The power of the Mediator complex-Expanding the genetic architecture and phenotypic spectrum of MED12-related disorders.

MED12 is a member of the large Mediator complex that controls cell growth, development, and differentiation. Mutations in MED12 disrupt neuronal gene expression and lead to at least three distinct X-linked intellectual disability syndromes (FG, Lujan-Fryns, and Ohdo). Here, we describe six families with missense variants in MED12 (p.(Arg815Gln), p.(Val954Gly), p.(Glu1091Lys), p.(Arg1295Cys), p.(Pro1371Ser), and p.(Arg1148His), the latter being first reported in affected females) associated with a continuum of symptoms rather than distinct syndromes. The variants expanded the genetic architecture and phenotypic spectrum of MED12-related disorders. New clinical symptoms included brachycephaly, anteverted nares, bulbous nasal tip, prognathism, deep set eyes, and single palmar crease. We showed that MED12 variants, initially implicated in X-linked recessive disorders in males, may predict a potential risk for phenotypic expression in females, with no correlation of the X chromosome inactivation pattern in blood cells. Molecular modeling (Yasara Structure) performed to model the functional effects of the variants strongly supported the pathogenic character of the variants examined. We showed that molecular modeling is a useful method for in silico testing of the potential functional effects of MED12 variants and thus can be a valuable addition to the interpretation of the clinical and genetic findings.

Large deletions play a minor but essential role in congenital coagulation factor VII and X deficiencies.

UNLABELLED: Congenital factor VII (FVII) and factor X (FX) deficiencies belong to the group of rare bleeding disorders which may occur in separate or combined forms since both the F7 and F10 genes are located in close proximity on the distal long arm of chromosome 13 (13q34). We here present data of 192 consecutive index cases with FVII and/or FX deficiency. 10 novel and 53 recurrent sequence alterations were identified in the F7 gene and 5 novel as well as 11 recurrent in the F10 gene including one homozygous 4.35 kb deletion within F7 (c.64+430_131-6delinsTCGTAA) and three large heterozygous deletions involving both the F7 and F10 genes. One of the latter proved to be cytogenetically visible as a chromosome 13q34 deletion and associated with agenesis of the corpus callosum and psychomotor retardation. CONCLUSIONS: Large deletions play a minor but essential role in the mutational spectrum of the F7 and F10 genes. Copy number analyses (e. g. MLPA) should be considered if sequencing cannot clarify the underlying reason of an observed coagulopathy. Of note, in cases of combined FVII/FX deficiency, a deletion of the two contiguous genes might be part of a larger chromosomal rearrangement.

Human Ring Chromosomes - New Insights for their Clinical Significance.

Twenty-nine as yet unreported ring chromosomes were characterized in detail by cytogenetic and molecular techniques. For FISH (fluorescence in situ hybridization) previously published high resolution approaches such as multicolor banding (MCB), subcentromere-specific multi-color-FISH (cenM-FISH) and two to three-color-FISH applying locus-specific probes were used. Overall, ring chromosome derived from chromosomes 4 (one case), 10 (one case), 13 (five cases), 14, (three cases), 18 (two cases), 21 (eight cases), 22 (three cases), X (five cases) and Y (one case) were studied. Eight cases were detected prenatally, eight due developmental delay and dysmorphic signs, and nine in connection with infertility and/or Turner syndrome. In general, this report together with data from the literature, supports the idea that ring chromosome patients fall into two groups: group one with (severe) clinical signs and symptoms due to the ring chromosome and group two with no obvious clinical problems apart from infertility.

Parental origin of de novo cytogenetically balanced reciprocal non-Robertsonian translocations.

De novo cytogenetically balanced reciprocal non-Robertsonian translocations are rare findings in clinical cytogenetics and might be associated with an abnormal phenotype. Knowledge of the parental origin and mechanisms of formation is still limited. By microdissection of the derivative chromosomes and their normal homologs from metaphases followed by microsatellite-mediated marker analysis we identified 7 cases of paternal and 3 cases of maternal origin in a cohort of 10 patients with de novo cytogenetically balanced reciprocal non-Robertsonian translocations. Neither in the maternal nor in the paternal group of our study parental age seems to be increased. Together with the data from the literature our results confirm that the majority of de novo cytogenetically balanced reciprocal translocations are of paternal origin, but the preponderance does not appear to be as distinct as previously thought and the paternal age does not seem to be necessarily a major contributing factor.

Mirror-image asymmetry in monozygotic twins with kabuki syndrome.

Kabuki syndrome (OMIM 147920) is a rare disorder characterised by moderate intellectual disability, growth retardation, microcephaly and characteristic facial dysmorphic features which comprise long palpebral fissures, eversion of the lateral third of the eyelids and arched eyebrows with lateral sparseness. Mutations in MLL2 are the most frequent cause of this disorder. More than 100 MLL2 point mutations have been reported, but large intragenic deletions comprising one or more exons have not yet been identified. We report on a pair of monozygotic twin brothers in whom a deletion of 2 neighbouring exons was detected. The twins had the characteristic facial features of Kabuki syndrome, and they suffered from microcephaly, cleft lip and palate and congenital heart disease. Cleft lip and palate were left-sided in the first twin and right-sided in the second twin, i.e. they represented a mirror-image asymmetry. The intragenic deletion in these brothers broadens the spectrum of MLL2 mutations, and they provide a rare example of mirror-image asymmetry of congenital malformations in monozygotic twins.

Mosaic trisomy 21/monosomy 21 in a living female infant.

Many autosomal monosomies are presumed to end in arrested growth in the first few mitoses, prior even to the time of implantation, with possibly some proceeding to the stage of occult abortion. The single exception may be monosomy 21, although this has been questioned, with most earlier reports of monosomy 21 recently re-interpreted as being due to an unbalanced translocation involving chromosome 21. Here we report a female infant with a mosaic trisomy 21/monosomy 21 karyotype. While the karyotype 46,XX,i(21)(q10) is detected in all metaphases investigated in lymphocytes, mosaicism with the karyotype 46,XX,i(21)(q10)[31]/45,XX, -21[12] is seen in fibroblasts from a skin biopsy. Dysmorphic facial features and multiple malformations remarkably resemble cases of monosomy 21 that have been described in the literature. This suggests a dominant phenotypic effect of loss of one chromosome 21. Detailed clinical description, results of gene dosage studies, and cytogenetic analysis will be presented.

Efficient in vitro generation of adult multipotent cells from mobilized peripheral blood CD133+ cells.

OBJECTIVES: To generate non-haematopoietic tissues from mobilized haematopoietic CD133(+) stem cells. MATERIALS AND METHODS: Mobilized peripheral blood CD133(+) cells from adult healthy donors were used. In vitro ability of highly enriched CD133(+) cells from mobilized peripheral blood to generate multipotent cells, and their potential to give rise to cells with characteristics of neuroectoderm, endoderm and mesoderm layers was investigated. RESULTS: We found that a recently identified population of CD45(+) adherent cells generated in vitro after culture of highly purified CD133(+) cells for 3-5 weeks with Flt3/Flk2 ligand and interleukin-6 can, in presence of the appropriate microenvironmental cues, differentiate into neural progenitor-like cells (NPLCs), hepatocyte-like cells and skeletal muscle-like cells. We have termed them to be adult multipotent haematopoietic cells (AMHCs). AMHC-derived NPLCs expressed morphological, phenotypic and molecular markers associated with primary neural progenitor cells. They can differentiate into astrocyte-like cells, neuronal-like cells and oligodendrocyte-like cells. Moreover, AMHC-derived NPLCs produced 3,4-dihydrophenylalanine and dopamine and expressed voltage-activated ion channels, suggesting their functional maturation. In addition, AMHC-derived hepatocyte-like cells and skeletal muscle-like cells, showed typical morphological features and expressed primary tissue-associated proteins. CONCLUSION: Our data demonstrate that AMHCs may therefore serve as a novel source of adult multipotent cells for autologous replacement cell therapies.

Specific transcriptional changes in human fetuses with autosomal trisomies.

Among full autosomal trisomies, only trisomies of chromosome 21 (Down syndrome), 18 (Edwards syndrome) and 13 (Patau syndrome) are compatible with postnatal survival. But the mechanisms, how a supernumerary chromosome disrupts the normal development and causes specific phenotypes, are still not fully explained. As an alternative to gene dosage effect due to the trisomic chromosome a genome-wide transcriptional dysregulation has been postulated. The aim of this study was to define the transcriptional changes in trisomy 13, 18, and 21 during early fetal development in order to obtain more insights into the molecular etiopathology of aneuploidy. Using oligonucleotide microarrays, we analyzed whole genome expression profiles in cultured amniocytes (AC) and chorionic villus cells (CV) from pregnancies with a normal karyotype and with trisomies of human chromosomes 13, 18 and 21. We observed a low to moderate up-regulation for a subset of genes of the trisomic chromosomes. Transcriptional levels of most of the genes on the supernumerary chromosome appeared similar to the respective chromosomal pair in normal karyotypes. A subset of chromosome 21 genes including the DSCR1 gene involved in fetal heart development was consistently up-regulated in different prenatal tissues (AC, CV) of trisomy 21 fetuses whereas only minor changes were found for genes of all other chromosomes. In contrast, in trisomy 18 vigorous downstream transcriptional changes were found. Global transcriptome analysis for autosomal trisomies 13, 18, and 21 supported a combination of the two major hypotheses.

Small supernumerary marker chromosomes (sSMC) in patients with a 45,X/46,X,+mar karyotype - 17 new cases and a review of the literature.

Small supernumerary marker chromosomes (sSMC) can appear in a numerically normal 'basic karyotype', but also in a numerically abnormal one like a Turner syndrome karyotype (= sSMC(T)). Here we present 17 new cases with such a mos 45,X/46,X,+mar karyotype. Moreover we reviewed all 512 cytogenetically similar cases available from the literature and supply for the first time data on occurrence, shapes and subgroups of this rare cytogenetic entity. sSMC(T) are very rare in the common population (1:100,000) - however, they can be observed with a 45- and even 60-times higher frequency in infertile and (develop)mentally retarded patients, respectively. Even though sSMC(T) derive from one of the gonosomes in >99% of the cases, there are also exceptional reports on sSMC(T) derived from one of the autosomes. The majority of sSMC(T)(X) form ring chromosomes, while most sSMC(T)(Y) are inverted duplicated/isodicentric chromosomes. Although >500 sSMC(T) are reported, a detailed characterization of the chromosomal breakpoints is only given for a minority. Thus, more cases with detailed (molecular) cytogenetic marker chromosome characterization are needed to provide information on formation and effects of an sSMC(T).

De novostructural chromosomal imbalances: molecular cytogenetic characterization of partial trisomies.

De novo structural chromosomal imbalances represent a major challenge in modern cytogenetic diagnostics. Based solely on conventional cytogenetic techniques it may be impossible to identify the chromosomal origin of additional chromosomal material. In these cases molecular cytogenetic investigations including multicolor-FISH (M-FISH), spectral karyotyping (SKY), multicolor banding (MCB) and cenM-FISH combined with appropriate single-locus FISH probes are highly suitable for the determination of the chromosomal origin and fine characterization of derivative chromosomes. Here we report on four patients with de novo chromosomal imbalances and distinct chromosomal phenotypes, three of them harboring pure partial trisomies: a mildly affected boy with pure partial trisomy 10q22.2-->q22.3 approximately 23.1 due to an interstitial duplication, a girl with pure trisomy 12p11.21-->pter and atypically moderate phenotype as the consequence of an X;autosome translocation, and a girl with multiple congenital abnormalities and severe developmental delay and a 46,XX,15p+ karyotype hiding a trisomy 17pter-->17q11.1. The fourth patient is a girl with minor phenotypic features and mental retardation with an inverted duplication 18q10-->p11.31 combined with a terminal deletion of 18p32. The clinical pictures are compared with previously described patients with focus on long term outcome.

Small supernumerary marker chromosomes--progress towards a genotype-phenotype correlation.

Small supernumerary marker chromosomes (sSMC) are still a major problem in clinical cytogenetics as they are too small to be characterized for their chromosomal origin by traditional banding techniques, but require molecular cytogenetic techniques for their identification. Apart from the correlation of about one third of the sSMC cases with a specific clinical picture, i.e. the i(18p), der(22), i(12p) (Pallister Killian syndrome) and inv dup(22) (cat-eye) syndromes, most of the remaining sSMC have not yet been correlated with clinical syndromes. Recently, we reviewed the available >1600 sSMC cases (Liehr T, sSMC homepage: http://mti-n.mti.uni-jena.de/~huwww/MOL_ZYTO/sSMC.htm). A total of 387 cases (including the 45 new cases reported here) have been molecularly cytogenetically characterized with regard to their chromosomal origin, the presence of euchromatin, heterochromatin and satellite material. Based on analysis of these cases we present the first draft of a basic genotype-phenotype correlation for sSMC for all human chromosomes apart from the chromosomes Y, 10, 11 and 13.

Dominant transmission of prepubertal gynecomastia due to serum estrone excess: hormonal, biochemical, and genetic analysis in a large kindred.

Increased extraglandular aromatization has been reported to cause the rare entity of familial gynecomastia. Recently heterozygous inversions at the p450 aromatase gene promotor locus were detected in two different families with this syndrome. We studied a family in which seven affected males over three generations had inherited prepubertal gynecomastia in an autosomal dominant manner. The proband developed gynecomastia at 11.5 yr, entered puberty at 12.5 yr, but was incompletely virilized at 19 yr. A similar development was observed in his affected stepbrother and one first-degree cousin. All three boys had acceleration of prepubertal growth and bone age. The older two had a diminished pubertal growth spurt and precocious growth arrest, but their final heights were within the range of their target height. In addition, the maternal grandfather and three maternal uncles were affected, who all had been mastectomized. The mother of the proband had normal age at menarche and no macromastia. Estrone levels of the proband and the other affected boys were elevated, 17beta-estradiol levels were high-normal, and testosterone levels were low. Hormonal analyses of the affected adults, who had all fathered children, revealed pathologically low serum testosterone levels but normal to high-normal levels of estradiol and estrone. The mother of the proband had elevated estrone levels. Treatment of the proband was more effective with anastrozole than with testolactone and increased the initially reduced testes volume to normal size, promoted virilization, and normalized serum estrone and testosterone levels. Neither preadipocytes from breast fat tissue of the affected stepbrother nor peripheral lymphocytes of the affected boys exhibited increased aromatase activity in culture. Therefore, these cells can be excluded from being the source of estrone excess. In addition, serum of the proband and his stepbrother did not contain factors promoting aromatase activity as assayed using preadipocytes from control individuals.A repeat polymorphism of the p450 aromatase gene cosegregated with the disease phenotype in the family, making a mutation of the p450 aromatase gene likely. Single-strand conformational polymorphism analysis of the known alternative untranslated exons and all coding exons of the p450 aromatase gene did not indicate any mutation. In addition, fluorescent in situ hybridization analysis using four probes covering the promotor region did not reveal the presence of any major inversion at this locus. In conclusion, preadipocytes and blood cells were excluded as the cell source of increased aromatization. Fluorescent in situ hybridization and single-strand conformational polymorphism analyses did not reveal any mutation of the p450 aromatase gene, but an intragenic polymorphic marker cosegregated with the disease phenotype. Excess of serum estrone in the presence of normal 17beta-estradiol levels may be the only indicative serum parameter of this mild manifestation of aromatase excess syndrome, which includes prepubertal gynecomastia and moderate hypogonadism in men but not necessarily short stature. In women, this mode of aromatase excess may remain clinically inapparent.

A complex chromosomal rearrangement with a translocation 4;10;14 in a fertile male carrier: ascertainment through an offspring with partial trisomy 14q13-->q24.1 and partial monosomy 4q27-->q28 [corrected].

Complex chromosomal rearrangements (CCRs) are usually associated with infertility or subfertility in male carriers. If fertility is maintained, there is a high risk of abnormal pregnancy outcome. Few male carriers have been identified by children presenting with mental retardation/congenital malformations (MR/CM) or by spontaneous abortions of the spouses. We report a de novo CCR with five breakpoints involving chromosomes 4, 10 and 14 in a male carrier who was ascertained through a son presenting with MR/CM due to an unbalanced karyotype with partial trisomy 14 and partial monosomy 4. The child has a healthy elder brother. In the family history no abortions were reported. No fertility treatment was necessary. Cytogenetic analysis from the affected son showed a reciprocal translocation t(4;10) with additional chromosomal material inserted between the translocation junctions in the derivative chromosome 10. The father showed the same derivative chromosome 10 but had additionally one aberrant chromosome 14. Further molecular cytogenetic analyses determined the inserted material in the aberrant chromosome 10 as derived from chromosome 14 and revealed a small translocation with material of chromosome 4 inserted into the derivative chromosome 14. Thus the phenotype of the son is supposed to be associated with a partial duplication 14q13-->q24.1 and a partial monosomy 4q27-->q28. Including our case we are aware of eleven CCR cases with fertile male carriers. In eight of these families normal offspring have been reported. We propose that exceptional CCRs in fertile male carriers might form comparatively simple pachytene configurations increasing the chance of healthy offspring.

[Microdeletion 22q11.2. A too rarely diagnosed genetic change in psychiatric illnesses]. / Mikrodeletion 22q11.2. Eine zu selten diagnostizierte genetische Veränderung bei psychiatrischen Erkrankungen.

Microdeletion 22q11.2 with an estimated incidence of 1:4000 is known to cause the DiGeorge syndrome (DGS) or the velocardiofacial syndrome (VCFS), both usually being diagnosed in the newborn period or childhood. Recent studies have shown that children suffering from VCFS frequently develop psychiatric disorders in late adolescence or adulthood. Here we report the case of a 30-year-old man presenting with slight facial dysmorphisms, hypoparathyreoidism, minor cardiac anomalies, and slight cognitive impairments who had developed a severe personality disorder which eventually led to the diagnosis of microdeletion 22q11.2 with maternal inheritance. Psychiatric patients should be thoroughly examined for typical signs associated with this chromosomal anomaly. Genetic diagnosis is necessary because of the 50% probability of inheritance with possibly severe congenital anomalies. In view of a prevalence of 2% in an unselected group of patients with schizophrenic psychosis, microdeletion 22q11.2 is likely to be underdiagnosed.

Hydrops fetalis in three male fetuses of a female with incontinentia pigmenti.

OBJECTIVES: Careful investigation of hydrops fetalis (HF) is important with regard to genetic counselling and prenatal diagnosis. HF is known to be associated with various genetic disorders. To date there has been only one report of a male fetus in whom incontinentia pigmenti (IP) was associated with generalised oedema. We describe a family who had a girl with clinical signs of IP after three consecutive miscarriages of three male fetuses due to HF. RESULTS: Molecular genetic analysis showed a mutation in the NEMO/IKK(chi) gene in the girl and the mother, which confirmed the diagnosis of IP in both cases. In the two fetuses that could be investigated, inheritance of the affected maternal X chromosome could be demonstrated retrospectively by linkage analysis. CONCLUSION: The present findings suggest that IP might be an X-linked dominant trait causing HF in male fetuses. In cases of recurrent HF in male fetuses, minimal signs of IP in the maternal line should therefore be carefully investigated in order to be able to perform mutational analysis and to offer appropriate genetic counselling.

Unusual chromosomal mosaicism as a cause of mental retardation and congenital malformations in a familial reciprocal translocation carrier, t(17;22)(q24.2;q11.23).

Familial reciprocal translocations are generally without phenotypic effect, although there is some evidence for a small excess of mental retardation and congenital malformations (MR/CM) in children carrying familial reciprocal translocations. Possible mechanisms whereby such translocations could have a phenotypic effect include cryptic unbalanced rearrangements, uniparental disomy, and disruption of putative genes at the breakpoints, unmasking recessive alleles on the normal homologs. Mosaicism for a supernumerary derivative chromosome in a carrier of a familial reciprocal translocation has not yet been described. We report a boy presenting with MR/CM and a familial reciprocal translocation, t(17;22)(q24.2;q11.23), inherited from the mother. Cytogenetic analysis of peripheral blood lymphocytes showed a balanced karyotype in all 32 analyzed metaphase spreads. Molecular genetic analysis was consistent with biparental origin of the normal homologs. In metaphase spreads from skin fibroblasts a supernumerary chromosome was found in all 24 cells analyzed and could be identified as der(22)t(17;22)(q24.2;q11.23). Several possible segregation modes at meiosis I followed by meiosis II or postzygotic nondisjunction of the der(22) might have led to this unusual chromosomal mosaicism. We propose hidden mosaicism as a possible cause for MR/CM in patients who apparently carry a balanced familial reciprocal translocation.

Partial tetrasomy 12pter-12p12.3 in a girl with Pallister-Killian syndrome: extraordinary finding of an analphoid, inverted duplicated marker.

Cytogenetic analysis in a girl with multiple congenital anomalies indicating Pallister-Killian syndrome (PKS) showed a supernumerary marker chromosome in 1/76 lymphocytes and 34/75 fibroblast metaphases. GTG-banding pattern was consistent with the chromosomal region 12pter-12q11. While fluorescence-in-situ hybridisation (FISH) with a whole chromosome 12 painting probe confirmed the origin of the marker, a chromosome 12 specific alpha-satellite probe did not hybridise to it. FISH analysis with a specific subtelomeric probe 12p showed hybridisation to both ends of the marker chromosome. High-resolution multicolour-banding (MCB) studies revealed the marker to be a der(12)(pter-->p12.3::p12.3-->pter). Summarising the FISH information, we defined the marker as an inverted duplication of 12pter-12p12.3 leading to partial tetrasomy of chromosome 12p. In skin fibroblasts, cultured at the patient's age of 1 year and 9 years, the marker chromosome was found in similar frequencies, even after several culture passages. Therefore, we consider the marker to have a functional centromere although it lacks detectable centromeric alpha-satellite sequences. To the best of our knowledge, this is the first proven analphoid marker of chromosome 12. Molecular genetic studies indicated that this marker is of paternal origin. The finding of partial tetrasomy 12pter-12p12.3 in our PKS patient allows to narrow down the critical region for PKS.

Tall stature, gonadal dysgenesis, and stigmata of Turner's syndrome caused by a structurally altered X chromosome.

Genetic analysis in a tall 14-year-old girl with gonadal dysgenesis and some stigmata of Turner's syndrome revealed a duplication of the short arm in a monocentric X chromosome with partial loss of Xq. We suggest that triple gene dosage of SHOX and estrogen deficiency caused the unique overgrowth.