cases of female uniparental disomy (UPD) of chromosome 15. This may reflect the fact that a proportion of UPD cases arise by "rescue" of a chromosomally abnormal conceptus and are therefore associated with CPM. A skewed pattern of X inactivation in CPM cases is hypothesized to result from a reduction in the size of the early-embryonic cell pool, because of either poor early growth or subsequent selection against the trisomic cells. Since approximately 2% of pregnancies detected by chorionic villus sampling are associated with CPM, this is likely a significant contributor to both skewed X inactivation observed in the newborn population and the expression of recessive X-linked diseases in females.     

De Novo Loss-of-Function Mutations in USP9X Cause a Female-Specific Recognizable Syndrome with Developmental Delay and Congenital Malformations

Mutations in more than a hundred genes have been reported to cause X-linked recessive intellectual disability (ID) mainly in males. In contrast, the number of identified X-linked genes in which de novo mutations specifically cause ID in females is limited. Here, we report 17 females with de novo loss-of-function mutations in USP9X, encoding a highly conserved deubiquitinating enzyme. The females in our study have a specific phenotype that includes ID/developmental delay (DD), characteristic facial features, short stature, and distinct congenital malformations comprising choanal atresia, anal abnormalities, post-axial polydactyly, heart defects, hypomastia, cleft palate/bifid uvula, progressive scoliosis, and structural brain abnormalities. Four females from our cohort were identified by targeted genetic testing because their phenotype was suggestive for USP9X mutations. In several females, pigment changes along Blaschko lines and body asymmetry were observed, which is probably related to differential (escape from) X-inactivation between tissues. Expression studies on both mRNA and protein level in affected-female-derived fibroblasts showed significant reduction of USP9X level, confirming the loss-of-function effect of the identified mutations. Given that some features of affected females are also reported in known ciliopathy syndromes, we examined the role of USP9X in the primary cilium and found that endogenous USP9X localizes along the length of the ciliary axoneme, indicating that its loss of function could indeed disrupt cilium-regulated processes. Absence of dysregulated ciliary parameters in affected female-derived fibroblasts, however, points toward spatiotemporal specificity of ciliary USP9X (dys-)function.     

Uniparental disomy of the entire X chromosome in a female with Duchenne muscular dystrophy.

Duchenne muscular dystrophy (DMD) is a severe, progressive, X-linked muscle-wasting disorder with an incidence of approximately 1/3,500 male births. Females are also affected, in rare instances. The manifestation of mild to severe symptoms in female carriers of dystrophin mutations is often the result of the preferential inactivation of the X chromosome carrying the normal dystrophin gene. The severity of the symptoms is dependent on the proportion of cells that have inactivated the normal X chromosome. A skewed pattern of X inactivation is also responsible for the clinical manifestation of DMD in females carrying X;autosome translocations, which disrupt the dystrophin gene. DMD may also be observed in females with Turner syndrome (45,X), if the remaining X chromosome carries a DMD mutation. We report here the case of a karyotypically normal female affected with DMD as a result of homozygosity for a deletion of exon 50 of the dystrophin gene. PCR analysis of microsatellite markers spanning the length of the X chromosome demonstrated that homozygosity for the dystrophin gene mutation was caused by maternal isodisomy for the entire X chromosome. This finding demonstrates that uniparental isodisomy of the X chromosome is an additional mechanism for the expression of X-linked recessive disorders. The proband's clinical presentation is consistent with the absence of imprinted genes (i.e., genes that are selectively expressed based on the parent of origin) on the X chromosome.     

Multiple de novo mutations in the MECP2 gene

Rett syndrome is an X-linked dominant disorder that usually arises following a single de novo mutation in the MECP2 gene. Point mutation testing and gene dosage analysis of a cohort of British Rett syndrome patients in our laboratory revealed four females who each had two different de novo causative mutations, presumed to be in cis because the patients showed no deviation from the classical Rett syndrome phenotype. Two of these cases had a point mutation and a small intraexonic deletion, a third had a whole exon deletion and a separate small intraexonic deletion, and a fourth case had a small intraexonic deletion and a large duplication. These findings highlight the necessity to perform both point mutation analysis and exon dosage analysis in such cases, particularly because of the possibility of undetected parental mosaicism and the implications for prenatal diagnosis in future pregnancies. These cases also suggest that the MECP2 gene may be particularly prone to multiple mutation events.     

A new Rett syndrome family consistent with X-linked inheritance expands the X chromosome exclusion map.

Although familial recurrences of Rett syndrome (RTT) comprise only approximately 1% of the reported cases, it is these cases that hold the key for the understanding of the genetic basis of the disorder. Families in which RTT occurs in mother and daughter, aunt and niece, and half sisters are consistent with dominant inheritance and variable expressivity of the phenotype. Recurrence of RTT in sisters is likely due to germ-line mosaicism in one of the parents, rather than to recessive inheritance. The exclusive occurrence of classic RTT in females led to the hypothesis that it is X-linked and may be lethal in males. In an X-linked dominant disorder, unaffected obligate-carrier females would be expected to show nonrandom or skewed inactivation of the X chromosome bearing the mutant allele. We investigated the X chromosome inactivation (XCI) patterns in the female members of a newly identified family with recurrence of RTT in a maternal aunt and a niece. Skewing of XCI is present in the obligate carrier in this family, supporting the hypothesis that RTT is an X-linked disorder. However, evaluation of the XCI pattern in the mother of affected half sisters shows random XCI, suggesting germ-line mosaicism as the cause of repeated transmission in this family. To determine which regions of the X chromosome were inherited concordantly/discordantly by the probands, we genotyped the individuals in the aunt-niece family and two previously reported pairs of half sisters. These combined exclusion-mapping data allow us to exclude the RTT locus from the interval between DXS1053 in Xp22.2 and DXS1222 in Xq22.3. This represents an extension of the previous exclusion map.     

Disruption of the serine/threonine kinase 9 gene causes severe X-linked infantile spasms and mental retardation

X-linked West syndrome, also called "X-linked infantile spasms" (ISSX), is characterized by early-onset generalized seizures, hypsarrhythmia, and mental retardation. Recently, we have shown that the majority of the X-linked families with infantile spasms carry mutations in the aristaless-related homeobox gene (ARX), which maps to the Xp21.3-p22.1 interval, and that the clinical picture in these patients can vary from mild mental retardation to severe ISSX with additional neurological abnormalities. Here, we report a study of two severely affected female patients with apparently de novo balanced X;autosome translocations, both disrupting the serine-threonine kinase 9 (STK9) gene, which maps distal to ARX in the Xp22.3 region. We show that STK9 is subject to X-inactivation in normal female somatic cells and is functionally absent in the two patients, because of preferential inactivation of the normal X. Disruption of the same gene in two unrelated patients who have identical phenotypes (consisting of early-onset severe infantile spasms, profound global developmental arrest, hypsarrhythmia, and severe mental retardation) strongly suggests that lack of functional STK9 protein causes severe ISSX and that STK9 is a second X-chromosomal locus for this disorder.     

Microphthalmia with linear skin defects (MLS), Aicardi,and Goltz syndromes: are they related X-linked dominant male-lethal disorders?

Gene identification for X-linked dominant sporadic disorders is challenging because no extended families exist that can be studied by linkage analysis. Therefore, classic positional cloning approaches are not possible, and other methods have to be used to search for candidate genes. These conditions present the next challenge for disease-gene identification of Mendelian disorders. The various issues and difficulties involved, such as male lethality, X chromosome inactivation, and analysis of phenotypic similarities among different conditions are illustrated through discussion of three X-linked developmental disorders: microphthalmia with linear skin defects (MLS) syndrome, Aicardi syndrome, and Goltz syndrome (focal dermal hypoplasia).     

Mary Lyon and the hypothesis of random X chromosome inactivation

The 50th anniversary of Mary Lyon’s 1961 Nature paper, proposing random inactivation in early embryonic life of one of the two X chromosomes in the cells of mammalian females, provides an opportunity to remember and celebrate the work of those involved. While the hypothesis was initially put forward by Lyon based on findings in the mouse, it was founded on earlier studies, notably the work of Susumu Ohno; it was also suggested independently by Beutler and colleagues using experimental evidence from a human X-linked disorder, glucose-6-phosphate dehydrogenase deficiency, and has proved to be of as great importance for human and medical genetics as it has for general mammalian genetics. Alongside the hypothesis itself, previous cytological studies of mouse and human chromosomes, and the observations on X-linked mutants in both species deserve recognition for their essential role in underpinning the hypothesis of random X-inactivation, while subsequent research on the X-inactivation centre and the molecular mechanisms underlying the inactivation process represent some of the most outstanding contributions to human and wider mammalian genetics over the past 50 years.