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.     

Temperature modulation of DHPLC analysis for detection of coexisting constitutional and mosaic sequence variants in TSC2

Somatic mosaicism is a frequent phenomenon in Mendelian disorders that exhibit a high proportion of new mutations. However, mutant alleles present at low frequency may escape detection. We have previously shown that denaturing high-performance liquid chromatography (DHPLC) at the recommended melt temperature can detect TSC1 and TSC2 mutations in tuberous sclerosis patients with low-level somatic mosaicism, even when direct sequencing cannot identify the causative lesion. Here, we report the use of temperature modulation in DHPLC analysis to facilitate the robust detection of a mosaic mutation, N1643K, in the presence of a coexisting constitutional polymorphism.     

Somatic mosaicism and variable expressivity

For more than 50 years geneticists have assumed that variations in phenotypic expression are caused by alterations in genotype. Recent evidence shows that 'simple' mendelian disorders or monogenic traits are often far from simple, exhibiting phenotypic variation (variable expressivity) that cannot be explained entirely by a gene or allelic alteration. In certain cases of androgen insensitivity syndrome caused by identical mutations in the androgen receptor gene, phenotypic variability is caused by somatic mosaicism, that is, somatic mutations that occur only in certain androgen-sensitive cells. Recently, more than 30 other genetic conditions that exhibit variable expressivity have been linked to somatic mosaicism. Somatic mutations have also been identified in diseases such as prostate and colorectal cancer. Therefore, the concept of somatic mutations and mosaicism is likely to have far reaching consequences for genetics, in particular in areas such as genetic counseling.     

Mosaike bei monogenen Erkrankungen

Mosaicism is defined as the simultaneous presence of cells with different genotypes that originate from a common zygote. Mutations can either be present in germline or somatic cells. Monogenic disorders apparently caused by a de novo mutation may show a recurrence risk due to germline mosaicism in a parent. Duchenne muscular dystrophy is a well investigated example with a high frequency of germline mosaicism and the estimation for the risk of recurrence is based on theoretical models and empirical data. Recently, somatic mutations have been uncovered in various syndromic disorders, such as Proteus syndrome or hemimegalencephaly and respective mutations often show gain-of-function properties. Genetic testing is mainly based on next generation sequencing technologies but still remains challenging; however, detection of somatic mosaicism is expected to be of increasing relevance in the diagnosis of monogenic disorders. Somatic mosaicism may also play a hitherto underestimated role in common disorders.     

Parental Somatic Mosaicism Is Underrecognized and Influences Recurrence Risk of Genomic Disorders

New human mutations are thought to originate in germ cells, thus making a recurrence of the same mutation in a sibling exceedingly rare. However, increasing sensitivity of genomic technologies has anecdotally revealed mosaicism for mutations in somatic tissues of apparently healthy parents. Such somatically mosaic parents might also have germline mosaicism that can potentially cause unexpected intergenerational recurrences. Here, we show that somatic mosaicism for transmitted mutations among parents of children with simplex genetic disease is more common than currently appreciated. Using the sensitivity of individual-specific breakpoint PCR, we prospectively screened 100 families with children affected by genomic disorders due to rare deletion copy-number variants (CNVs) determined to be de novo by clinical analysis of parental DNA. Surprisingly, we identified four cases of low-level somatic mosaicism for the transmitted CNV in DNA isolated from parental blood. Integrated probabilistic modeling of gametogenesis developed in response to our observations predicts that mutations in parental blood increase recurrence risk substantially more than parental mutations confined to the germline. Moreover, despite the fact that maternally transmitted mutations are the minority of alleles, our model suggests that sexual dimorphisms in gametogenesis result in a greater proportion of somatically mosaic transmitting mothers who are thus at increased risk of recurrence. Therefore, somatic mosaicism together with sexual differences in gametogenesis might explain a considerable fraction of unexpected recurrences of X-linked recessive disease. Overall, our results underscore an important role for somatic mosaicism and mitotic replicative mutational mechanisms in transmission genetics.     

De novo facioscapulohumeral muscular dystrophy: frequent somatic mosaicism, sex-dependent phenotype, and the role of mitotic transchromosomal repeat interaction between chromosomes 4 and 10

Autosomal dominant facioscapulohumeral muscular dystrophy (FSHD) is caused by deletion of most copies of the 3.3-kb subtelomeric D4Z4 repeat array on chromosome 4q. The molecular mechanisms behind the deletion and the high proportion of new mutations have remained elusive. We surveyed 35 de novo FSHD families and found somatic mosaicism in 40% of cases, in either the patient or an asymptomatic parent. Mosaic males were typically affected; mosaic females were more often the unaffected parent of a nonmosaic de novo patient. A genotypic-severity score, composed of the residual repeat size and the degree of somatic mosaicism, yields a consistent relationship with severity and age at onset of disease. Mosaic females had a higher proportion of somatic mosaicism than did mosaic males. The repeat deletion is significantly enhanced by supernumerary homologous repeat arrays. In 10% of normal chromosomes, 4-type repeat arrays are present on chromosome 10. In mosaic individuals, 4-type repeats on chromosome 10 are almost five times more frequent. The reverse configuration, also 10% in normal chromosomes, was not found, indicating that mutations may arise from transchromosomal interaction, to which the increase in 4-type repeat clusters is a predisposing factor. The somatic mosaicism suggests a mainly mitotic origin; mitotic interchromosomal gene conversion or translocation between fully homologous 4-type repeat arrays may be a major mechanism for FSHD mutations.     

Prenatal diagnosis and molecular cytogenetic characterization of mosaicism for a small supernumerary marker chromosome derived from chromosome 22 associated with cat eye syndrome

We present prenatal diagnosis of mosaicism for a small supernumerary marker chromosome (sSMC) derived from chromosome 22 associated with cat eye syndrome (CES) using cultured amniocytes in a pregnancy with fetal microcephaly, intrauterine growth restriction, left renal hypoplasia, total anomalous pulmonary venous return with dominant right heart and right ear deformity. The sSMC was bisatellited and dicentric, and was characterized by multiplex ligation-dependent probe amplification (MLPA) and array comparative genomic hybridization (aCGH). The SALSA MLPA P250-B1 DiGeorge Probemix showed duplication of gene dosage in the CES region. aCGH showed a 1.26-Mb duplication at 22q11.1–q11.21 encompassing CECR1–CECR7. The sSMC was likely inv dup(22) (q11.21). Prenatal diagnosis of an sSMC(22) at amniocentesis should alert CES. MLPA, aCGH and fetal ultrasound are useful for rapid diagnosis of CES in case of prenatally detected sSMC(22).     

Small Supernumerary Marker Chromosomes and Uniparental Disomy Have a Story to Tell

Small supernumerary maker chromosomes (sSMC) and uniparental disomy (UPD) are rare, and a combination of both is rarely encountered. Accordingly, only 46 sSMC cases UPD have been reported. Despite of its rareness, UPD has to be considered, especially in prenatal cases with sSMC. Here, the authors reviewed all sSMC cases with UPD (sSMC^U+) and compared them to sSMC without UPD (sSMC^U−), which resulted in the following correlations: 1) every sSMC, irrespective of its chromosomal origin, may be principally connected with UPD; 2) mixed hetero- and iso-UPD (hUPD/iUPD) can be observed most often in sSMC^U+ cases followed by complete iUPD, complete hUPD, and segmental iUPD; 3) UPD of chromosomes 6, 7, 14, 15, 16, and 20 is most often reported in sSMC^U+; 4) maternal UPD was approximately nine times more frequent than paternal UPD; 5) if mosaic with a normal cell line, acrocentric-derived sSMC had a three times higher chance of occurrence than the corresponding nonmosaic sSMC cases; 6) UPD in connection with a parentally inherited sSMC is, if existent at all, a rare event; and 7) the gender type and shape of sSMC had no effect on UPD formation. Overall, sSMC^U+ cases may have a story to tell about chromosome number control mechanisms in early embryogenesis.     

Unusual small supernumerary marker chromosome (sSMC) 9 in a Klinefelter patient

Small supernumerary marker chromosomes (sSMC) are small additional chromosomes characterizable for their origin only by molecular cytogenetic approaches. sSMC have been reported previously in four types of syndromes associated with chromosomal imbalances: in approximately 150 cases with Turner syndrome, 26 cases with Down syndrome and only one case each with Klinefelter syndrome and "Triple-X"-syndrome. Here we report the second case with an sSMC detected in addition to a Klinefelter karyotype. Molecular cytogenetics applying centromere-specific multicolor FISH (cenM-FISH) and a specific subcentromere-specific (subcenM-FISH) probe set characterized the sSMC as a dic(9)(:p12-->q11.1::q11.1--> p11.1:). The reported patient was described with hypogonadism, gynaecomastia plus a bronchial carcinoma. The patient's clinical features are discussed in connection with other Klinefelter cases and possible consequences of presence of the sSMC(9). Furthermore, a suggestion is made for the mode of sSMC-formation in this case.     

Mosaic Uniparental Disomies and Aneuploidies as Large Structural Variants of the Human Genome

Mosaicism is defined as the coexistence of cells with different genetic composition within an individual, caused by postzygotic somatic mutation. Although somatic mosaicism for chromosomal abnormalities is a well-established cause of developmental and somatic disorders and has also been detected in different tissues, its frequency and extent in the adult normal population are still unknown. We provide here a genome-wide survey of mosaic genomic variation obtained by analyzing Illumina 1M SNP array data from blood or buccal DNA samples of 1991 adult individuals from the Spanish Bladder Cancer/EPICURO genome-wide association study. We found mosaic abnormalities in autosomes in 1.7% of samples, including 23 segmental uniparental disomies, 8 complete trisomies, and 11 large (1.5–37 Mb) copy-number variants. Alterations were observed across the different autosomes with recurrent events in chromosomes 9 and 20. No case-control differences were found in the frequency of events or the percentage of cells affected, thus indicating that most rearrangements found are not central to the development of bladder cancer. However, five out of six events tested were detected in both blood and bladder tissue from the same individual, indicating an early developmental origin. The high cellular frequency of the anomalies detected and their presence in normal adult individuals suggest that this type of mosaicism is a widespread phenomenon in the human genome. Somatic mosaicism should be considered in the expanding repertoire of inter- and intraindividual genetic variation, some of which may cause somatic human diseases but also contribute to modifying inherited disorders and/or late-onset multifactorial traits.     

High rate of mosaicism in tuberous sclerosis complex.

Six families with mosaicism are identified in a series of 62 unrelated families with a mutation in one of the two tuberous sclerosis complex (TSC) genes, TSC1 or TSC2. In five families, somatic mosaicism was present in a mildly affected parent of an index patient. In one family with clinically unaffected parents, gonadal mosaicism was detected after TSC was found in three children. The detection of mosaicism has consequences for genetic counseling of the families involved, as changed risks apply to individuals with mosaicism, both siblings and parents. Clinical investigation of parents of patients with seemingly sporadic mutations is essential to determine their residual chance of gonadal and/or somatic mosaicism, unless a mosaic pattern is detected in the index patient, proving a de novo event. In our data set, the exclusion of signs of TSC in the parents of a patient with TSC reduced the chance of one of the parents to be a (mosaic) mutation carrier from 10% to 2%. In the five families with somatic mosaicism, the parent was given the diagnosis after the diagnosis was made in the child.     

Somatic and germline mosaic mutations in the doublecortin gene are associated with variable phenotypes

Mutations in the X-linked gene doublecortin lead to "double cortex" syndrome (DC) in females and to X-linked lissencephaly (XLIS) in males. Because most patients with DC and XLIS are sporadic, representing de novo doublecortin mutations, we considered that some of these patients could be somatic or germline mosaics. Among a population of 20 patients and their families, we found evidence for mosaic doublecortin mutations in 6 individuals. Germline mosaicism was identified in two unaffected women, each with two affected children. Additionally, one affected male with DC was found to be a somatic mosaic, which presumably spared him from the more severe phenotype of lissencephaly. The high rate of mosaicism indicates that there may be a significant recurrence risk for DC/XLIS in families at risk, even when the mother is unaffected.     

Germinal and Somatic Trisomy 21 Mosaicism: How Common is it,What are the Implications for Individual Carriers and How Does it Come About?

It is well known that varying degrees of mosaicism for Trisomy 21, primarily a combination of normal and Trisomy 21 cells within individual tissues, may exist in the human population. This involves both Trisomy 21 mosaicism occurring in the germ line and Trisomy 21 mosaicism documented in different somatic tissues, or indeed a combination of both in the same subjects. Information on the incidence of Trisomy 21 mosaicism in different tissue samples from people with clinical features of Down syndrome as well as in the general population is, however, still limited. One of the main reasons for this lack of detailed knowledge is the technological problem of its identification, where in particular low grade/cryptic Trisomy 21 mosaicism, i.e. occurring in less than 3-5% of the respective tissues, can only be ascertained by fluorescence in situ hybridization (FISH) methods on large cell populations from the different tissue samples.In this review we summarize current knowledge in this field with special reference to the question on the likely incidence of germinal and somatic Trisomy 21 mosaicism in the general population and its mechanisms of origin. We also highlight the reproductive and clinical implications of this type of aneuploidy mosaicism for individual carriers. We conclude that the risk of begetting a child with Trisomy 21 Down syndrome most likely is related to the incidence of Trisomy 21 cells in the germ line of any carrier parent. The clinical implications for individual carriers may likewise be dependent on the incidence of Trisomy 21 in the relevant somatic tissues. Remarkably, for example, there are indications that Trisomy 21 mosaicism will predispose carriers to conditions such as childhood leukemia and Alzheimer's Disease but there is on the other hand a possibility that the risk of solid cancers may be substantially reduced.     

Somatic mosaicism and cancer: inference based on a conditional Luria-Delbrück distribution

Somatic mosaicism for mutations in disease-causing genes has been reported in several recent studies. Examples include hemophilia A, many skin disorders, and several cancers such as retinoblastoma and familial adenomatous polyposis. Many of these disorders require multiple mutations in order to express the disease phenotype. For example, two recessive mutations to the retinoblastoma locus are required to initiate retinoblastomal tumors. I develop a mathematical framework for somatic mosaicism in which two recessive mutations cause disease. With my framework, I analyse the following question: Given an observed frequency of cells with two mutations and an easily scored aberrant phenotype, what is the conditional frequency distribution of cells carrying one mutation and therefore susceptible to transformation by a second mutation? This question is important because a high frequency of carrier cells can cause genetic counselors to misdiagnose a mosaic as an inherited heterozygote carrier and because widespread mosaicism can lead to some germline transmission. As more data accumulate, the observed distribution of mosaics can be compared against my predicted distribution. These sorts of studies will contribute to a broader understanding of the distribution of somatic mutations, a central topic in the study of cancer.     

Severe psychomotor retardation in a boy with a small supernumerary marker chromosome 19p

We describe the clinical case of a nine-year-old boy with psychomotor retardation and a small supernumerary marker chromosome (sSMC) present in mosaic form. Fluorescence in situ hybridization (FISH) using centromere cross-hybridizing probes D1/5/19Z (pZ5.1), the whole chromosome paint probe 19, pool YACs19p (839B1, 872G3, 728C8), and pool YACs19q (767C4, 761C1, 786G6) demonstrated that the sSMC was derived from chromosome 19p. Based on GTG-banding and FISH analyses, the patient's karyotype was interpreted as: 47,XY,+mar.ish der(19) (:p13.3-->p11:)(839B1+, 872G3+,728C8+, D1/5/19Z+) de novo[52]/46,XY[48]. To our knowledge, only two other similar cases have been reported. This case helps to better delineate karyotype-phenotype correlations between sSMC 19p and associated clinical phenomena.     

Neocentric small supernumerary marker chromosomes (sSMC)--three more cases and review of the literature

Here we report on three new patients with neocentric small supernumerary marker chromosomes (sSMC) derived from chromosome 2, 13 and 15, respectively. The sSMC(13) and sSMC(15) had inverted duplicated shapes and the sSMC(2) a ring chromosome shape. All three cases were clinically severely abnormal. A review of the available sSMC literature revealed that up to the present 73 neocentric sSMC cases including these three new cases have been reported. Seven of these cases were not characterized morphologically; in the remainder, 80% had an inverted duplication, 17% a ring and 3% a minute shape. 81% of the reported neocentric sSMC carriers showed severe, 12% moderate and 8% no clinical abnormalities. In summary, we report three more neocentric sSMC cases, provide a review on all up to now published cases, highlight their special characteristics and compare them to centric sSMC.     

Neurofibromatosis type 2 attributable to gonosomal mosaicism in a clinically normal mother, and identification of seven novel mutations in the NF2 gene

Neurofibromatosis type 2 (NF2) is an autosomal dominant cancer syndrome that predisposes to the development of bilateral vestibular schwannomas sometimes associated with schwannomas at other locations, meningiomas, ependymomas and juvenile posterior subcapsular lenticular opacities. This disease is caused by inactivating mutations in the NF2 tumour-suppressor gene, located in 22q12. Recently, somatic mosaicism has been demonstrated in some "de novo" NF2 patients. We here report the genetic study of 33 NF2 patients from 33 unrelated Italian families. Twelve mutations were characterised, including seven newly identified mutations and five recurrent ones. Furthermore, we describe one patient with an inactivating mutation that lies in exon 13 but that is present in only a portion of the lymphocytes and, more importantly, a clinically normal individual carrying a somatic/germinal mosaicism for a nonsense mutation in exon 10 of the NF2 gene. Our results confirm the relatively high percentage of mosaicism for mutations in the NF2 gene and establish the importance of evaluating genomic DNA from several tissues, in addition to lymphocytes, so as to identify mosaicism in "de novo" NF2 patients and their relatives. In addition, the demonstration of somatic and/or gonadal mosaicism is an important tool for accurate genetic counselling in families with sporadic cases of NF2.     

New immortalized cell lines of patients with small supernumerary marker chromosome: towards the establishment of a cell bank.

Sixteen newly established cell lines with small supernumerary marker chromosomes (sSMC) derived from chromosomes 1, 2, 4, 6, 7, 8, 14, 15, 16, 18, 19, 21, and 22 are reported. Two sSMC are neocentric and derived from 15q24.1-qter and 2q35-q36, respectively. Two further cases each present with two sSMC of different chromosomal origin. sSMC were characterized by multicolor fluorescence in situ hybridization for their chromosomal origin and genetic content. Moreover, uniparental disomy of the sister chromosomes of the sSMC was excluded in all nine cases studied for that reason. The 16 cases provide information to establish a refined genotype-phenotype correlation of sSMC and are available for future studies.     

Somatic mosaicism for copy-neutral loss of heterozygosity and DNA copy number variations in the human genome

Background

Somatic mosaicism denotes the presence of genetically distinct populations of somatic cells in one individual who has developed from a single fertilised oocyte. Mosaicism may result from a mutation that occurs during postzygotic development and is propagated to only a subset of the adult cells. Our aim was to investigate both somatic mosaicism for copy-neutral loss of heterozygosity (cn-LOH) events and DNA copy number variations (CNVs) in fully differentiated tissues.

Results

We studied panels of tissue samples (11–12 tissues per individual) from four autopsy subjects using high-resolution Illumina HumanOmniExpress-12 BeadChips to reveal the presence of possible intra-individual tissue-specific cn-LOH and CNV patterns.We detected five mosaic cn-LOH regions >5 Mb in some tissue samples in three out of four individuals. We also detected three CNVs that affected only a portion of the tissues studied in one out of four individuals. These three somatic CNVs range from 123 to 796 kb and are also found in the general population. An attempt was made to explain the succession of genomic events that led to the observed somatic genetic mosaicism under the assumption that the specific mosaic patterns of CNV and cn-LOH changes reflect their formation during the postzygotic embryonic development of germinal layers and organ systems.

Conclusions

Our results give further support to the idea that somatic mosaicism for CNVs, and also cn-LOHs, is a common phenomenon in phenotypically normal humans. Thus, the examination of only a single tissue might not provide enough information to diagnose potentially deleterious CNVs within an individual. During routine CNV and cn-LOH analysis, DNA derived from a buccal swab can be used in addition to blood DNA to get information about the CNV/cn-LOH content in tissues of both mesodermal and ectodermal origin. Currently, the real frequency and possible phenotypic consequences of both CNVs and cn-LOHs that display somatic mosaicism remain largely unknown. To answer these questions, future studies should involve larger cohorts of individuals and a range of tissues.

Electronic supplementary material

The online version of this article (doi:10.1186/s12864-015-1916-3) contains supplementary material, which is available to authorized users.