Institut für Humangenetik
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- Institut für Humangenetik (237)
- Theodor-Boveri-Institut für Biowissenschaften (49)
- Deutsches Zentrum für Herzinsuffizienz (DZHI) (6)
- Kinderklinik und Poliklinik (6)
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Sonstige beteiligte Institutionen
- Comprehensive Hearing Center, Department of ORL, Plastic, Aesthetic and Reconstructive Head and Neck Surgery, Würzburg, Germany (1)
- DNA Analytics Core Facility, Biocenter, University of Würzburg, Würzburg, Germany (1)
- Department of Animal Ecology and Tropical Biology, University of Würzburg, Würzburg, Germany (1)
- Maastricht University, Maastricht, the Netherlands (1)
Background
The spondylodysplastic Ehlers-Danlos subtype (OMIM #130070) is a rare connective tissue disorder characterized by a combination of connective tissue symptoms, skeletal features and short stature. It is caused by variants in genes encoding for enzymes involved in the proteoglycan biosynthesis or for a zinc transporter.
Presentation of cases
We report two brothers with a similar phenotype of short stature, joint hypermobility, distinct craniofacial features, developmental delay and severe hypermetropia indicative for a spondylodysplastic Ehlers-Danlos subtype. One also suffered from a recurrent pneumothorax. Gene panel analysis identified two compound heterozygous variants in the B4GALT7 gene: c.641G > A and c.723 + 4A > G. B4GALT7 encodes for galactosyltransferase I, which is required for the initiation of glycosaminoglycan side chain synthesis of proteoglycans.
Conclusions
This is a first full report on two cases with spondylodysplastic Ehlers-Danlos syndrome and the c.723 + 4A > G variant of B4GALT7. The recurrent pneumothoraces observed in one case expand the variable phenotype of the syndrome.
Fibroblasts isolated from a skin biopsy of a healthy 46-year-old female were infected with Sendai virus containing the Yamanaka factors to produce transgene-free human induced pluripotent stem cells (iPSCs). CRISPR/Cas9 was used to generate isogenic cell lines with a gene dose-dependent deficiency of CDH13, a risk gene associated with neurodevelopmental and psychiatric disorders. Thereby, a heterozygous CDH13 knockout (CDH13\(^{+/-}\)) and a CDH13 null mutant (CDH13\(^{-/-}\)) iPSC line was obtained. All three lines showed expression of pluripotency-associated markers, the ability to differentiate into cells of the three germ layers in vitro, and a normal female karyotype.
Arrhythmogenic cardiomyopathy (ACM) is characterized by fibro-fatty replacement of the myocardium, heart failure and life-threatening ventricular arrhythmias. Causal mutations were identified in genes encoding for proteins of the desmosomes, predominantly plakophilin-2 (PKP2) and desmoglein-2 (DSG2). We generated gene-edited knock-out iPSC lines for PKP2 (JMUi001-A-2) and DSG2 (JMUi001-A-3) using the CRISPR/Cas9 system in a healthy control iPSC background (JMUi001A). Stem cell-like morphology, robust expression of pluripotency markers, embryoid body formation and normal karyotypes confirmed the generation of high quality iPSCs to provide a novel isogenic human in vitro model system mimicking ACM when differentiated into cardiomyocytes.
BRCA1 is a major gatekeeper of genomic stability. Acting in multiple central processes like double-strand break repair, centrosome replication, and checkpoint control, BRCA1 participates in maintaining genomic integrity and protects the cell against genomic instability. Chromosomal instability (CIN) as part of genomic instability is an inherent characteristic of most solid tumors and is also involved in breast cancer development. In this study, we determined the extent of CIN in 32 breast cancer tumors of women with a BRCA1 germline mutation compared to 62 unselected breast cancers. We applied fluorescence in situ hybridization (FISH) with centromere-specific probes for the chromosomes 1, 7, 8, 10, 17, and X and locus-specific probes for 3q27 (BCL6), 5p15.2 (D5S23), 5q31 (EGR1), 10q23.3 (PTEN), and 14q32 (IGH@) on formalin-fixed paraffin-embedded tissue microarray sections. Our hypothesis of an increased level of CIN in BRCA1-associated breast cancer could not be confirmed by this approach. Surprisingly, we detected no significant difference in the extent of CIN in BRCA1-mutated versus sporadic tumors. The only exception was the CIN value for chromosome 1. Here, the extent of CIN was slightly higher in the group of sporadic tumors.
This review summarizes important information on the ectoenzyme tissue-nonspecific alkaline phosphatase (TNAP) and gives a brief insight into the symptoms, diagnostics, and treatment of the rare disease Hypophosphatasia (HPP), which is resulting from mutations in the TNAP encoding ALPL gene. We emphasize the role of TNAP beyond its well-known contribution to mineralization processes. Therefore, above all, the impact of the enzyme on central molecular processes in the nervous system and on inflammation is presented here.
Untersuchungen zur Informationsweitergabe in Familien mit erblichem Brust- und Eierstockkrebs
(2021)
Für die hier beschriebene Studie wurde ein Fragebogen erstellt, welcher von 80 Trägerinnen und Trägern einer pathogenen Mutation in den Genen BRCA1 oder BRCA2 ausgefüllt wurde. Die Befragung sollte untersuchen, ob den Befragten das Risiko ihrer Verwandten, ebenso Mutationsträger zu sein, bewusst war. Weiterhin sollte ermittelt werden, ob sie die jeweiligen Risikopersonen darüber informierten. Es zeigte sich, dass den meisten Befragten dieses Risiko bekannt war. Einigen Personen schienen jedoch nicht genau zu wissen, welche Verwandten als „Risikopersonen“ zählen. Insbesondere war nicht allen Befragten die Möglichkeit bewusst, dass auch Männer die Mutation tragen und an ihre Kinder weitergeben sowie selbst an Brustkrebs erkranken können. Weiterhin gaben mehr als ein Viertel der Befragten an, dass sie mindestens ein Familienmitglied, obwohl es ihnen als Risikoperson bekannt war, nicht informierten. Als häufigste Grund hierfür wurde mangelnder Kontakt genannt. Vor dem Hintergrund der Angaben der Befragten sowie der aktuellen Forschungslage werden in der vorliegenden Arbeit Möglichkeiten diskutiert, wie die Anzahl der informierten Angehörigen verbessert werden könnte.
Isolation of a Cancer-Associated Microchromosome in the Sperm-Dependent Parthenogen Poecilia formosa
(2011)
In the asexual all-female fish species Poecilia formosa, the Amazon molly, supernumerary chromosomes have frequently been found in both laboratory-reared and wild-caught individuals. While wild-caught individuals with B chromosomes are phenotypically indifferent from conspecifics, individuals carrying B chromosomes from recent introgression events in the laboratory show phenotypic changes. Former analyses showed that the expression of a pigment cell locus is associated with the presence of these B chromosomes. In addition, they contain a so far unidentified locus that confers a higher susceptibility to tumor formation in the presence of pigmentation pattern. Isolation by microdissection and hybridization to metaphase chromosomes revealed that they contain one or several sequences with similarity to a highly repetitive pericentromeric and subtelomeric sequence in A chromosomes. Isolation of one particular sequence by AFLP showed that the B chromosomes contain at least 1 copy of an A-chromosomal region which is highly conserved in the whole genus Poecilia, i.e. more than 5 million years old. We propose it to be a single copy sequence.
Trisomy 22 is a common trisomy in spontaneous abortions. In contrast, live-born trisomy 22 is rarely seen due to severe organ malformations associated with this condition. Here, we report on a male infant with complete, non-mosaic trisomy 22 born at 35 + 5 weeks via caesarean section. Peripheral blood lymphocytes and fibroblasts showed an additional chromosome 22 in all metaphases analyzed (47,XY,+22). In addition, array CGH confirmed complete trisomy 22. The patient’s clinical features included dolichocephalus, hypertelorism, flattened nasal bridge, dysplastic ears with preauricular sinuses and tags, medial cleft palate, anal atresia, and coronary hypospadias with scrotum bipartitum. Essential treatment was implemented in close coordination with the parents. The child died 29 days after birth due to respiratory insufficiency and deterioration of renal function. Our patient’s history complements other reports illustrating that children with complete trisomy 22 may survive until birth and beyond.
Supernumerary (B) chromosomes are dispensable elements found in many eukaryote genomes in addition to standard (A) chromosomes. In many respects, B chromosomes resemble sex chromosomes, so that a common ancestry for them has frequently been suggested. For instance, B chromosomes in grasshoppers, and other insects, show a pycnotic cycle of condensation-decondensation during meiosis remarkably similar to that of the X chromosome. In some cases, B chromosome size is even very similar to that of the X chromosome. These resemblances have led to suggest the X as the B ancestor in many cases. In addition, sex chromosome origin from B chromosomes has also been suggested. In this article, we review the existing evidence for both evolutionary pathways, as well as sex differences for B frequency at adult and embryo progeny levels, B chromosome effects or B chromosome transmission. In addition, we review cases found in the literature showing sex-ratio distortion associated with B chromosome presence, the most extreme case being the paternal sex ratio (PSR) chromosomes in some Hymenoptera. We finally analyse the possibility of B chromosome regularisation within the host genome and, as a consequence of it, whether B chromosomes can become regular members of the host genome.
Xenopus laevis (XLA) is an allotetraploid species which appears to have undergone whole-genome duplication after the interspecific hybridization of 2 diploid species closely related to Silurana/Xenopus tropicalis (XTR). Previous cDNA fluorescence in situ hybridization (FISH) experiments have identified 9 sets of homoeologous chromosomes in X. laevis, in which 8 sets correspond to chromosomes 1-8 of X. tropicalis (XTR1-XTR8), and the last set corresponds to a fusion of XTR9 and XTR10. In addition, recent X. laevis genome sequencing and BAC-FISH experiments support this physiological relationship and show no gross chromosome translocation in the X. laevis karyotype. Therefore, for the benefit of both comparative cytogenetics and genome research, we here propose a new chromosome nomenclature for X. laevis based on the phylogenetic relationship and chromosome length, i.e. XLA1L, XLA1S, XLA2L, XLA2S, and so on, in which the numbering of XLA chromosomes corresponds to that in X. tropicalis and the postfixes ‘L' and ‘S' stand for ‘long' and ‘short' chromosomes in the homoeologous pairs, which can be distinguished cytologically by their relative size. The last chromosome set is named XLA9L and XLA9S, in which XLA9 corresponds to both XTR9 and XTR10, and hence, to emphasize the phylogenetic relationship to X. tropicalis, XLA9_10L and XLA9_10S are also used as synonyms.
Two 5-methylcytosine (5-MeC)-rich heterochromatic regions were demonstrated in metaphase chromosomes of the Indian muntjac by indirect immunofluorescence using a monoclonal anti-5-MeC antibody. The metaphases were obtained from diploid and triploid cell lines. A major region is located in the ‘neck' of the 3;X fusion chromosome and can be detected after denaturation of the chromosomal DNA with UV-light irradiation for 1 h. It is located exactly at the border of the X chromosome and the translocated autosome 3. A minor region is found in the centromeric region of the free autosome 3 after denaturing the chromosomal DNA for 3 h or longer. The structure and possible function of the major hypermethylated region as barrier against spreading of the X-inactivation process into the autosome 3 is discussed.
Multicolor spectral analysis (spectral karyotyping) was applied to mitotic and male diakinetic chromosomes of hybrid mice carrying a unique system of 18 autosomal Robertsonian translocation chromosomes with alternating arm homologies. Only the autosomes 19 and the XY sex chromosomes are excluded from these Robertsonian translocations. The translocations, previously identified by conventional banding analyses, could be verified by spectral karyotyping. Besides the Robertsonian translocations, no other interchromosomal rearrangements were detected. In diakineses of male meiosis, the 18 metacentric Robertsonian translocation chromosomes form a very large meiotic ‘superring'. The predictable, specific order of the chromosomes along this ‘superring' was completely confirmed by multicolor spectral analysis. In the majority of diakineses analyzed, the free autosomal bivalent 19 and the XY sex bivalent form a conspicuous complex which tightly associates with the 12;14 Robertsonian translocation chromosome in the ‘superring'.
Polyploidy in Amphibia
(2015)
This review summarizes the current status of the known extant genuine polyploid anuran and urodelan species, as well as spontaneously originated and/or experimentally produced amphibian polyploids. The mechanisms by which polyploids can originate, the meiotic pairing configurations, the diploidization processes operating in polyploid genomes, the phenomenon of hybridogenesis, and the relationship between polyploidization and sex chromosome evolution are discussed. The polyploid systems in some important amphibian taxa are described in more detail.
Sites and amounts of 5-methylcytosine (5-MeC)-rich chromosome regions were detected in the karyotypes of 9 Brazilian species of Characiformes fishes by indirect immunofluorescence using a monoclonal anti-5-MeC antibody. These species, belonging to the genera Leporinus, Triportheus and Hoplias, are characterized by highly differentiated and heteromorphic ZW and XY sex chromosomes. In all species, the hypermethylated regions are confined to constitutive heterochromatin. The number and chromosome locations of hypermethylated heterochromatic regions in the karyotypes are constant and species-specific. Generally, heterochromatic regions that are darkly stained by the C-banding technique are distinctly hypermethylated, but several of the brightly fluorescing hypermethylated regions merely exhibit moderate or faint C-banding. The ZW and XY sex chromosomes of all 9 analyzed species also show species-specific heterochromatin hypermethylation patterns. The analysis of 5-MeC-rich chromosome regions contributes valuable data for comparative cytogenetics of closely related species and highlights the dynamic process of differentiation operating in the repetitive DNA fraction of sex chromosomes.
An experimental approach using monoclonal anti-5-methylcytosine (5-MeC) antibodies and indirect immunofluorescence was elaborated for detecting 5-MeC-rich chromosome regions in anuran chromosomes. This technique was applied to mitotic metaphases of 6 neotropical frog species belonging to 6 genera and 4 families. The hypermethylation patterns were compared with a variety of banding patterns obtained by conventional banding techniques. The hypermethylated DNA sequences are species-specific and located exclusively in constitutive heterochromatin. They are found in centromeric, pericentromeric, telomeric, and interstitial positions of the chromosomes and adjacent to nucleolus organizer regions. 5-MeC-rich DNA sequences can be embedded both in AT- and GC-rich repetitive DNA. The experimental parameters that have major influence on the reproducibility and quality of the anti-5-MeC antibody labeling are discussed.
The mitotic chromosomes of 11 species from the anuran families Centrolenidae and Allophrynidae were analyzed by means of conventional staining, banding techniques, and in situ hybridization. The amount, location, and fluorochrome affinities of constitutive heterochromatin, the number and positions of nucleolus organizer regions, and the patterns of telomeric DNA sequences were determined for most of the species. The karyotypes were found to be highly conserved with a low diploid chromosome number of 2n = 20 and morphologically similar chromosomes. The sister group relationship between the Centrolenidae and Allophrynidae (unranked taxon Allocentroleniae) is clearly corroborated by the cytogenetic data. The existence of heteromorphic XY♂/XX♀ sex chromosomes in an initial stage of morphological differentiation was confirmed in Vitreorana antisthenesi. The genome sizes of 4 centrolenid species were determined using flow cytometry. For completeness and for comparative purposes, all previously published cytogenetic data on centrolenids are included.
Mitotic chromosomes of 16 species of the frog genus Xenopus were prepared from kidney and lung cell cultures. In the chromosomes of 7 species, high-resolution replication banding patterns could be induced by treating the cultures with 5-bromodeoxyuridine (BrdU) and deoxythymidine (dT) in succession, and in 6 of these species the BrdU/dT-banded chromosomes could be arranged into karyotypes. In the 3 species of the clade with 2n = 20 and 4n = 40 chromosomes (X. tropicalis, X. epitropicalis, X. new tetraploid 1), as well as in the 3 species with 4n = 36 chromosomes (X. laevis, X. borealis, X. muelleri), the BrdU/dT-banded karyotypes show a high degree of homoeology, though differences were detected between these groups. Translocations, inversions, insertions or sex-specific replication bands were not observed. Minor replication asynchronies found between chromosomes probably involve heterochromatic regions. BrdU/dT replication banding of Xenopus chromosomes provides the landmarks necessary for the exact physical mapping of genes and repetitive sequences. FISH with an X. laevis 5S rDNA probe detected multiple hybridization sites at or near the long-arm telomeric regions in most chromosomes of X. laevis and X. borealis, whereas in X. muelleri, the 5S rDNA sequences are located exclusively at the long-arm telomeres of a single chromosome pair. Staining with the AT base pair-specific fluorochrome quinacrine mustard revealed brightly fluorescing heterochromatic regions in the majority of X. borealis chromosomes which are absent in other Xenopus species.
The mitotic chromosomes of 4 anuran species were examined by various classical banding techniques and by fluorescence in situ hybridization using a (TTAGGG)\(_n\) repeat. Large intrachromosomal telomeric sequences (ITSs) were demonstrated in differing numbers and chromosome locations. A detailed comparison of the present results with numerous published and unpublished data allowed a consistent classification of the various categories of large ITSs present in the genomes of anurans and other vertebrates. The classification takes into consideration the total numbers of large ITSs in the karyotypes, their chromosomal locations and their specific distribution patterns. A new category of large ITSs was recognized to exist in anuran species. It consists of large clusters of ITSs located in euchromatic chromosome segments, which is in clear contrast to the large ITSs in heterochromatic chromosome regions known in vertebrates. The origin of the different categories of large ITSs in heterochromatic and euchromatic chromosome regions, their mode of distribution in the karyotypes and evolutionary fixation in the genomes, as well as their cytological detection are discussed.
Correct imprinting is crucial for normal fetal and placental development in mammals. Experimental evidence in animal models and epidemiological studies in humans suggest that assisted reproductive technologies (ARTs) can interfere with imprinted gene regulation in gametogenesis and early embryogenesis. Bos taurus is an agriculturally important species in which ARTs are commonly employed. Because this species exhibits a similar preimplantation development and gestation length as humans, it is increasingly being used as a model for human germ-cell and embryo development. However, in contrast to humans and mice, there is relatively little information on bovine imprinted genes. Here, we characterized the bovine intergenic IGF2-H19 imprinting control region (ICR) spanning approximately 3 kb. We identified a 300-bp differentially methylated region (DMR) approximately 6 kb upstream of the H19 promoter, containing a CpG island with CTCF-binding site and high sequence similarity with the human intergenic ICR. Additional differentially methylated CpG islands lie –6 kb to –3 kb upstream of the promoter, however these are less conserved. Both classical bisulfite sequencing and bisulfite pyrosequencing demonstrated complete methylation of the IGF2-H19 ICR in sperm, complete demethylation in parthenogenetic embryos having only the female genome, and differential methylation in placental and somatic tissues. In addition, we established pyrosequencing assays for the previously reported bovine SNRPN and PEG3 DMRs. The observed methylation patterns were consistent with genomic imprinting in all analyzed tissues/cell types. The identified IGF2-H19 ICR and the developed quantitative methylation assays may prove useful for further studies on the relationship between ARTs and imprinting defects in the bovine model.
Mitotic and meiotic chromosomes of 5 species of the reptile genus Gonatodes are described by means of conventional staining, banding analyses and in situ hybridization using a synthetic telomeric DNA probe. The amount, location and fluorochrome affinities of constitutive heterochromatin, the number and positions of nucleolus organizer regions, and the patterns of telomeric DNA sequences were determined for most of the species. The karyotypes of G. falconensis and G. taniae from northern Venezuela are distinguished by their extraordinarily reduced diploid chromosome number of 2n = 16, which is the lowest value found so far in reptiles. In contrast to most other reptiles, both species have exclusively large biarmed (meta- and submetacentric) chromosomes. Comparison of the karyotypes of G. falconensis and G. taniae with those of other Gonatodes species indicates that the exceptional 2n = 16 karyotype originated by a series of 8 centric fusions. The karyotypes of G. falconensis and G. taniae are further characterized by the presence of considerable amounts of (TTAGGG)<sub>n</sub> telomeric sequences in the centromeric regions of all chromosomes. These are probably not only relics of the centric fusion events, but a component of the highly repetitive DNA in the constitutive heterochromatin of the chromosomes. The genome sizes of 4 Gonatodes species were determined using flow cytometry. For comparative purposes, all previously published cytogenetic data on Gonatodes and other sphaerodactylids are included and discussed.