Filtern
Gehört zur Bibliographie
- ja (237)
Erscheinungsjahr
Dokumenttyp
- Artikel / Aufsatz in einer Zeitschrift (127)
- Dissertation (109)
- Buch (1)
Schlagworte
- Fanconi-Anämie (13)
- Molekulargenetik (10)
- DNA methylation (9)
- Fanconi Anämie (9)
- DNS-Reparatur (8)
- Erbkrankheit (8)
- Epigenetik (7)
- Fanconi anemia (7)
- Brustkrebs (6)
- Mutation (6)
- breast cancer (6)
- BRCA1 (5)
- DNA repair (5)
- Genetik (5)
- Genmutation (5)
- genetics (5)
- heterochromatin (5)
- mutations (5)
- ovarian cancer (5)
- spectral karyotyping (5)
- Altern (4)
- Anura (4)
- BMD (4)
- Fanconi Anemia (4)
- Humangenetik (4)
- Makuladegeneration (4)
- Methylierung (4)
- Muskeldystrophie (4)
- Netzhaut (4)
- Zellzyklus (4)
- cancer (4)
- consortium (4)
- epigenetics (4)
- gene (4)
- mutation (4)
- sex chromosomes (4)
- zebrafish (4)
- 5-Methylcytosine (3)
- B chromosomes (3)
- DMD (3)
- Duchenne-Syndrom (3)
- Durchflusszytometrie (3)
- FISH (3)
- FSHD (3)
- Fanconi Anaemia (3)
- Genanalyse (3)
- Hereditäres Angioödem (3)
- Hämophilie A (3)
- Medizin (3)
- VKORC1 (3)
- VMD2 (3)
- Vitamin K (3)
- duchenne muscular dystrophy (3)
- fanconi anemia (3)
- genetic modifiers (3)
- genome-wide association (3)
- immunofluorescence (3)
- investigators (3)
- methylation (3)
- mosaicism (3)
- next generation sequencing (3)
- screening (3)
- sperm (3)
- susceptibility loci (3)
- whole exome sequencing (3)
- 2B (2)
- 5-Azadeoxycytidin (2)
- 5-azadeoxycytidine micronucleus (2)
- ADHD (2)
- AMD (2)
- Angiogenese (2)
- BRCA2 (2)
- C1-Inhibitor (2)
- Candida albicans (2)
- Chromosomenverlust (2)
- Cytogenetik (2)
- DM2 (2)
- DNA (2)
- DNA Reparatur (2)
- DNA-Repair (2)
- DNA-Reparatur (2)
- DNA-Reparaturgene (2)
- Duchenne (2)
- Duchenne muscular dystrophy (2)
- Eierstockkrebs (2)
- Epigenetics (2)
- Exomsequenzierung (2)
- Fabry disease (2)
- Fabry genotype (2)
- Fabry phenotype (2)
- Gen (2)
- Gentherapie (2)
- Geschlechtschromosomen (2)
- Gliedergürtelmuskeldystrophie (2)
- HAE (2)
- HPP (2)
- Hypophosphatasie (2)
- Hämophilie (2)
- Hörstörung (2)
- Ionisierende Strahlung (2)
- Keimzellmosaik (2)
- Konsanguinität (2)
- Maus (2)
- Mausmodell (2)
- Mikronukleus (2)
- Mitomycin C (2)
- Molekularbiologie (2)
- Morbus Best (2)
- Mosaik (2)
- Mosaizismus (2)
- Mutationsanalyse (2)
- Mutationsraten (2)
- Next generation sequencing (2)
- PCR (2)
- Positionsklonierung (2)
- Progeria adultorum (2)
- Proteine (2)
- RAD51C (2)
- RPE (2)
- Retinoschisis (2)
- Reversion (2)
- SLX4 (2)
- STR profile (2)
- Screening (2)
- Spermium (2)
- TNAP (2)
- VKORC1L1 (2)
- Vitamin-K-Gruppe (2)
- Warfarin (2)
- Würzburg / Institut für Humangenetik (2)
- Zellzyklusanalyse (2)
- ageing (2)
- angiogenesis (2)
- association (2)
- bisulfite pyrosequencing (2)
- blood (2)
- cardiogenetics (2)
- case report (2)
- cell cycle analysis (2)
- chromosome loss (2)
- common variants (2)
- congenital myopathy (2)
- copy number variation (2)
- cytogenetics (2)
- deafness (2)
- degeneration (2)
- diagnosis (2)
- disease genetics (2)
- exome sequencing (2)
- extracellular matrix (2)
- fetal programming (2)
- fibrosis (2)
- flow cytometry (2)
- flowcytometry (2)
- gene therapy (2)
- genetic diagnostics (2)
- genetic heterogeneity (2)
- genome (2)
- genomic imprinting (2)
- genotype-phenotype correlation (2)
- gestational diabetes mellitus (2)
- gridle muscular-dystrophy (2)
- growth hormone deficiency (2)
- hearing impairment (2)
- hearing loss (2)
- hemophilia (2)
- hepatic stellate cell (2)
- hereditary angioedema (2)
- hereditary breast and ovarian cancer (2)
- hypophosphatasia (2)
- immunodeficiency (2)
- incidence (2)
- intellectual disability (2)
- liver (2)
- macular degeneration (2)
- microcephaly (2)
- mineralization (2)
- modifiers (2)
- mouse model (2)
- myofibrillar myopathy (2)
- myofibroblast (2)
- myopathy (2)
- nervous system (2)
- next generation sequencing (NGS) (2)
- positional cloning (2)
- protein (2)
- recombination (2)
- retina (2)
- risk (2)
- sensory neuropathy (2)
- single-nucleotide polymorphisms (2)
- skeletal dysplasia (2)
- somatic mosaicism (2)
- sperm DNA methylation (2)
- survival (2)
- variants (2)
- vitamin K (2)
- whole-exome sequencing (2)
- (classical and atypical) Werner syndrome (1)
- - (1)
- 16q22 (1)
- 3D modeling (1)
- 3R (1)
- 5-methylcytosine (1)
- A chromosomes (1)
- AFLP (1)
- AKT-signaling (1)
- ALPL (1)
- ART outcome (1)
- ATM (1)
- ATM gene (1)
- Achondroplasie (1)
- Adulthood (1)
- Age-related macular degeneration (1)
- Alburnus alburnus (1)
- Alkalische Phosphatase (1)
- Allelfrequenzen (1)
- Allgemeinmedizin (1)
- Allophrynidae (1)
- Alter (1)
- Alternatives Spleißen (1)
- Alzheimer's disease (1)
- Alzheimerkrankheit (1)
- Anticoagulants (1)
- Antralfollikel (1)
- Apolipoprotein (1)
- Array-Methoden (1)
- Arrhythmie (1)
- Asisted Reproduction (1)
- Aspergillus fumigatus (1)
- Assoziation (1)
- Ataxia telangiectasia (1)
- Ataxia telangiectasia (AT) (1)
- Ataxia teleangiectatica (1)
- Autism (1)
- Autism spectrum disorders (1)
- Autismus (1)
- B Chromosomen (1)
- B-Chromosom (1)
- B4GALT7 gene (1)
- BRCA1/2 (1)
- BRIP1 gene (1)
- Becker (1)
- Becker-Kiener-Syndrom (1)
- Berechnung (1)
- Best Disease (1)
- Best's Disease (1)
- Best-Krankheit (1)
- Bioinformatics (1)
- Blasentumor (1)
- Blutgefäß (1)
- Blutgerinnung (1)
- Brachmann-de-Lange-Syndrom (1)
- Brachmann-de-Lange-syndrome (1)
- BrdU replication banding pattern (1)
- BrdU/dT replication banding (1)
- Breastcancer (1)
- Brüder Grimm (1)
- C1 Inhibitor (1)
- C1-INH (1)
- C1-inhibitor (1)
- C1INH (1)
- C282Y (1)
- CAGSSS (1)
- CCM (1)
- CCM1/Krit1 (1)
- CCM2/Malcavernin (1)
- CCM3 (1)
- CCM3/PDCD10 (1)
- CDC14A (1)
- CIN (1)
- CLRN2 (1)
- CNV (1)
- COVID-19 (1)
- CRISPR-Cas Systems (1)
- CRISPR/Cas9 (1)
- Calcyon (1)
- Carcinogenese (1)
- Caretaker gene syndromes (1)
- Caretaker-Gen-Syndrome (1)
- Cathepsin (1)
- Cellcycle (1)
- Cellcycle blockage (1)
- Central Core Disease (1)
- Centrolenidae (1)
- Charcot-Marie-Tooth neuropathy 1A (1)
- Checkpoints (1)
- Chromosomal instability (1)
- Chromosomenaberration (1)
- Chromosomenaberrationen (1)
- Chromosomenbruch (1)
- Chromosomenbruchanalyse (1)
- Chromosomenbruchsyndrome (1)
- Chromosomeninstabilitätssyndrome (1)
- Chromosomenkondensation (1)
- Coagulation factor IX (1)
- Coexpression (1)
- Cognitive control (1)
- Copy number variation (1)
- Cornelia-de-Lange-Syndrom (1)
- Cornelia-de-Lange-syndrome (1)
- Cortex (1)
- Coumarin (1)
- Cranial sutures (1)
- Craniosynostosis (1)
- Creatin Kinase (1)
- Cyrillic 2.13 (1)
- D4Z4 partial deletion (1)
- DFNB32 (1)
- DFNB68 (1)
- DLX5/6 (1)
- DM1 (1)
- DNA Methylation (1)
- DNA damage (1)
- DNA damage response (1)
- DNA double-strand break (1)
- DNA methylation (DNAm) age (1)
- DNA methylation dynamics (1)
- DNA methyltransferase gene (1)
- DNA purification (1)
- DNA repair defect (1)
- DNA repair gene (1)
- DNA repair genes (1)
- DNA sequencing (1)
- DNA-Instabilitätssyndrom (1)
- DNA-Methylation (1)
- DNA-Methylierung (1)
- DNA-Quervernetzung (1)
- DNA-Sequenz (1)
- DNA-repair (1)
- DNA-repair genes (1)
- DNER (1)
- DNMT3B (1)
- DNS-Methyltransferase (1)
- DYNC1I1 (1)
- Damage (1)
- Deflazacort (1)
- Degeneration (1)
- Deletion <Genetik> (1)
- Deutschland (1)
- Deutschland / Stammzellgesetz (1)
- Development (1)
- Diagnostik (1)
- Dickdarmkrebs (1)
- Dickkopf-1 (1)
- Domäne <Biochemie> (1)
- Down Syndrom (1)
- Down syndrome (1)
- Down´s Syndrome (1)
- Duchenne dystrphy (1)
- Duchennesche Muskeldystrophie (1)
- Duchflußzytophotometrie (1)
- Durchflusscytometrie (1)
- Durchflußzytometrie (1)
- Dystrophin Gene (1)
- Dystrophin-Gen (1)
- ELISA (1)
- ENaC (1)
- ERCC1-XPF (1)
- ERCC4 (1)
- EU (1)
- Embryos (1)
- Endothelzelle (1)
- Endothelzellen (1)
- Enhancer elements (1)
- Entwicklung (1)
- Environment (1)
- Epigenetische Uhr (1)
- Epigenotypus (1)
- Epimutation (1)
- Epimutationen (1)
- Erbkrankheiten (1)
- Erfolg (1)
- Erkrankungswahrscheinlcihkeit (1)
- Erwachsener (1)
- Evans syndrome (1)
- Excelsior Cyrillic (1)
- Extracellular matrix (1)
- FA (1)
- FAAP100 (1)
- FADS1 (1)
- FADS2 (1)
- FADS3 (1)
- FANCA (1)
- FANCD2 (1)
- FANCO (1)
- FANCP (1)
- FAP-1/PTPN13 (1)
- Fabry (1)
- Fabry-Krankheit (1)
- Fallbeispiele (1)
- Familial Beckwith-Wiedemann syndrome (1)
- Familial breast cancer (1)
- Familienanamnese (1)
- Familiärer Brustkrebs (1)
- Fanconi Anämie (FA) (1)
- Fanconi Anämie A (1)
- Fanconi anaemia (1)
- Fanconi anemia (FA) (1)
- Fanconi anemia A (1)
- Fanconi-anemia subtype (1)
- Fetal brain development (1)
- Fetale Programmierung (1)
- Fettsäuredesaturasen (1)
- Fluorescence in situ hybridization (1)
- Fluorescence-in-situ-hybridization (1)
- Fluoreszenz-in-situ-Hybridisierung (1)
- Fourthcorner analysis (1)
- Frequency (1)
- Frontal cortex (1)
- Fruchtwasserzellen (1)
- Frühgeburt (1)
- Förderung (1)
- G0/G1 (1)
- G2 Phase (1)
- GTL2 (1)
- Gamma-Carboxylase (1)
- Gecko (1)
- Gefäßkrankheit (1)
- Gehirn (1)
- Gen BRCA 1 (1)
- Gen BRCA 2 (1)
- Gendefekt (1)
- Gene-expression (1)
- Genes (1)
- Genetische Variabilität (1)
- Genetisches Imprinting (1)
- Genetisches Modell (1)
- Genom (1)
- Genome (1)
- Genort (1)
- Genotyp (1)
- Genotyp-Phänotyp Korrelation (1)
- Genotype-phenotype association (1)
- Genotype–phenotype correlations (1)
- Genotypisierung (1)
- Genpanel (1)
- Genregulation (1)
- Gerinnungsfaktor VIII (1)
- Gesundheitsstatus (1)
- Gesundheitssystem (1)
- Gewebe-unspezifische Alkalische Phosphatase (1)
- Glaukom (1)
- H63D (1)
- Hereditary Angioedem (1)
- Hereditary breast cancer (1)
- Herzmuskelkrankheit (1)
- High throughput screening (1)
- High-throughput data (1)
- Hindbrain (1)
- Histologic grade (1)
- Holliday junction reolvass (1)
- Holstein <Rind> (1)
- Human genetics (1)
- Human prefrontal cortex (1)
- Hypermutabilität (1)
- Hypophosphatasia (1)
- Hämatopoese (1)
- Hämochromatose (1)
- Hörstörungen (1)
- IARS2 (1)
- ICF2 (1)
- ICL (1)
- ICSI (1)
- IGF2-H19 (1)
- II citrullinemia (1)
- IMSI (1)
- Immunfluoreszenz (1)
- Imprinting (1)
- Indian muntjac (1)
- Induced Pluripotent Stem Cells (1)
- Infertilität (1)
- Informationsweitergabe (1)
- Instability (1)
- Instabilität (1)
- Inzidenz <Medizin> (1)
- Inzidenzschätzung (1)
- Jugendliche und erwachsene Probanden (1)
- KRAS (1)
- Kandidatengen (1)
- Kasuistik (1)
- Kathepsin (1)
- Kathepsin B (1)
- Kathepsin L (1)
- Kavernom (1)
- Keimzell- und Embryonalentwicklung (1)
- Knochenmarksversagen (1)
- Koagulopathie (1)
- Kolorektales Karzinom (1)
- Koppelungsanalyse (1)
- Korrelation (1)
- Kraniosynostosen (1)
- Krebserkrankungen (1)
- LGMDR5 (1)
- Landouzy-Déjerine-Atrophie (1)
- Langdon Down (1)
- Lautwahrnehmung (1)
- Legasthenie (1)
- Lese-Rechtschreibstörung (1)
- Limb development (1)
- Limb girdle muscular dystrophy (LGMD) (1)
- Long-term follow-up (1)
- MCPH1 (1)
- MEK/ERK-signaling (1)
- MFM (1)
- MPP4 (1)
- MPP5 (1)
- MTM (1)
- Makula (1)
- Makuladystrophie (1)
- Male breast cancer (1)
- Malignant Hyperthermia (1)
- Malignant neoplasms (1)
- Maligne Hyperthermie (1)
- Massive parallele Sequenzierung (1)
- Medizinisches Versorgungszentrum (1)
- Meiose (1)
- Membranproteine (1)
- Mensch (1)
- Mentale Retardierung (1)
- Methylome (1)
- Microarray (1)
- Microarray analysis (1)
- Microdeletions (1)
- Mikroarray (1)
- Mikrodeletionen (1)
- Mikrozephalie (1)
- Missbildung (1)
- Miyoshi myopathy (1)
- Molecular approaches (1)
- Monogen (1)
- Morbidität (1)
- Mortalität (1)
- Mucopolysaccharidosis IIIa (1)
- Muenke-Syndrom (1)
- Multiple displacement amplification (1)
- Multiproteinkomplex (1)
- Multivariate analysis (1)
- Muscular Dystrophy (1)
- Muskeldystrophie Becker (1)
- Muskeldystrophie Duchenne (1)
- Muskelentwicklung (1)
- Muskelkrankheit (1)
- Mutationen (1)
- Mutationrate (1)
- Mutationsrate (1)
- Mutationswahrscheinlichkeit (1)
- Myotone Dystrophie (1)
- Myotone Dystrophy (1)
- Myotonische Dystrophie (1)
- Myotubular Myopathy (1)
- Myotubuläre Myopathie (1)
- N170 (1)
- NEIL2 (1)
- NLS-Sequenz (1)
- Na-K-ATPase (1)
- Nachtblindheit (1)
- Neanderthal (1)
- Neisseria meningitidis (1)
- Nervendegeneration (1)
- Netzhautdegeneration (1)
- Neue Fanconi Anämie Gene (1)
- Neurodegeneration (1)
- Neuroepigenomics (1)
- Neuromuskuläre Erkrankungen (1)
- Neuromuskuläre Krankheit (1)
- Neurons (1)
- Next Generation Sequencing (1)
- Next Generation Sequencing (NGS) (1)
- Next-Generation Sequencing (1)
- Nibrin (1)
- Nijmegen Breakage Syndrom (1)
- Nijmegen Breakage Syndrom (NBS) (1)
- Nijmegen breakage syndrome (1)
- Nijmegen breakage syndrome (NBS) (1)
- Nonsensmutation (1)
- North Carolina Makuladystrophie (1)
- North Carolina Makuladystrophy (1)
- OGG1 (1)
- Oogenese (1)
- Oozyte (1)
- Ordination methods (1)
- Organisationsform (1)
- Ovarian (1)
- Ovariancancer (1)
- Ovarielle Alterung (1)
- P100 (1)
- PCC-Syndrom (1)
- PDZ domain (1)
- PROMM (1)
- PSD95 (1)
- Pakistan (1)
- Pankreaskarzinom (1)
- Parameter (1)
- Parameter <Mathematik> (1)
- Parent-of-origin (1)
- Paternal age effect (1)
- Pathology (1)
- Patterns (1)
- Penetranz (1)
- Pfeiffer-Syndrom (1)
- Phi29 Polymerase (1)
- Phänotyp (1)
- Phänotypische Heterogenität (1)
- Pigmentepithel (1)
- Plasma DNA (1)
- Pompe disease (1)
- Postovulatorische Alterung (1)
- Prenatal diagnosis (1)
- Primary Microcephaly (1)
- Primary care (1)
- Proteininteraktion (1)
- Proteomanalyse (1)
- Proximale myotone Myopathie (1)
- Pränataldiagnostik (1)
- Pränatale Diagnostik (1)
- Psychosoziale Beratung (1)
- Psychosoziale Situation (1)
- Punktmutation (1)
- Qualitätskontrolle (1)
- RAD50-Defizienz (1)
- RAD52 (1)
- RLQ analysis (1)
- RNA metabolism (1)
- RNA-Seq analysis (1)
- RPE specific genes (1)
- Rab14 (1)
- Radiosensitivität (1)
- Radiäre Drusen (1)
- Rechenprogramme (1)
- Regeneration (1)
- Rekombination (1)
- Remission (1)
- Renal abnormalities (1)
- Repair (1)
- Reproduktionsmedizin (1)
- Retina (1)
- Ribbon-Synapse (1)
- Risiko (1)
- Risikoberatung (1)
- Risikoberechnung (1)
- Risikoberechnungen (1)
- Risikofaktoren (1)
- Risk estimation (1)
- Robertsonian translocation chromosomes (1)
- Robertsonsche Translokation (1)
- Robin-Syndrom (1)
- Ryanodin-Rezeptor Gen (1)
- Ryanodine receptor gene (1)
- Ryr 1 (1)
- S1PR2 (1)
- SARS-CoV-2 (1)
- SH3 domain (1)
- SHFM (1)
- SKY analysis (1)
- SLC2A3 (1)
- SMA (1)
- SMN1-Kopien (1)
- SMN1-copies (1)
- SNP array (1)
- SNP-Array (1)
- SNP-microarray (1)
- SOX9 (1)
- STK25/SOK-1/YSK-1 (1)
- Sanger Sequenzierung (1)
- Sanger sequencing (1)
- Schizophrenia (1)
- Schwangerschaft (1)
- Schwangerschaftsdiabetes (1)
- Segregation (1)
- Selective attention (1)
- Self-renewal (1)
- Senile Makuladegeneration (1)
- Senile Makuladegeneration / Pigmentepithel / Genexpression (1)
- Sequenzdaten (1)
- Serpin (1)
- Skull (1)
- Somites (1)
- Spectral Karyotyping (1)
- Spermatogenese (1)
- Spermien (1)
- Spinale Muskelatrophie (1)
- Stability (1)
- Stammzelltransplantation (1)
- Staphylococcus aureus (1)
- Sterilität (1)
- Strahleninduzierte Genominstabilität (1)
- Strahlensensitivität (1)
- Suicidal behavior (1)
- Summe G2/GF (1)
- Susceptibility (1)
- TMEM43 (1)
- TRIM32 (1)
- TYPE-2 (1)
- Terminal 4q deletion syndrome (1)
- Th17 (1)
- Thrombozytopenie (1)
- Tight junction (1)
- Tissue Nonspecific Alkaline Phosphatase (1)
- Tnap (1)
- Transcription (1)
- Transcription regulation (1)
- Transcriptome (1)
- Transcriptomics (1)
- Transkription <Genetik> (1)
- Transkriptome (1)
- Tregs (1)
- Trisomie 21 (1)
- Trisomy 21 (1)
- UBZ (1)
- Ukelei (1)
- Ungarn (1)
- Urodela (1)
- Usher syndrome (1)
- VKCFD (1)
- Vater (1)
- Vektoren (1)
- Verteilung (1)
- Verwandtenehe (1)
- Visualization (1)
- Vitamin K epoxide reductase (1)
- Vitamin-K-Epoxid-Reduktase (1)
- Väterliches Alter (1)
- WDR62 mutation (1)
- Wachstumshormonmangel (1)
- Whole Genome Amplification (1)
- Working memory (1)
- X chromosome (1)
- X-Chromosom (1)
- X-Inaktivierung (1)
- X-chromosomal inactivation (1)
- X-gebundene juvenile Retinoschisis (1)
- X-linked juvenile retinoschisis (1)
- X. laevis-type karyotype (1)
- X. tropicalis-type karyotype (1)
- Xenopus (1)
- Xenopus laevis (1)
- Xenopus tropicalis (1)
- Y chromosome degeneration (1)
- ZBTB24 (1)
- ZBTB24 mutations (1)
- ZNF365 (1)
- ZW sex chromosomes (1)
- Zahnentwicklung (1)
- Zebrabärbling (1)
- Zebrafish (1)
- Zellkern (1)
- Zellkultur (1)
- Zellzyklus-Analyse (1)
- Zellzyklusdiagnostik (1)
- Zellzykluseffekte (1)
- Zentralfibrillen-Myopathie (1)
- Zytogenetik (1)
- abnormalities (1)
- adenoma (1)
- adrenal insufficiency (1)
- age (1)
- age at onset (1)
- age-related (1)
- age-related differentially methylated regions (ageDMRs) (1)
- aldehydes (1)
- allelefrequences (1)
- alleles (1)
- allopolyploidy (1)
- alternative methods (1)
- alternative splicing (1)
- altersabhängig (1)
- alu elements (1)
- alzheimers disease (1)
- amniotic fluid cells (1)
- amphiphysin-2 BIN1 (1)
- amplicon sequencing (1)
- antidepressants (1)
- anxiety disorders (1)
- apoptosis (1)
- array-CGH (1)
- arrhythmogenic cardiomyopathy (1)
- asexual reproduction (1)
- assistierte Reproduktion (1)
- ataxia telangiectasia (1)
- autism (1)
- autophagy (1)
- autopolyploidy (1)
- autosomal dominant (1)
- autosomal recessive (1)
- autosomal recessive hearing loss (1)
- autosomal recessive heredity (1)
- autosomal recessive non-synstromic hearing loss (1)
- autosomal rezessiver Erbgang (1)
- autosomal-dominant (1)
- autosomal-rezessiv (1)
- banding analyses (1)
- bcl-2 associated athanogene protein 3 (1)
- becker muscular dystrophy (1)
- behavior (1)
- bestrophin (1)
- bioinformatics and computational biology (1)
- bivariate Durchflußzytometrie (1)
- bleding disorders other than hemophilia (1)
- blood coagulation (1)
- blood plasma (1)
- bone development (1)
- bone marrow failure syndrome (1)
- bone-marrow failure (1)
- bovine (1)
- breakage (1)
- breast cancer predisposition (1)
- breast cancer predisposition genes (1)
- breast neoplasms (1)
- brothers grimm (1)
- cDNA Selektion (1)
- cDNA selection (1)
- cadherins (1)
- calculation (1)
- cancer treatment (1)
- candida genome database (1)
- candidate genes (1)
- cardiomyopathy (1)
- case reports (1)
- cataracts (1)
- cavernoma (1)
- cell (1)
- cell culture (1)
- cell cycle (1)
- cell cycle studies (1)
- cell death (1)
- cell division (1)
- cell wall (1)
- cellcycle (1)
- cellcycle effects (1)
- centromeric instability (1)
- childhood ataxia (1)
- childhood cancer (1)
- children (1)
- chip-seq (1)
- chromosomal aberrations (1)
- chromosomal abnormality (1)
- chromosomal breakage (1)
- chromosomal instability (1)
- chromosomal instability disorder (1)
- chromosomale Instabilität (1)
- chromosome (1)
- chromosome aberration (1)
- chromosome condensation (1)
- chromosome evolution (1)
- chromosome inversion (1)
- chromosome staining (1)
- chromsomal instability (1)
- classification (1)
- colorectal cancer (1)
- combined retinal dystrophy (1)
- comparative genomics (1)
- complex chromosome rearrangements (1)
- complex disorders (1)
- complex traits (1)
- computational prediction (1)
- congenital (1)
- congenital heart-deffects (1)
- connective tissue disorder (1)
- consanguinity (1)
- copy number variation (CNV) (1)
- copy-number variation (1)
- coumarin (1)
- craniosynostosis (1)
- cross-link repair (1)
- cytokinesis (1)
- cytometrie (1)
- cytotoxicity (1)
- damage (1)
- danazol (1)
- danio rerio (1)
- database (1)
- deep bisulfite sequencing (1)
- deficiency (1)
- delayed radiation effects (1)
- deletion (1)
- department of human genetics (1)
- design (1)
- development (1)
- developmental origins hypothesis (1)
- diagnostic delay (1)
- differentially methylated region (1)
- differentiation (1)
- diploidization (1)
- disease (1)
- disruption project (1)
- double trouble (1)
- double-strand DNA breaks (1)
- duplication (1)
- duplication-deficiency (1)
- dysferlinopathy (1)
- dyslexia (1)
- dystrophin (1)
- eExons (1)
- early respiratory-failure (1)
- elective surgery (1)
- elements (1)
- embryos (1)
- emotional disorders (1)
- endonuclease (1)
- endothelial cells (1)
- epigenetic heterogeneity (1)
- epimutation (1)
- epithelial cells (1)
- erblicher Brustkrebs (1)
- establishment (1)
- etiology (1)
- euchromatin (1)
- evolutionary biology (1)
- evolutionary fixation (1)
- expansion (1)
- expression signature (1)
- extreme phenotypes (1)
- facial anomalies (1)
- facio-scapulo-humeral dystrophie (1)
- facio-scapulo-humeralen Muskeldystrophie (1)
- facioscapulohumeral muscular dystrophy (1)
- factor VIII (1)
- familial breast cancer (1)
- families (1)
- family history of glaucoma (1)
- fanconi anaemia (1)
- fanconi-anemia (1)
- fatty acid desaturases (1)
- features (1)
- female (1)
- female Fabry patients (1)
- fetal brain development (1)
- fetal cord blood (1)
- fetal overnutrition (1)
- fibroblasts (1)
- filamin C (1)
- fine-scale mapping (1)
- fish (1)
- flies (1)
- flow (1)
- flow-cytometry (1)
- frameshift (1)
- frontal cortex (1)
- früher vorzeitiger Blasensprung (1)
- functional analysis (1)
- functional modules (1)
- g2 phase (1)
- gamma-carboxylation (1)
- geckos (1)
- gene defect (1)
- gene mutations (1)
- gene panel (1)
- genetic (1)
- genetic causes of cancer (1)
- genetic counselling (1)
- genetic diagnosis (1)
- genetic diseases (1)
- genetic interaction networks (1)
- genetic loci (1)
- genetic model (1)
- genetic risk (1)
- genetic skeletal disorders (1)
- genetic susceptibility (1)
- genetic testing (1)
- genetic variants (1)
- genetische Beratung (1)
- genetische Erkrankungen (1)
- genome sequencing (1)
- genome-wide association study (GWAS) (1)
- genome-wide linkage analysis (1)
- genomic analysis (1)
- genomic instability (1)
- genomic libraries (1)
- genomics (1)
- genotyping arrays (1)
- geprägte Gene (1)
- germ-line mosaicism (1)
- germinal mosaicism (1)
- germline mosaicism (1)
- germline mutations (1)
- glaucoma (1)
- global DNA methylation (1)
- glycogenin 1 (1)
- gonads (1)
- granulomas (1)
- great dane (1)
- growth (1)
- growth failure (1)
- gynogenesis (1)
- haemophilia a (1)
- hand/foot malformation (1)
- haplogroups (1)
- haploinsufficiency (1)
- helicase BRIP1 (1)
- hematology (1)
- hemostasis and thrombosis (1)
- hereditary breast cancer (1)
- hereditary hearing loss (1)
- hereditary hemochromatosis (1)
- hereditary motor (1)
- hereditary motor and sensory neuropathy (1)
- heterogeneity (1)
- heteromorphic sex chromosomes (1)
- heterozygote (1)
- histological subtype (1)
- homoeologous chromosomes (1)
- hormone-related protein (1)
- host cells (1)
- human disease (1)
- human evolution (1)
- human medicine (1)
- humane Keimzellen (1)
- humans (1)
- hybridogenesis (1)
- hypermethylated DNA (1)
- hypermethylation (1)
- hypermutable (1)
- hämatopoetisches Mosaik (1)
- illumina (1)
- immune thrombocytopenia (1)
- imprinting control region (1)
- in situ hybridization (1)
- in vitro model (1)
- induced pluripotent stem cells (1)
- infections (1)
- infinium HumanOmni1-Quad (1)
- inflammation (1)
- inflammatory diseases (1)
- inherited disorders (1)
- inherited myopathy (1)
- insulin treatment (1)
- integrierte Versorgung (1)
- intergenerational contraction (1)
- interolog (1)
- interstrand crosslink (1)
- intrachromosomal telomeric sequences (1)
- inversion (1)
- italian patients (1)
- juvenile and adult patients (1)
- karyotype evolution (1)
- kinase signaling (1)
- kongenitale Myopathie (1)
- limb development (1)
- limb-girdle dystrophie (1)
- limb-girdle muscular dystrophies (1)
- linkage analysis (1)
- linked myotubular myopathy (1)
- live-born (1)
- loss of chromosome Y (LOY); (1)
- lymphoma (1)
- lyso‐Gb3 (1)
- macrophages (1)
- macula dystrophy (1)
- macular (1)
- maintenance (1)
- makula (1)
- male breast cancer (1)
- malformations (1)
- malignant hyperthermia (1)
- maligne Hyperthermie (1)
- mammalian male germline (1)
- mammographic density (1)
- medical genetics (1)
- meiosis (1)
- meiotic chromosomes (1)
- meiotic ‘superring’ (1)
- membrane curvature (1)
- membrane repair (1)
- mental retardation (1)
- metabolic disease (1)
- metabolism (1)
- methylation array (1)
- methylation array analysis (1)
- mice (1)
- microdeletion syndrome (1)
- microdissection (1)
- midbody (1)
- middle aged (1)
- mineraliztion (1)
- missense mutations (1)
- mitomycin c (1)
- mitotic chromosomes (1)
- mixed hearing loss (1)
- mixed mutation mechanisms (1)
- miyoshi myopathy (1)
- model (1)
- moderate-penetrance genes (1)
- molecular analysis (1)
- molecular cloning (1)
- monogeneic (1)
- monoubiquitination (1)
- monozygotic twins (1)
- morbidity (1)
- mortality (1)
- mouse (1)
- multi protein complex (1)
- multiple diseases (1)
- multiple myeloma (1)
- muscle disease (1)
- muscle strength (1)
- muscular dystrophy (1)
- muscular-dystrophy (1)
- mutation analysis (1)
- mutation mechanism (1)
- mutation rate (1)
- mutationrate (1)
- myotonic dystrophie (1)
- myotonic dytsrophy (1)
- natural variation (1)
- network analysis (1)
- network inference (1)
- networks (1)
- neurodegeneration (1)
- neurodevelopmental disorders / genetics (1)
- neuromuscular disease (1)
- neurotransmission (1)
- neutrophils (1)
- next-generation sequencing (1)
- next-generation-sequencing (1)
- nibrin (1)
- night blindness (1)
- nomenclature (1)
- non-mosaic (1)
- non-sense mediated mRNA decay (1)
- nonspecific alkaline-phosphae (1)
- oocytes (1)
- organ toxicity (1)
- overlapping syndrome (1)
- oxidative stress (1)
- p.R245H (1)
- p.S298P (1)
- p53-deficient mice overexpressing TGFalpha (1)
- painful (1)
- pancreatic carcinoma (1)
- panel sequencing (1)
- panic disorder (1)
- paternal age (1)
- paternal age effect (1)
- paternal introgression (1)
- pathogen-host interaction (PHI) (1)
- pathogenicity (1)
- pathway (1)
- patient (1)
- penetrance (1)
- phalloidin stain (1)
- phenotype (1)
- phenotypic heterogeneity (1)
- phenotypic spectrum (1)
- phosphoproteome (1)
- photoreceptor synapse (1)
- phylloquinone (1)
- plasma (1)
- platelet disorders (1)
- polymerase chain reaction (1)
- polymorphism (1)
- polyneuropathy (1)
- polyploidization (1)
- polyploidy (1)
- population (1)
- potential role (1)
- prednisone (1)
- premature aging (1)
- premutation (1)
- prenatal diagnosis (1)
- preterm labor (1)
- preterm prelabor rupture of the membranes (1)
- prevalence (1)
- promoter methylation (1)
- promotes (1)
- protein aggregation (1)
- protein interaction (1)
- protein interaction database (1)
- protein-protein interaction (1)
- proteins (1)
- proteomic approach (1)
- proteomic signature (1)
- protocadherin gamma cluster (1)
- proximal myotonic myopathy (1)
- pseudotetraploidisierung (1)
- pseudotetraploidization (1)
- psychische Probleme (1)
- psychosocial aspects (1)
- psychosoziale Aspekte (1)
- qPCR (1)
- quality control (1)
- quantitative Polymerasekettenreaktion (1)
- radial drusen (1)
- radiation-induced genome instability (RIGI) (1)
- radiosensitivity (1)
- radiosensivitity (1)
- recessive inheritance (1)
- reciprocal translocation (1)
- recurrence (1)
- regulatory T cells (1)
- regulatory mutations (1)
- reproduction (1)
- requency (1)
- reversion (1)
- rhodamine–phalloidin stain (1)
- risk assesment (1)
- risk factors (1)
- robertsonian translocation (1)
- ryr 1 (1)
- sarcoglycanopathy (1)
- sarcotubular myopathy (1)
- secondary cancer (1)
- segmental progeria (1)
- selection (1)
- sensorineural (1)
- sensorineural hearing loss (1)
- sequence alignment (1)
- sequence assembly tools (1)
- sex chromosome evolution (1)
- sex linked pigmentation pattern (1)
- sex ratio (1)
- sexual antagonistic genes (1)
- short stature (1)
- skeletal muscle (1)
- skeletal myopathy (1)
- skewed (1)
- sky kinases (1)
- species-specific epigenetic marks (1)
- spektrale Karyotypenanalyse (1)
- spektrale karyotypisierung (1)
- spinal muscular atrophy (1)
- splicing (1)
- spondylodysplastic Ehlers-Danlos syndrome (1)
- stem cell transplantation (1)
- steroids (1)
- stress fibers (1)
- structural genome variations (1)
- submicroscopic chromosome rearrangement (1)
- subtypes (1)
- success (1)
- suppression (1)
- susceptibility (1)
- susceptibility alleles (1)
- susceptibility gene (1)
- synaptonemal complex (1)
- systemic sclerosis (1)
- targeted gene panel (1)
- teeth (1)
- telomere length (1)
- telomeres (1)
- temperature (1)
- testes (1)
- testosterone (1)
- therapy (1)
- throat (1)
- thrombozytopenia (1)
- tinnitus (1)
- tissue-specific enhancers (1)
- tooth development (1)
- transcription deficiency (1)
- transcriptional regulation (1)
- transcriptome (1)
- transgene TGFalpha-Mäuse (1)
- transgenic animals (1)
- transporter gene SLC2A3 (1)
- transposable elements (1)
- treatment (1)
- treatment guidelines (1)
- triple-negative breast cancer (1)
- triplosufficiency (1)
- trisomy 21 (1)
- trisomy 22 (1)
- tumor subtypes (1)
- tumor-suppressor (1)
- twin study (1)
- two-parameter flow cytometry (1)
- type II esophageal achalasia (1)
- ubiquitin (1)
- vacuolar myopathy (1)
- vascular malformation (1)
- vaskuläre Malformation (1)
- vectors (1)
- ventilation (1)
- vertebrate (1)
- werner syndrom (1)
- werner syndrome (1)
- whole genome amplification (1)
- whole genome sequencing (1)
- whole genome sequencing (WGS) (1)
- wuerzburg (1)
- zytogenetik (1)
- Änderung (1)
- Ätiologie (1)
- Ödem (1)
- α‐GalA 3D‐structure (1)
Institut
- Institut für Humangenetik (237) (entfernen)
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)
Western societies are steadily becoming older undergoing a clear trend of delayed parenthood. Children of older fathers have an undeniably higher risk for certain neurodevelopmental disorders and other medical conditions. Changes in the epigenetic landscape and especially in DNA methylation patterns are likely to account for a portion of this inherited disease susceptibility. DNA methylation changes during the ageing process are a well-known epigenetic feature. These so-called age-DMRs exist in developmentally important genes in the methylome of several mammalian species. However, there is only a minor overlap between the age-DMR datasets of different studies. We therefore replicated age-DMRs (which were obtained from a genome wide technique) by applying a different technical approach in a larger sample number. Here, this study confirmed 10 age-DMRs in the human and 4 in the bovine sperm epigenome from a preliminary candidate list based on RRBS. For this purpose, we used bisulphite Pyrosequencing in 94 human and 36 bovine sperm samples. These Pyrosequencing results confirm RRBS as an effective and reliable method to screen for age-DMRs in the vertebrate genome. To decipher whether paternal age effects are an evolutionary conserved feature of mammalian development, we compared methylation patterns between human and bovine sperm in orthologous regulatory regions. We discovered that the level of methylation and the age effect are both species-specific and speculate that these methylation marks reflect the lineage-specific development of each species to hit evolutionary requirements and adaptation processes. Different methylation levels between species in developmentally important genes also imply a differing mutational burden, representing a potential driver for point mutations and consequently deviations in the underlying DNA sequence of different species. Using the example of different haplotypes, this study showed the great effect of single base variations on the methylation of adjacent CpGs. Nonetheless, this study could not provide further evidence or a mechanism for the transfer of epigenetic marks to future generations. Therefore, further research in tissues from the progeny of old and young fathers is required to determine if the observed methylation changes are transmitted to the next generation and if they are associated with altered transcriptional activity of the respective genes. This could provide a direct link between the methylome of sperm from elderly fathers and the development potential of the next generation.
Laut des aktuellen Reports der Weltgesundheitsorganisation sind ca. 466 Millionen Menschen weltweit von einer Hörstörung (HS) betroffen. Durch die enorme Heterogenität und die klinische Variabilität, die diese Erkrankung ausmacht, und viele bisher nicht mit HS assoziierte Gene, bleibt ein großer Teil der erblich bedingten HS in vielen Familien unaufgeklärt. Die Entwicklung moderner Techniken, wie die Next-Generation Sequenzierung (NGS) und der Fortschritt bei der Untersuchung von Modellorganismen trugen jedoch in den letzten Jahren immens dazu bei, neue Gene zu identifizieren, die innerhalb des auditorischen Signalwegs oder damit assoziierten Strukturen beteiligt sind. Die vorliegende Arbeit umfasst Ergebnisse dreier Veröffentlichungen, in denen iranische und pakistanische Familien und eine deutsche Familie mit erblich bedingter HS untersucht und neue, krankheitsverursachende Varianten identifiziert und funktionell charakterisiert wurden. Im ersten Abschnitt konnten zwei neue rezessive Varianten im CDC14A-Gen als krankheitsverursachend identifiziert werden, die zu einem potentiellen Funktionsverlust des kodierten Proteins in einer iranischen und einer pakistanischen Familie führen. Mit Hilfe einer funktionellen Charakterisierung auf RNA-Ebene (Spleiß-Assay und RT-qPCR) konnte der Funktionsverlust beider Varianten bestätigt werden. Der zweite Abschnitt umfasst eine deutsche Familie mit sieben von einer HS betroffenen Familienmitgliedern, in der eine heterozygote missense Variante in MYO3A identifiziert wurde. In der vorliegenden Arbeit konnte somit die erste autosomal dominante Variante in einer europäischen Familie mit einer bilingualen, sensorineuralen Hochtonschwerhörigkeit beschrieben werden und der dominante Charakter von MYO3A bestätigt werden. Im dritten Abschnitt konnten die krankheitsverursachenden Varianten in 13 Familien aus einer Kohorte mit 21 pakistanischen Familien mit einer syndromalen und nicht-syndromalen HS ausfindig gemacht werden. Hierbei wurden sowohl bekannte, als auch bisher nicht beschriebene Varianten detektiert. Die Aufklärungsrate innerhalb dieser Kohorte betrug 61,9% und es konnte somit das Spektrum syndromaler und nicht-syndromaler HS erweitert werden. Der letzte Abschnitt dieser Arbeit beschreibt eine iranische Familie mit einer milden HS und milden Intelligenzminderung, in der eine homozygote missense Variante im Kandidatengen DBN1 ausfindig gemacht wurde. Um die Funktion und die Auswirkungen eines potentiellen Verlusts des codierten Proteins Drebrin zu untersuchen, wurden immunhistochemische Färbungen und auditorische Messungen an Dbn1 Knockout (KO)-Mäusen durchgeführt. Hierbei konnte eine Expression innerhalb der Nervenfasern, die innere Haarzellen innervieren, nachgewiesen werden. Eine leicht verlängerte Latenz für die ABR-Welle IV in KO-Mäusen im Vergleich zum Wildtyp ergab den Hinweis auf einen Defekt innerhalb des zentralen auditorischen Signalwegs, der möglicherweise mit einer Sprachverarbeitungsstörung im Menschen korreliert.
Die Auswirkungen der X-Inaktivierung auf den klinischen Phänotyp bei Patientinnen mit Morbus Fabry
(2023)
M. Fabry ist eine X-chromosomal vererbte Stoffwechselerkrankung. Die Mutation im α-Galactosidase A Gen führt zur reduzierten Aktivität des Enzyms und zur Akkumulation der Stoffwechselprodukte im gesamten Körper. Von der daraus resultierenden Multiorganerkrankung sind sowohl Männer, als auch Frauen betroffen. Als Grund hierfür steht eine verschobene X-Inaktivierung zur Diskussion.
In der vorliegenden Arbeit wurden 104 Frauen rekrutiert und die X-Inaktivierungsmuster in Mundschleimhautepithel, Blut und Hautfibroblasten untersucht. Es wurden umfangreiche klinische und laborchemische Untersuchungen durchgeführt, sodass von jeder Patientin ein klinischer Phänotyp vorlag, der mit Hilfe eines numerischen Scores klassifiziert wurde.
Es zeigte sich, dass Blut ein leicht zu asservierendes Biomaterial mit einer hohen Prävalenz an verschobenen X-Inaktivierungsmustern darstellt. Eine signifikante Korrelation mit dem klinischen Phänotyp konnte in keinem der drei untersuchten Gewebe nachgewiesen werden.
The research that is compiled in this thesis can be divided in two parts. The first part, consisting of four chapters, is centered around the role of epigenetic dysregulation in the etiopathophysiology of sporadic alzheimer's disease (sAD). In addition to providing insights into the most recent developments in neuroepigenomic studies of this disease, the first part of the thesis also touches upon remaining challenges, and provides a future outlook on possible developments in the field. The second part, which includes three more chapters, is focused on the application of induced pluripotent stem cell (iPSC)-based disease models for the study of AD, including but not limited to mechanistic studies on epigenetic dysregulation using this platform. Aside from outlining the research that has been conducted using iPSC-based models for sAD to date, the second part of the thesis also provides insights into the acquisition of disease-relevant neural cultures based on directed differentiation of iPSCs, and furthermore includes an experimental approach for the establishment of such a model system.
Fungal infections are a major global health burden where Candida albicans is among the most common fungal pathogen in humans and is a common cause of invasive candidiasis. Fungal phenotypes, such as those related to morphology, proliferation and virulence are mainly driven by gene expression, which is primarily regulated by kinase signaling cascades. Serine-arginine (SR) protein kinases are highly conserved among eukaryotes and are involved in major transcriptional processes in human and S. cerevisiae. Candida albicans harbors two SR protein kinases, while Sky2 is important for metabolic adaptation, Sky1 has similar functions as in S. cerevisiae. To investigate the role of these SR kinases for the regulation of transcriptional responses in C. albicans, we performed RNA sequencing of sky1Δ and sky2Δ and integrated a comprehensive phosphoproteome dataset of these mutants. Using a Systems Biology approach, we study transcriptional regulation in the context of kinase signaling networks. Transcriptomic enrichment analysis indicates that pathways involved in the regulation of gene expression are downregulated and mitochondrial processes are upregulated in sky1Δ. In sky2Δ, primarily metabolic processes are affected, especially for arginine, and we observed that arginine-induced hyphae formation is impaired in sky2Δ. In addition, our analysis identifies several transcription factors as potential drivers of the transcriptional response. Among these, a core set is shared between both kinase knockouts, but it appears to regulate different subsets of target genes. To elucidate these diverse regulatory patterns, we created network modules by integrating the data of site-specific protein phosphorylation and gene expression with kinase-substrate predictions and protein-protein interactions. These integrated signaling modules reveal shared parts but also highlight specific patterns characteristic for each kinase. Interestingly, the modules contain many proteins involved in fungal morphogenesis and stress response. Accordingly, experimental phenotyping shows a higher resistance to Hygromycin B for sky1Δ. Thus, our study demonstrates that a combination of computational approaches with integration of experimental data can offer a new systems biological perspective on the complex network of signaling and transcription. With that, the investigation of the interface between signaling and transcriptional regulation in C. albicans provides a deeper insight into how cellular mechanisms can shape the phenotype.
Die Fanconi-Anämie (FA) ist eine seltene, heterogene Erbkrankheit. Sie weist ein sehr variables klinisches Erscheinungsbild auf, das sich aus angeborenen Fehlbildungen, hämatologischen Funktionsstörungen, einem erhöhten Risiko für Tumorentwicklung und endokrinen Pathologien zusammensetzt. Die Erkrankung zählt zu den genomischen Instabilitätssyndromen, welche durch eine fehlerhafte DNA-Schadensreparatur gekennzeichnet sind. Bei der FA zeigt sich dies vor allem in einer charakteristischen Hypersensitivität gegenüber DNA-quervernetzenden Substanzen (z. B. Mitomycin C, Cisplatin). Der zelluläre FA-Phänotyp zeichnet sich durch eine erhöhte Chromosomenbrüchigkeit und einen Zellzyklusarrest in der G2-Phase aus. Diese Charakteristika sind bereits spontan vorhanden und werden durch Induktion mit DNA-quervernetzenden Substanzen verstärkt. Der Gendefekt ist dabei in einem der 22 bekannten FA-Gene (FANCA, -B, -C, -D1, -D2, -E, -F, -G, -I, -J, -L, -M, -N, -O, -P, -Q, -R, -S, -T, -U, -V, -W) oder in noch unbekannten FA-Genen zu finden. Die FA-Gendefekte werden mit Ausnahme von FANCR (dominant-negative de novo Mutationen) und FANCB (X-chromosomal) autosomal rezessiv vererbt. Die FA-Genprodukte bilden zusammen mit weiteren Proteinen den FA/BRCA-Signalweg. Das Schlüsselereignis dieses Signalwegs stellt die Monoubiquitinierung von FANCD2 und FANCI (ID2-Komplex) dar. Ausgehend davon lässt sich zwischen upstream- und downstream-gelegenen FA-Proteinen unterscheiden. Letztere sind direkt an der DNA-Schadensreparatur beteiligt. Zu den upstream-gelegenen Proteinen zählt der FA-Kernkomplex, der sich aus bekannten FA-Proteinen und aus FA-assoziierten-Proteinen (FAAPs) zusammensetzt und für die Monoubiquitinierung des ID2-Komplexes verantwortlich ist. Für FAAPs wurden bisher keine pathogenen humanen Mutationen beschrieben. Zu diesen Proteinen gehört auch FAAP100, das mit FANCB und FANCL innerhalb des FA-Kernkomplexes den Subkomplex LBP100 bildet.
Durch die vorliegende Arbeit wurde eine nähere Charakterisierung dieses Proteins erreicht. In einer Amnion-Zelllinie konnte eine homozygote Missense-Mutation identifiziert werden. Der Fetus zeigte einen typischen FA-Phänotyp und auch seine Zellen wiesen charakteristische FA-Merkmale auf. Der zelluläre Phänotyp ließ sich durch FAAP100WT komplementieren, sodass die Pathogenität der Mutation bewiesen war. Unterstützend dazu wurden mithilfe des CRISPR/Cas9-Systems weitere FAAP100-defiziente Zelllinien generiert. Diese zeigten ebenfalls einen typischen FA-Phänotyp, welcher sich durch FAAP100WT komplementieren ließ. Die in vitro-Modelle dienten als Grundlage dafür, die Funktion des FA-Kernkomplexes im Allgemeinen und die des Subkomplexes LBP100 im Besonderen besser zu verstehen. Dabei kann nur durch intaktes FAAP100 das LBP100-Modul gebildet und dieses an die DNA-Schadensstelle transportiert werden. Dort leistet FAAP100 einen essentiellen Beitrag für den FANCD2-Monoubiquitinierungsprozess und somit für die Aktivierung der FA-abhängigen DNA-Schadensreparatur. Um die Funktion von FAAP100 auch in vivo zu untersuchen, wurde ein Faap100-/--Mausmodell generiert, das einen mit anderen FA-Mausmodellen vergleichbaren, relativ schweren FA-Phänotyp aufwies. Aufgrund der Ergebnisse lässt sich FAAP100 als neues FA-Gen klassifizieren. Zudem wurde die Rolle des Subkomplexes LBP100 innerhalb des FA-Kernkomplexes weiter aufgeklärt. Beides trägt zu einem besseren Verständnis des FA/BRCA-Signalweges bei. Ein weiterer Teil der vorliegenden Arbeit beschäftigt sich mit der Charakterisierung von FAAP100138, einer bisher nicht validierten Isoform von FAAP100. Durch dieses Protein konnte der zelluläre FA-Phänotyp von FAAP100-defizienten Zelllinien nicht komplementiert werden, jedoch wurden Hinweise auf einen dominant-negativen Effekt von FAAP100138 auf den FA/BRCA-Signalweg gefunden. Dies könnte zu der Erklärung beitragen, warum und wie der Signalweg, beispielsweise in bestimmtem Gewebearten, herunterreguliert wird. Zudem wäre eine Verwendung in der Krebstherapie denkbar.
Usher syndrome, the most prevalent cause of combined hereditary vision and hearing impairment, is clinically and genetically heterogeneous. Moreover, several conditions with phenotypes overlapping Usher syndrome have been described. This makes the molecular diagnosis of hereditary deaf-blindness challenging. Here, we performed exome sequencing and analysis on 7 Mexican and 52 Iranian probands with combined retinal degeneration and hearing impairment (without intellectual disability). Clinical assessment involved ophthalmological examination and hearing loss questionnaire. Usher syndrome, most frequently due to biallelic variants in MYO7A (USH1B in 16 probands), USH2A (17 probands), and ADGRV1 (USH2C in 7 probands), was diagnosed in 44 of 59 (75%) unrelated probands. Almost half of the identified variants were novel. Nine of 59 (15%) probands displayed other genetic entities with dual sensory impairment, including Alström syndrome (3 patients), cone-rod dystrophy and hearing loss 1 (2 probands), and Heimler syndrome (1 patient). Unexpected findings included one proband each with Scheie syndrome, coenzyme Q10 deficiency, and pseudoxanthoma elasticum. In four probands, including three Usher cases, dual sensory impairment was either modified/aggravated or caused by variants in distinct genes associated with retinal degeneration and/or hearing loss. The overall diagnostic yield of whole exome analysis in our deaf-blind cohort was 92%. Two (3%) probands were partially solved and only 3 (5%) remained without any molecular diagnosis. In many cases, the molecular diagnosis is important to guide genetic counseling, to support prognostic outcomes and decisions with currently available and evolving treatment modalities.
Aufgrund mangelnder Aktivität der Gewebe-unspezifischen Phosphatase (tissue-nonspecific alkaline phosphatase, TNAP) kommt es zum Krankheitsbild der Hypophosphatasie (HPP). Neben skelettalen und neuronalen Symptomen leiden Patienten mit HPP häufig an einem vorzeitigen Verlust der Milchzähne und weiteren dentalen Manifestationen, wie Zahnhartsubstanzdefekten, Eruptionsstörungen, erweiterte Pulpenkammern oder einer verringerten alveolären Knochenhöhe.
Ziel der Arbeit war es, den Einfluss der TNAP auf die Zahnentwicklung von Zebrafischlarven zu untersuchen, um ein neues in-vivo Modell für die dentalen Auswirkungen bei Hypophosphatasie etablieren zu können. Um die sehr kleinen Zähne der Zebrafischlarven auch in frühen Entwicklungsstadien darzustellen, wurden mittels verschiedener histologischer Färbungen die Zahnstrukturen angefärbt und die Larven danach in JB4®, einen polymeren Kunststoff, eingebettet. Im Anschluss wurden histologische Schnitte angefertigt und am Fluoreszenzmikroskop ausgewertet.
Einerseits konnte durch In-situ-Hybridisierung die Expression verschiedener Gene, wie z.B. alpl (welches für die Tnap im Zebrafisch kodiert), im Bereich von dentalen Strukturen in verschiedenen Entwicklungsstadien nachgewiesen werden. Außerdem zeigte die Analyse der dentalen Strukturen nach Inhibition der Tnap mittels Levamisol bei fünf Tage alten Zebrafischlarven eine Veränderung von Form, Größe und Struktur der ersten Zähne. Die TNAP-Inhibition führte auch zur quantitativ nachweisbaren Steigerung des Fluoreszenzsignals von ß-Catenin, welches eine zentrale Funktion im Wnt/ß-Catenin-Signalweg besitzt und essenziell in verschiedenen zellulären Prozessen während der Embryogenese ist.
Zusammenfassend zeigen die Ergebnisse der Arbeit, dass der Zebrafisch großes Potenzial als in-vivo Modell für die dentalen Symptome bei HPP bietet. Außerdem eröffnen sich neue interessante Fragen in Bezug auf den Einfluss von ß-Catenin bei den frühen pathophysiologischen Prozessen der Erkrankung.
Background
Mucopolysaccharidosis type III (Sanfilippo syndrome) is a lysosomal storage disorder, caused by a deficiency in the heparan-N-sulfatase enzyme involved in the catabolism of the glycosaminoglycan heparan sulfate. It is characterized by early nonspecific neuropsychiatric symptoms, followed by progressive neurocognitive impairment in combination with only mild somatic features. In this patient group with a broad clinical spectrum a significant genotype-phenotype correlation with some mutations leading to a slower progressive, attenuated course has been demonstrated.
Case presentation
Our patient had complications in the neonatal period and was diagnosed with Mucopolysaccharidosis IIIa only at the age of 28 years. He was compound heterozygous for the variants p.R245H and p.S298P, the latter having been shown to lead to a significantly milder phenotype.
Conclusions
The diagnostic delay is even more prolonged in this patient population with comorbidities and a slowly progressive course of the disease.
Male breast cancer (mBC) is associated with a high prevalence of pathogenic variants (PVs) in the BRCA2 gene; however, data regarding other BC predisposition genes are limited. In this retrospective multicenter study, we investigated the prevalence of PVs in BRCA1/2 and 23 non-BRCA1/2 genes using a sample of 614 patients with mBC, recruited through the centers of the German Consortium for Hereditary Breast and Ovarian Cancer. A high proportion of patients with mBC carried PVs in BRCA2 (23.0%, 142/614) and BRCA1 (4.6%, 28/614). The prevalence of BRCA1/2 PVs was 11.0% in patients with mBC without a family history of breast and/or ovarian cancer. Patients with BRCA1/2 PVs did not show an earlier disease onset than those without. The predominant clinical presentation of tumor phenotypes was estrogen receptor (ER)-positive, progesterone receptor (PR)-positive, and HER2-negative (77.7%); further, 10.2% of the tumors were triple-positive, and 1.2% were triple-negative. No association was found between ER/PR/HER2 status and BRCA1/2 PV occurrence. Comparing the prevalence of protein-truncating variants (PTVs) between patients with mBC and control data (ExAC, n = 27,173) revealed significant associations of PTVs in both BRCA1 and BRCA2 with mBC (BRCA1: OR = 17.04, 95% CI = 10.54–26.82, p < 10\(^{−5}\); BRCA2: OR = 77.71, 95% CI = 58.71–102.33, p < 10\(^{−5}\)). A case-control investigation of 23 non-BRCA1/2 genes in 340 BRCA1/2-negative patients and ExAC controls revealed significant associations of PTVs in CHEK2, PALB2, and ATM with mBC (CHEK2: OR = 3.78, 95% CI = 1.59–7.71, p = 0.002; PALB2: OR = 14.77, 95% CI = 5.02–36.02, p < 10\(^{−5}\); ATM: OR = 3.36, 95% CI = 0.89–8.96, p = 0.04). Overall, our findings support the benefit of multi-gene panel testing in patients with mBC irrespective of their family history, age at disease onset, and tumor phenotype.
In der vorliegenden Arbeit wurden mittels 5-Methylcytosin Immunofärbung zytogenetische Analysen an Metaphasechromosomen aus der Mitose, an Interphase-Zellen verschiedener Organe und an Meiose-Stadien der Maus (Mus musculus) zur Detektion hypermethylierter DNA durchgeführt. Zusätzlich erfolgte eine C-Bänderung an Metaphasechromosomen und Meiose-Stadien zum Nachweis von konstitutivem Heterochromatin.
Background
Dystrophinopathies caused by variants in the DMD gene are a well‐studied muscle disease. The most common type of variant in DMD are large deletions. Very rarely reported forms of variants are chromosomal translocations, inversions and deep intronic variants (DIVs) because they are not detectable by standard diagnostic techniques (sequencing of coding sequence, copy number variant detection). This might be the reason that some clinically and histologically proven dystrophinopathy cases remain unsolved.
Methods
We used whole genome sequencing (WGS) to screen the entire DMD gene for variants in one of two brothers suffering from typical muscular dystrophy with strongly elevated creatine kinase levels.
Results
Although a pathogenic DIV could not be detected, we were able to identify a pericentric inversion with breakpoints in DMD intron 44 and Xq13.3, which could be confirmed by Sanger sequencing in the index as well as in his brother and mother. As this variation affects a major part of DMD it is most likely disease causing.
Conclusion
Our findings elucidate that WGS is capable of detecting large structural rearrangements and might be suitable for the genetic diagnostics of dystrophinopathies in the future. In particular, inversions might be a more frequent cause for dystrophinopathies as anticipated and should be considered in genetically unsolved dystrophinopathy cases.
Immortalized hepatic stellate cells (HSCs) established from mouse, rat, and humans are valuable in vitro models for the biomedical investigation of liver biology. These cell lines are homogenous, thereby providing consistent and reproducible results. They grow more robustly than primary HSCs and provide an unlimited supply of proteins or nucleic acids for biochemical studies. Moreover, they can overcome ethical concerns associated with the use of animal and human tissue and allow for fostering of the 3R principle of replacement, reduction, and refinement proposed in 1959 by William M. S. Russell and Rex L. Burch. Nevertheless, working with continuous cell lines also has some disadvantages. In particular, there are ample examples in which genetic drift and cell misidentification has led to invalid data. Therefore, many journals and granting agencies now recommend proper cell line authentication. We herein describe the genetic characterization of the rat HSC line HSC-T6, which was introduced as a new in vitro model for the study of retinoid metabolism. The consensus chromosome markers, outlined primarily through multicolor spectral karyotyping (SKY), demonstrate that apart from the large derivative chromosome 1 (RNO1), at least two additional chromosomes (RNO4 and RNO7) are found to be in three copies in all metaphases. Additionally, we have defined a short tandem repeat (STR) profile for HSC-T6, including 31 species-specific markers. The typical features of these cells have been further determined by electron microscopy, Western blotting, and Rhodamine-Phalloidin staining. Finally, we have analyzed the transcriptome of HSC-T6 cells by mRNA sequencing (mRNA-Seq) using next generation sequencing (NGS).
Arrhythmogenic cardiomyopathy (ACM) is an inherited heart muscle disease caused by heterozygous missense mutations within the gene encoding for the nuclear envelope protein transmembrane protein 43 (TMEM43). The disease is characterized by myocyte loss and fibro-fatty replacement, leading to life-threatening ventricular arrhythmias and sudden cardiac death. However, the role of TMEM43 in the pathogenesis of ACM remains poorly understood. In this study, we generated cardiomyocyte-restricted transgenic zebrafish lines that overexpress eGFP-linked full-length human wild-type (WT) TMEM43 and two genetic variants (c.1073C>T, p.S358L; c.332C>T, p.P111L) using the Tol2-system. Overexpression of WT and p.P111L-mutant TMEM43 was associated with transcriptional activation of the mTOR pathway and ribosome biogenesis, and resulted in enlarged hearts with cardiomyocyte hypertrophy. Intriguingly, mutant p.S358L TMEM43 was found to be unstable and partially redistributed into the cytoplasm in embryonic and adult hearts. Moreover, both TMEM43 variants displayed cardiac morphological defects at juvenile stages and ultrastructural changes within the myocardium, accompanied by dysregulated gene expression profiles in adulthood. Finally, CRISPR/Cas9 mutants demonstrated an age-dependent cardiac phenotype characterized by heart enlargement in adulthood. In conclusion, our findings suggest ultrastructural remodeling and transcriptomic alterations underlying the development of structural and functional cardiac defects in TMEM43-associated cardiomyopathy.
Rare variants in at least 10 genes, including BRCA1, BRCA2, PALB2, ATM, and CHEK2, are associated with increased risk of breast cancer; however, these variants, in combination with common variants identified through genome-wide association studies, explain only a fraction of the familial aggregation of the disease. To identify further susceptibility genes, we performed a two-stage whole-exome sequencing study. In the discovery stage, samples from 1528 breast cancer cases enriched for breast cancer susceptibility and 3733 geographically matched unaffected controls were sequenced. Using five different filtering and gene prioritization strategies, 198 genes were selected for further validation. These genes, and a panel of 32 known or suspected breast cancer susceptibility genes, were assessed in a validation set of 6211 cases and 6019 controls for their association with risk of breast cancer overall, and by estrogen receptor (ER) disease subtypes, using gene burden tests applied to loss-of-function and rare missense variants. Twenty genes showed nominal evidence of association (p-value < 0.05) with either overall or subtype-specific breast cancer. Our study had the statistical power to detect susceptibility genes with effect sizes similar to ATM, CHEK2, and PALB2, however, it was underpowered to identify genes in which susceptibility variants are rarer or confer smaller effect sizes. Larger sample sizes would be required in order to identify such genes.
New techniques in molecular genetic diagnostics now allow for accurate diagnosis in a large proportion of patients with muscular diseases. Nevertheless, many patients remain unsolved, although the clinical history and/or the muscle biopsy give a clear indication of the involved genes. In many cases, there is a strong suspicion that the cause must lie in unexplored gene areas, such as deep-intronic or other non-coding regions. In order to find these changes, next-generation sequencing (NGS) methods are constantly evolving, making it possible to sequence entire genomes to reveal these previously uninvestigated regions. Here, we present a young woman who was strongly suspected of having a so far genetically unsolved sarcoglycanopathy based on her clinical history and muscle biopsy. Using short read whole genome sequencing (WGS), a homozygous inversion on chromosome 13 involving SGCG and LINC00621 was detected. The breakpoint in intron 2 of SGCG led to the absence of γ-sarcoglycan, resulting in the manifestation of autosomal recessive limb-girdle muscular dystrophy 5 (LGMDR5) in the young woman.
Hepatic stellate cells (HSCs) are also known as lipocytes, fat-storing cells, perisinusoidal cells, or Ito cells. These liver-specific mesenchymal cells represent about 5% to 8% of all liver cells, playing a key role in maintaining the microenvironment of the hepatic sinusoid. Upon chronic liver injury or in primary culture, these cells become activated and transdifferentiate into a contractile phenotype, i.e., the myofibroblast, capable of producing and secreting large quantities of extracellular matrix compounds. Based on their central role in the initiation and progression of chronic liver diseases, cultured HSCs are valuable in vitro tools to study molecular and cellular aspects of liver diseases. However, the isolation of these cells requires special equipment, trained personnel, and in some cases needs approval from respective authorities. To overcome these limitations, several immortalized HSC lines were established. One of these cell lines is CFSC, which was originally established from cirrhotic rat livers induced by carbon tetrachloride. First introduced in 1991, this cell line and derivatives thereof (i.e., CFSC-2G, CFSC-3H, CFSC-5H, and CFSC-8B) are now used in many laboratories as an established in vitro HSC model. We here describe molecular features that are suitable for cell authentication. Importantly, chromosome banding and multicolor spectral karyotyping (SKY) analysis demonstrate that the CFSC-2G genome has accumulated extensive chromosome rearrangements and most chromosomes exist in multiple copies producing a pseudo-triploid karyotype. Furthermore, our study documents a defined short tandem repeat (STR) profile including 31 species-specific markers, and a list of genes expressed in CFSC-2G established by bulk mRNA next-generation sequencing (NGS).
The conspicuous colour sexual dimorphism of guppies has made them paradigmatic study objects for sex-linked traits and sex chromosome evolution. Both the X- and Y-chromosomes of the common guppy (Poecilia reticulata) are genetically active and homomorphic, with a large homologous part and a small sex specific region. This feature is considered to emulate the initial stage of sex chromosome evolution. A similar situation has been documented in the related Endler’s and Oropuche guppies (P. wingei, P. obscura) indicating a common origin of the Y in this group. A recent molecular study in the swamp guppy (Micropoecilia. picta) reported a low SNP density on the Y, indicating Y-chromosome deterioration. We performed a series of cytological studies on M. picta to show that the Y-chromosome is quite small compared to the X and has accumulated a high content of heterochromatin. Furthermore, the Y-chromosome stands out in displaying CpG clusters around the centromeric region. These cytological findings evidently illustrate that the Y-chromosome in M. picta is indeed highly degenerated. Immunostaining for SYCP3 and MLH1 in pachytene meiocytes revealed that a substantial part of the Y remains associated with the X. A specific MLH1 hotspot site was persistently marked at the distal end of the associated XY structure. These results unveil a landmark of a recombining pseudoautosomal region on the otherwise strongly degenerated Y chromosome of M. picta. Hormone treatments of females revealed that, unexpectedly, no sexually antagonistic color gene is Y-linked in M. picta. All these differences to the Poecilia group of guppies indicate that the trajectories associated with the evolution of sex chromosomes are not in parallel.
A growing number of sperm methylome analyses have identified genomic loci that are susceptible to paternal age effects in a variety of mammalian species, including human, bovine, and mouse. However, there is little overlap between different data sets. Here, we studied whether or not paternal age effects on the sperm epigenome have been conserved in mammalian evolution and compared methylation patterns of orthologous regulatory regions (mainly gene promoters) containing both conserved and non-conserved CpG sites in 94 human, 36 bovine, and 94 mouse sperm samples, using bisulfite pyrosequencing. We discovered three (NFKB2, RASGEF1C, and RPL6) age-related differentially methylated regions (ageDMRs) in humans, four (CHD7, HDAC11, PAK1, and PTK2B) in bovines, and three (Def6, Nrxn2, and Tbx19) in mice. Remarkably, the identified sperm ageDMRs were all species-specific. Most ageDMRs were in genomic regions with medium methylation levels and large methylation variation. Orthologous regions in species not showing this age effect were either hypermethylated (>80%) or hypomethylated (<20%). In humans and mice, ageDMRs lost methylation, whereas bovine ageDMRs gained methylation with age. Our results are in line with the hypothesis that sperm ageDMRs are in regions under epigenomic evolution and may be part of an epigenetic mechanism(s) for lineage-specific environmental adaptations and provide a solid basis for studies on downstream effects in the genes analyzed here.
Background
Although Fabry disease (FD) is an X-linked lysosomal storage disorder caused by mutations in the α-galactosidase A gene (GLA), women may develop severe symptoms. We investigated X-chromosomal inactivation patterns (XCI) as a potential determinant of symptom severity in FD women.
Patients and Methods
We included 95 women with mutations in GLA (n = 18 with variants of unknown pathogenicity) and 50 related men, and collected mouth epithelial cells, venous blood, and skin fibroblasts for XCI analysis using the methylation status of the androgen receptor gene. The mutated X-chromosome was identified by comparison of samples from relatives. Patients underwent genotype categorization and deep clinical phenotyping of symptom severity.
Results
43/95 (45%) women carried mutations categorized as classic. The XCI pattern was skewed (i.e., ≥75:25% distribution) in 6/87 (7%) mouth epithelial cell samples, 31/88 (35%) blood samples, and 9/27 (33%) skin fibroblast samples. Clinical phenotype, α-galactosidase A (GAL) activity, and lyso-Gb3 levels did not show intergroup differences when stratified for X-chromosomal skewing and activity status of the mutated X-chromosome.
Conclusions
X-inactivation patterns alone do not reliably reflect the clinical phenotype of women with FD when investigated in biomaterial not directly affected by FD. However, while XCI patterns may vary between tissues, blood frequently shows skewing of XCI patterns.
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.
Hereditäre Kardiomyopathien sind durch klinische und genetische Heterogenität gekennzeichnet, welche die Kardiogenetik vor Herausforderungen stellt. In dieser Arbeit wurden manche dieser Herausforderungen angegangen, indem anhand einer Kohorte von 61 Patienten mit Kardiomyopathie bzw. primärer Arrhythmie eine Exom-Diagnostik mit anschließender stufenweiser Datenanalyse vorgenommen wurde.
Ein Ziel der Arbeit war, die aktuellen diagnostischen Detektionsraten zu prüfen sowie zu bewerten, ob eine erweiterte Exom-Diagnostik im Vergleich zur üblichen Genpanel-Analyse einen diagnostischen Zugewinn bringt. Zudem sollten potenzielle Krankheitsgene sowie komplexe Genotypen identifiziert werden.
Die Ergebnisse zeigten, dass bei insgesamt 64% der Patienten eine Variante von Interesse gefunden wurde. Hervorzuheben ist die hohe Detektionsrate in der größten Subkohorte, die aus Patienten mit dilatativer bzw. linksventrikulärer Non-Compaction Kardiomyopathie bestand: 69% und damit höher im Vergleich zur in der Literatur berichteten Detektionsrate von bis zu 50%.
Im Rahmen der stufenweisen Daten-Auswertung zeigte sich zwar, dass die meisten kausalen Varianten in den phänotypspezifischen Panels zu finden waren, die Analyse eines erweiterten Panels mit 79 Genen sowie der Gesamtexom-Daten aber zu einer zusätzlichen Aufklärungsquote von 13% bzw. 5% führte. Durch die Erweiterung der Diagnostik konnten interessante, teilweise neue Assoziationen zwischen Genotyp und Phänotyp sowie neue Kandidatengene identifiziert werden. Das beste Beispiel dafür ist eine trunkierende Variante im STK38-Gen, das an der Phosphorylierung eines Regulators der Expression kardialer Gene beteiligt ist.
Zusammenfassend konnte gezeigt werden, dass, obwohl die Detektionsrate von Genpanels für die Routine-Diagnostik akzeptabel ist, die Anwendung von Exom-Diagnostik einen diagnostischen Zugewinn, die Entdeckung von interessanten Genotyp-Phänotyp-Korrelationen sowie die Identifizierung von Kandidatengenen ermöglicht.
Background
Patients with coronavirus disease 2019 (COVID-19) who undergo surgery have impaired postoperative outcomes and increased mortality. Consequently, elective and semi-urgent operations on the increasing number of patients severely affected by COVID-19 have been indefinitely postponed.in many countries with unclear implications on disease progression and overall survival. The purpose of this study was to evaluate whether the establishment of a standardized screening program for acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is sufficient to ensure high-quality medical and surgical treatment of COVID-19 and non-COVID-19 patients while minimizing in-hospital SARS-CoV-2 transmission.
Methods
The screening program comprised polymerase chain reaction (PCR) testing of nasopharyngeal swabs and a standardized questionnaire about potential symptoms for SARS-CoV-2 infection. All elective and emergency patients admitted to the surgical department of a tertiary-care hospital center in Lower Franconia, Germany, between March and May 2020 were included and their characteristics were recorded.
Results
Out of the study population (n = 657), 509 patients (77.5%) had at least one risk factor for a potentially severe course of COVID-19 and 164 patients (25%) were active smokers. The average 7-day incidence in Lower Franconia was 24.0/100,000 during the observation period. Preoperative PCR testing revealed four asymptomatic positive patients out of the 657 tested patients. No postoperative SARS-CoV-2 infection or transmission could be detected.
Conclusion
The implementation of a standardized preoperative screening program to both COVID-19 and non-COVID-19 patients can ensure high-quality surgical care while minimizing infection risk for healthcare workers and potential in-hospital transmission.
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.
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.
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.
Copy number variants of SLC2A3, which encodes the glucose transporter GLUT3, are associated with several neuropsychiatric and cardiac diseases. Here, we report the successful reprogramming of peripheral blood mononuclear cells from two SLC2A3 duplication and two SLC2A3 deletion carriers and subsequent generation of two transgene-free iPSC clones per donor by Sendai viral transduction. All eight clones represent bona fide hiPSCs with high expression of pluripotency genes, ability to differentiate into cells of all three germ layers and normal karyotype. The generated cell lines will be helpful to enlighten the role of glucometabolic alterations in pathophysiological processes shared across organ boundaries.
Prerequisite to any biological laboratory assay employing living animals is consideration about its necessity, feasibility, ethics and the potential harm caused during an experiment. The imperative of these thoughts has led to the formulation of the 3R-principle, which today is a pivotal scientific standard of animal experimentation worldwide. The rising amount of laboratory investigations utilizing living animals throughout the last decades, either for regulatory concerns or for basic science, demands the development of alternative methods in accordance with 3R to help reduce experiments in mammals. This demand has resulted in investigation of additional vertebrate species displaying favourable biological properties. One prominent species among these is the zebrafish (Danio rerio), as these small laboratory ray-finned fish are well established in science today and feature outstanding biological characteristics. In this review, we highlight the advantages and general prerequisites of zebrafish embryos and larvae before free-feeding stages for toxicological testing, with a particular focus on cardio-, neuro, hepato- and nephrotoxicity. Furthermore, we discuss toxicokinetics, current advances in utilizing zebrafish for organ toxicity testing and highlight how advanced laboratory methods (such as automation, advanced imaging and genetic techniques) can refine future toxicological studies in this species.
Deafness, the most frequent sensory deficit in humans, is extremely heterogeneous with hundreds of genes involved. Clinical and genetic analyses of an extended consanguineous family with pre-lingual, moderate-to-profound autosomal recessive sensorineural hearing loss, allowed us to identify CLRN2, encoding a tetraspan protein, as a new deafness gene. Homozygosity mapping followed by exome sequencing identified a 14.96 Mb locus on chromosome 4p15.32p15.1 containing a likely pathogenic missense variant in CLRN2 (c.494C > A, NM_001079827.2) segregating with the disease. Using in vitro RNA splicing analysis, we show that the CLRN2 c.494C > A variant leads to two events: (1) the substitution of a highly conserved threonine (uncharged amino acid) to lysine (charged amino acid) at position 165, p.(Thr165Lys), and (2) aberrant splicing, with the retention of intron 2 resulting in a stop codon after 26 additional amino acids, p.(Gly146Lysfs*26). Expression studies and phenotyping of newly produced zebrafish and mouse models deficient for clarin 2 further confirm that clarin 2, expressed in the inner ear hair cells, is essential for normal organization and maintenance of the auditory hair bundles, and for hearing function. Together, our findings identify CLRN2 as a new deafness gene, which will impact future diagnosis and treatment for deaf patients.
Trotz der rasanten Entwicklung molekulargenetischer Analysemethoden sind die Auslöser vieler Erbrankheiten bislang ungeklärt. Eine Identifikation der genetischen Ursache einer Erkrankung ist jedoch essenziell, um zusätzliche invasive Tests vermeiden, adäquate Therapiemaßnahmen in die Wege leiten, akkurate Prognosen stellen und eine entsprechende genetische Beratung anbieten zu können. Next Generation Sequencing (NGS)-basierte Techniken wie die Whole Exome Sequenzierung (WES) haben die humangenetische Forschung und Diagnostik in den letzten Jahren revolutioniert. Die WES ermöglicht die Sequenzierung der Exons aller proteincodierenden Gene von mehreren Individuen gleichzeitig und stellt ein hilfreiches Werkzeug bei der Suche nach neuen kranheitsrelevanten Genen im Menschen dar.
Die vorliegende Arbeit beschäftigt sich mit der Aufklärung genetischer Ursachen verschiedenster Erkrankungen in konsanguinen Familien aus dem nahen und mittleren Osten mittels WES. Insgesamt wurden 43 Patienten mit unterschiedlichen Krankheitsbildern untersucht, darunter viele mit Skelettdysplasien oder Neuropathien. In 22 Fällen (51%) konnte die entsprechende krankheitsverursachende Mutation ausfindig gemacht werden. In 21% der aufgeklärten Fälle wurden Sequenzvarianten detektiert, die in der Literatur bereits als pathogen beschrieben wurden, während 63% bisher noch unbekannte Mutationen in bereits als krankheitsrelevant beschriebenen Genen darstellten. Zudem konnten im Rahmen dieser Arbeit drei neue, für den Menschen krankheitsrelevante Gene identifiziert werden, solute carrier family 10 member 7 (SLC10A7), T-box 4 (TBX4) und MIA SH3 domain ER export factor 3 (MIA3). SLC10A7 codiert für einen Transporter aus der Familie der solute carrier, der in der Plasmamembran verankert ist. In dieser Arbeit geleistete Analyseergebnisse konnten zu der Erstbeschreibung von homozygoten pathogenen SLC10A7-Mutationen als Ursache für eine Skelettdysplasie mit Amelogenesis imperfecta beitragen. Bei TBX4 handelt es sich um einen hochkonservierten Transkriptionsfaktor, der während der embryonalen Entwicklung an der Ausbildung der unteren Extremitäten beteiligt ist. Homozygote pathogene TBX4-Mutationen wurden im Kontext dieser Arbeit erstmalig mit einer posterioren Amelie mit Becken- und Lungenhypoplasie in Verbindung gebracht. MIA3 ist ein Transmembranprotein des endoplasmatischen Retikulums, das eine essenzielle Rolle bei der Proteinsekretion spielt. Die hier vorgestellten Patienten mit homozygoten pathogenen MIA3-Mutationen zeigen eine komplexe syndromale Erkrankung, die sich hauptsächlich in einer Kollagenopathie, Diabetes mellitus und milder mentaler Retardierung manifestiert und ein neues Krankheitsbild darstellt.
Die im Rahmen dieser Arbeit erzielten Ergebnisse erweitern somit zum einen das Mutationsspektrum verschiedener bekannter Krankheitsbilder und offenbaren zum anderen neue krankheitsrelevante Gene im Menschen.
The effect of late parenthood on the offspring´s physical and mental health status has recently become an increasingly important topic of discussion. Studies on neurodevelopmental disorders in children of older parents (Naserbakht et al., 2011) outline the negative consequences of aging fathers as unpredictable compared to the better-understood unfavorable maternal influences (Cedars et al. 2015). This may be due to the fact that lifelong production of male gametes becomes more susceptible to error, not only for somatic mutations. Non-genomic mechanisms such as epigenetic methylation also alter DNA dynamically throughout life (Jones et al., 2015) and influence the aging human sperm DNA (Jenkins et al., 2014). These methylation changes may be transmitted to the next generation via epigenetic inheritance mechanisms (Milekic et al., 2015), which may negatively impact the sensitive epigenetic regulation of cell differentiation in the embryonic period (Curley et al., 2011; Spiers et al., 2015). Accordingly, Nardone et al. (2014) reported several hypomethylated regions in autistic patients, illustrating potential epigenetic influences on the multifactorial pathogenesis of neuropsychiatric disorders. In the present study, the methylation status of five gene regions in the sperm DNA of males of different ages was analyzed by two techniques - pyrosequencing and deep bisulfite sequencing. Two gene regions, FOXK1 and DMPK, showed a highly significant age-related methylation loss and FOXK1 a reduced methylation variation at the level of single alleles. In addition, the examined gene region of FOXK1 showed significant methylation changes in the fetal cord blood DNA of the respective offspring of the sperm donor. This fact suggests a transfer of age-related methylation loss to the next generation. Interestingly, a methylation analysis at the level of single alleles showed that the methylation loss was inherited exclusively by the father. FOXK1 is a transcription factor that plays an important role in the epigenetic regulation of the cell cycle during embryonic neuronal development (Huang et al., 2004; Wijchers et al., 2006). For this reason, the methylation status of FOXK1 in the blood of autistic patients and an age- and sex-matched control group was investigated. While both groups showed age-associated FOXK1 methylation loss, a faster dynamics of methylation change was observed in the autistic group. Although further studies are needed to uncover inheritance mechanisms of epigenetic information, the present results show an evident influence of age-related methylation changes on offspring. When advising future fathers, it is important to consider how the paternal epigenome is altered by aging and can have a negative impact on the developing embryo.
Fanconi-Anämie (FA) ist, mit Ausnahme von Mutationen in FANCR/RAD51, eine autosomal-rezessive oder X-chromosomal vererbte Krankheit, die sich durch eine ausgesprochene klinische als auch genetische Heterogenität auszeichnet. Neben einem fortschreitenden Knochenmarksversagen zählen zu den typischen Merkmalen eine Vielzahl an angeborenen Fehlbildungen, wie beispielsweise Radialstrahlanomalien, Minderwuchs oder Pigmentierungsstörungen. Zudem besteht für FA-Patienten ein überdurchschnittlich hohes Risiko bereits in jungen Jahren an akuter myeloischer Leukämie oder soliden Tumoren zu erkranken. Bislang konnten in 21 FA-Genen (FANCA, -B, -C, - D1, -D2, -E, -F, -G, -I, -J, -L, -M, -N, -O, -P, -Q, -R, -S, -T, -U oder -V) krankheitsverursachende Mutationen identifiziert werden, deren Proteinprodukte maßgeblich an der Aufrechterhaltung der Genomstabilität beteiligt sind und Komponenten des FA/BRCA-DNA-Reparaturweges darstellen. In der klassischen FA-Mutationsanalyse kommen meist Sanger-Sequenzierungen sowie MLPA- und Immunblot-Analysen zum Einsatz. Da im Wesentlichen keine Genotyp-Phänotyp-Korrelation besteht, gestaltet sich, gerade bei seltenen FA-Komplementationsgruppen, der Nachweis von krankheitsverursachenden Mutationen oftmals sehr zeit- und kostenintensiv. Während der letzten Jahre wurden verschiedene Strategien zur Anreicherung und Sequenzierung entwickelt, welche die parallele Sequenzanalyse einzelner ausgewählter Gene, ganzer Exome oder sogar des gesamten Genoms und somit eine kosten- und zeiteffiziente Mutationsanalyse ermöglichen. In der vorliegenden Arbeit wurden unterschiedliche Anreicherungsmethoden mit anschließender Hochdurchsatzsequenzierung auf ihre Anwendbarkeit in der molekulargenetischen FA-Diagnostik getestet, um klassische Mutationsanalyse-Methoden zu ergänzen oder möglicherweise sogar ganz ersetzen zu können.
Der erste Teil der Arbeit befasste sich mit der Etablierung eines FA-spezifischen Genpanels zur Genotypisierung von FA-Patienten. Nachdem die Methode zunächst anhand von FA-Patienten mit bekannten Mutationen optimiert werden musste, erwies sie sich als effizienter Ansatz zum Nachweis krankheitsverursachender Mutationen bei FA-Patienten unbekannter Komplementationsgruppe. Durch die FA-Panelanalyse konnten 37 von 47 unklassifizierten Patienten einer FA-Komplementationsgruppe zugeordnet werden, indem deren kausalen Mutationen bestimmt wurden. In einem weiteren Ansatz sollte die Anwendbarkeit eines kommerziellen Anreicherungspanels zur FA-Diagnostik untersucht werden. Auch hier konnte ein Großteil der krankheitsverursachenden Mutationen von fünf bekannten wie auch 13 nicht zugeordneten FA-Patienten detektiert und somit eine molekulargenetische Diagnose bei neun weiteren, zuvor unklassifizierten FA-Patienten, gestellt werden. Ferner wurden sechs ausgewählte Patienten, zusätzlich zur Panelanreicherung, per Exomanalyse untersucht. Zum einen konnten Mutationen in bekannten FA-Genen bestätigt oder neu identifiziert werden. Zum anderen wurden auch potentiell pathogene Mutationen in DNA-Reparaturgenen außerhalb des FA/BRCA-Signalweges bei zwei Patienten mit unbestätigter Verdachtsdiagnose FA verifiziert. So wurde bei mehreren Mitgliedern einer Familie mit unterschiedlichen Tumorerkrankungen eine zuvor unbeschriebene homozygote Nonsense-Mutation in der BER-Glykosylase NTHL1 nachgewiesen, für welche bislang erst zwei pathogene Mutationen als Auslöser eines neuen Krebssyndroms bekannt sind. Bei einem weiteren Patienten wurden compound-heterozygote Mutationen in RPA1 detektiert, ein Gen für das bislang noch kein Krankheitsbild bekannt ist. Mit Hilfe der drei verschiedenen Anreicherungsstrategien konnten insgesamt 47 von 60 unklassifizierten FA-Patienten 13 verschiedenen Komplementationsgruppen eindeutig zugeordnet werden. Es zeigte sich dabei ein breites Spektrum an neuen, bislang unbeschriebenen FA-Mutationen. Den größten Anteil an der Gesamtzahl der nachgewiesenen Mutationen hatten Spleißmutationen, die auf eine Auswirkung auf das kanonische Spleißmuster untersucht wurden, um einen pathogenen Effekt nachweisen zu können.
Weiterhin schloss die Arbeit die Charakterisierung einzelner FA-Patienten bzw. Komplementationsgruppen mit ein. Dazu zählen die seltenen Untergruppen FA-T und FA-Q, für die jeweils ein neuer Patient identifiziert werden konnte. Durch die funktionelle Charakterisierung der dritten jemals beschriebenen FA-Q-Patientin konnten Einblicke in das Zusammenspiel der Reparatur von DNA-Quervernetzungen und der Nukleotidexzisionsreparatur gewonnen und die phänotypische Variabilität von FA durch die subjektive als auch zelluläre UV-Sensitivität der Patientin ergänzt werden. Darüber hinaus konnte das Mutationsspektrum in FA-I sowie FA-D2 erweitert werden. Eine genauere Untersuchung der Pseudogenregionen von FANCD2 ermöglichte dabei die gezielte Mutationsanalyse des Gens.
Insgesamt konnten die Ergebnisse dieser Arbeit dazu beitragen, das Mutationsspektrum in FA zu erweitern und durch die Identifizierung und Charakterisierung einzelner Patienten neue Einblicke in verschiedene Komponenten des FA/BRCA-Signalweges zu erhalten. Es zeigte sich, dass neue DNA-Sequenzierungsstrategien in der FA-Diagnostik eingesetzt werden können, um eine effiziente Mutationsanalyse zu gewährleisten und klassische Methoden in Teilbereichen zu ersetzen.
CDC14A encodes the Cell Division Cycle 14A protein and has been associated with autosomal recessive non-syndromic hearing loss (DFNB32), as well as hearing impairment and infertile male syndrome (HIIMS) since 2016. To date, only nine variants have been associated in patients whose initial symptoms included moderate-to-profound hearing impairment. Exome analysis of Iranian and Pakistani probands who both showed bilateral, sensorineural hearing loss revealed a novel splice site variant (c.1421+2T>C, p.?) that disrupts the splice donor site and a novel frameshift variant (c.1041dup, p.Ser348Glnfs*2) in the gene CDC14A, respectively. To evaluate the pathogenicity of both loss-of-function variants, we analyzed the effects of both variants on the RNA-level. The splice variant was characterized using a minigene assay. Altered expression levels due to the c.1041dup variant were assessed using RT-qPCR. In summary, cDNA analysis confirmed that the c.1421+2T>C variant activates a cryptic splice site, resulting in a truncated transcript (c.1414_1421del, p.Val472Leufs*20) and the c.1041dup variant results in a defective transcript that is likely degraded by nonsense-mediated mRNA decay. The present study functionally characterizes two variants and provides further confirmatory evidence that CDC14A is associated with a rare form of hereditary hearing loss.
Background
Fabry disease (FD) is an X‐linked lysosomal storage and multi‐system disorder due to mutations in the α‐galactosidase A (α‐GalA) gene. We investigated the impact of individual amino acid exchanges in the α‐GalA 3D‐structure on the clinical phenotype of FD patients.
Patients and methods
We enrolled 80 adult FD patients with α‐GalA missense mutations and stratified them into three groups based on the amino acid exchange location in the α‐GalA 3D‐structure: patients with active site mutations, buried mutations and other mutations. Patient subgroups were deep phenotyped for clinical and laboratory parameters and FD‐specific treatment.
Results
Patients with active site or buried mutations showed a severe phenotype with multi‐organ involvement and early disease manifestation. Patients with other mutations had a milder phenotype with less organ impairment and later disease onset. α‐GalA activity was lower in patients with active site or buried mutations than in those with other mutations (P < 0.01 in men; P < 0.05 in women) whilst lyso‐Gb3 levels were higher (P < 0.01 in men; <0.05 in women).
Conclusions
The type of amino acid exchange location in the α‐GalA 3D‐structure determines disease severity and temporal course of symptom onset. Patient stratification using this parameter may become a useful tool in the management of FD patients.
Hypophosphatasia (HPP) is a rare genetic disease with diverse symptoms and a heterogeneous severity of onset with underlying mutations in the ALPL gene encoding the ectoenzyme Tissue-nonspecific alkaline phosphatase (TNAP). Considering the establishment of zebrafish (Danio rerio) as a new model organism for HPP, the aim of the study was the spatial and temporal analysis of alpl expression in embryos and adult brains. Additionally, we determined functional consequences of Tnap inhibition on neural and skeletal development in zebrafish. We show that expression of alpl is present during embryonic stages and in adult neuronal tissues. Analyses of enzyme function reveal zones of pronounced Tnap-activity within the telencephalon and the mesencephalon. Treatment of zebrafish embryos with chemical Tnap inhibitors followed by axonal and cartilage/mineralized tissue staining imply functional consequences of Tnap deficiency on neuronal and skeletal development. Based on the results from neuronal and skeletal tissue analyses, which demonstrate an evolutionary conserved role of this enzyme, we consider zebrafish as a promising species for modeling HPP in order to discover new potential therapy strategies in the long-term.
Altered autophagy accompanied by abnormal autophagic (rimmed) vacuoles detectable by light and electron microscopy is a common denominator of many familial and sporadic non‐inflammatory muscle diseases. Even in the era of next generation sequencing (NGS), late‐onset vacuolar myopathies remain a diagnostic challenge. We identified 32 adult vacuolar myopathy patients from 30 unrelated families, studied their clinical, histopathological and ultrastructural characteristics and performed genetic testing in index patients and relatives using Sanger sequencing and NGS including whole exome sequencing (WES). We established a molecular genetic diagnosis in 17 patients. Pathogenic mutations were found in genes typically linked to vacuolar myopathy (GNE, LDB3/ZASP, MYOT, DES and GAA), but also in genes not regularly associated with severely altered autophagy (FKRP, DYSF, CAV3, COL6A2, GYG1 and TRIM32) and in the digenic facioscapulohumeral muscular dystrophy 2. Characteristic histopathological features including distinct patterns of myofibrillar disarray and evidence of exocytosis proved to be helpful to distinguish causes of vacuolar myopathies. Biopsy validated the pathogenicity of the novel mutations p.(Phe55*) and p.(Arg216*) in GYG1 and of the p.(Leu156Pro) TRIM32 mutation combined with compound heterozygous deletion of exon 2 of TRIM32 and expanded the phenotype of Ala93Thr‐caveolinopathy and of limb‐girdle muscular dystrophy 2i caused by FKRP mutation. In 15 patients no causal variants were detected by Sanger sequencing and NGS panel analysis. In 12 of these cases, WES was performed, but did not yield any definite mutation or likely candidate gene. In one of these patients with a family history of muscle weakness, the vacuolar myopathy was eventually linked to chloroquine therapy. Our study illustrates the wide phenotypic and genotypic heterogeneity of vacuolar myopathies and validates the role of histopathology in assessing the pathogenicity of novel mutations detected by NGS. In a sizable portion of vacuolar myopathy cases, it remains to be shown whether the cause is hereditary or degenerative.
Filamin C (encoded by the FLNC gene) is a large actin‐cross‐linking protein involved in shaping the actin cytoskeleton in response to signaling events both at the sarcolemma and at myofibrillar Z‐discs of cross‐striated muscle cells. Multiple mutations in FLNC are associated with myofibrillar myopathies of autosomal‐dominant inheritance. Here, we describe for the first time a boy with congenital onset of generalized muscular hypotonia and muscular weakness, delayed motor development but no cardiac involvement associated with a homozygous FLNC mutation c.1325C>G (p.Pro442Arg). We performed ultramorphological, proteomic, and functional investigations as well as immunological studies of known marker proteins for dominant filaminopathies. We show that the mutant protein is expressed in similar quantities as the wild‐type variant in control skeletal muscle fibers. The proteomic signature of quadriceps muscle is altered and ultrastructural perturbations are evident. Moreover, filaminopathy marker proteins are comparable both in our homozygous and a dominant control case (c.5161delG). Biochemical investigations demonstrate that the recombinant mutant protein is less stable and more prone to degradation by proteolytic enzymes than the wild‐type variant. The unusual congenital presentation of the disease clearly demonstrates that homozygosity for mutations in FLNC severely aggravates the phenotype.
Exon-4 Mutations in KRAS Affect MEK/ERK and PI3K/AKT Signaling in Human Multiple Myeloma Cell Lines
(2020)
Approximately 20% of multiple myeloma (MM) cases harbor a point mutation in KRAS. However, there is still no final consent on whether KRAS-mutations are associated with disease outcome. Specifically, no data exist on whether KRAS-mutations have an impact on survival of MM patients at diagnosis in the era of novel agents. Direct blockade of KRAS for therapeutic purposes is mostly impossible, but recently a mutation-specific covalent inhibitor targeting KRAS\(^{p.G12C}\) entered into clinical trials. However, other KRAS hotspot-mutations exist in MM patients, including the less common exon-4 mutations. For the current study, the coding regions of KRAS were deep-sequenced in 80 newly diagnosed MM patients, uniformely treated with three cycles of bortezomib plus dexamethasone and cyclophosphamide (VCD)-induction, followed by high-dose chemotherapy and autologous stem cell transplantation. Moreover, the functional impact of KRAS\(^{p.G12A}\) and the exon-4 mutations p.A146T and p.A146V on different survival pathways was investigated. Specifically, KRAS\(^{WT}\), KRAS\(^{p.G12A}\), KRAS\(^{p.A146T}\), and KRAS\(^{p.A146V}\) were overexpressed in HEK293 cells and the KRAS\(^{WT}\) MM cell lines JJN3 and OPM2 using lentiviral transduction and the Sleeping Beauty vector system. Even though KRAS-mutations were not correlated with survival, all KRAS-mutants were found capable of potentially activating MEK/ERK- and sustaining PI3K/AKT-signaling in MM cells.
Objective
The biological interpretation of gene expression measurements is a challenging task. While ordination methods are routinely used to identify clusters of samples or co-expressed genes, these methods do not take sample or gene annotations into account. We aim to provide a tool that allows users of all backgrounds to assess and visualize the intrinsic correlation structure of complex annotated gene expression data and discover the covariates that jointly affect expression patterns.
Results
The Bioconductor package covRNA provides a convenient and fast interface for testing and visualizing complex relationships between sample and gene covariates mediated by gene expression data in an entirely unsupervised setting. The relationships between sample and gene covariates are tested by statistical permutation tests and visualized by ordination. The methods are inspired by the fourthcorner and RLQ analyses used in ecological research for the analysis of species abundance data, that we modified to make them suitable for the distributional characteristics of both, RNA-Seq read counts and microarray intensities, and to provide a high-performance parallelized implementation for the analysis of large-scale gene expression data on multi-core computational systems. CovRNA provides additional modules for unsupervised gene filtering and plotting functions to ensure a smooth and coherent analysis workflow.
The current molecular genetic diagnostic rates for hereditary hearing loss (HL) vary considerably according to the population background. Pakistan and other countries with high rates of consanguineous marriages have served as a unique resource for studying rare and novel forms of recessive HL. A combined exome sequencing, bioinformatics analysis, and gene mapping approach for 21 consanguineous Pakistani families revealed 13 pathogenic or likely pathogenic variants in the genes GJB2, MYO7A, FGF3, CDC14A, SLITRK6, CDH23, and MYO15A, with an overall resolve rate of 61.9%. GJB2 and MYO7A were the most frequently involved genes in this cohort. All the identified variants were either homozygous or compound heterozygous, with two of them not previously described in the literature (15.4%). Overall, seven missense variants (53.8%), three nonsense variants (23.1%), two frameshift variants (15.4%), and one splice-site variant (7.7%) were observed. Syndromic HL was identified in five (23.8%) of the 21 families studied. This study reflects the extreme genetic heterogeneity observed in HL and expands the spectrum of variants in deafness-associated genes.
Tissue-nonspecific alkaline phosphatase (TNAP) is a ubiquitously expressed enzyme that is best known for its role during mineralization processes in bones and skeleton. The enzyme metabolizes phosphate compounds like inorganic pyrophosphate and pyridoxal-5′-phosphate to provide, among others, inorganic phosphate for the mineralization and transportable vitamin B6 molecules. Patients with inherited loss of function mutations in the ALPL gene and consequently altered TNAP activity are suffering from the rare metabolic disease hypophosphatasia (HPP). This systemic disease is mainly characterized by impaired bone and dental mineralization but may also be accompanied by neurological symptoms, like anxiety disorders, seizures, and depression. HPP characteristically affects all ages and shows a wide range of clinical symptoms and disease severity, which results in the classification into different clinical subtypes. This review describes the molecular function of TNAP during the mineralization of bones and teeth, further discusses the current knowledge on the enzyme’s role in the nervous system and in sensory perception. An additional focus is set on the molecular role of TNAP in health and on functional observations reported in common laboratory vertebrate disease models, like rodents and zebrafish.
Inherited cardiomyopathies are characterized by clinical and genetic heterogeneity that challenge genetic diagnostics. In this study, we examined the diagnostic benefit of exome data compared to targeted gene panel analyses, and we propose new candidate genes. We performed exome sequencing in a cohort of 61 consecutive patients with a diagnosis of cardiomyopathy or primary arrhythmia, and we analyzed the data following a stepwise approach. Overall, in 64% of patients, a variant of interest (VOI) was detected. The detection rate in the main sub-cohort consisting of patients with dilated cardiomyopathy (DCM) was much higher than previously reported (25/36; 69%). The majority of VOIs were found in disease-specific panels, while a further analysis of an extended panel and exome data led to an additional diagnostic yield of 13% and 5%, respectively. Exome data analysis also detected variants in candidate genes whose functional profile suggested a probable pathogenetic role, the strongest candidate being a truncating variant in STK38. In conclusion, although the diagnostic yield of gene panels is acceptable for routine diagnostics, the genetic heterogeneity of cardiomyopathies and the presence of still-unknown causes favor exome sequencing, which enables the detection of interesting phenotype–genotype correlations, as well as the identification of novel candidate genes.
Defects of platelet intracellular signaling can result in severe platelet dysfunction. Several mutations in each of the linked genes FERMT3 and RASGRP2 on chromosome 11 causing a Glanzmann‐like bleeding phenotype have been identified so far. We report on novel variants in two unrelated pediatric patients with severe bleeding diathesis—one with leukocyte adhesion deficiency type III due to a homozygous frameshift in FERMT3 and the other with homozygous variants in both, FERMT3 and RASGRP2 . We focus on the challenging genetic and functional variant assessment and aim to accentuate the risk of obtaining misleading results due to the phenomenon of genetic linkage.
Die Arbeit zeigt, dass die Symptome der HPP sehr variabel und unterschiedlich stark auftreten können. Dies erschwert die klinische Diagnosestellung der Erkrankung. Nahezu alle Patienten berichteten von starken Knochen-, Gelenk,- und Muskelschmerzen, von Karies und Parodontose sowie von vermehrten Frakturen, die zum Teil weitere chronische Schmerzen und Wiederholungsfrakturen erzeugen. Eine deutlich verminderte Leistungsfähigkeit im Vergleich zu Gleichaltrigen wurde ebenso häufig angegeben. Es konnte keine eindeutige Phänotyp - Genotyp Korrelation gefunden werden, allerdings geben die Daten einen deutlichen Hinweis, dass Patienten mit zwei Mutationen am stärksten symptomatisch betroffen sind.
Ebenfalls konnten keine Unterschiede zwischen dominant negativen Mutationen und nicht dominant negativen Mutationen gefunden werden.
Werner Syndrome (WS) is an adult‐onset segmental progeroid syndrome. Bisulfite pyrosequencing of repetitive DNA families revealed comparable blood DNA methylation levels between classical (18 WRN‐mutant) or atypical WS (3 LMNA‐mutant and 3 POLD1‐mutant) patients and age‐ and sex‐matched controls. WS was not associated with either age‐related accelerated global losses of ALU, LINE1, and α‐satellite DNA methylations or gains of rDNA methylation. Single CpG methylation was analyzed with Infinium MethylationEPIC arrays. In a correspondence analysis, atypical WS samples clustered together with the controls and were clearly separated from classical WS, consistent with distinct epigenetic pathologies. In classical WS, we identified 659 differentially methylated regions (DMRs) comprising 3,656 CpG sites and 613 RefSeq genes. The top DMR was located in the HOXA4 promoter. Additional DMR genes included LMNA, POLD1, and 132 genes which have been reported to be differentially expressed in WRN‐mutant/depleted cells. DMRs were enriched in genes with molecular functions linked to transcription factor activity and sequence‐specific DNA binding to promoters transcribed by RNA polymerase II. We propose that transcriptional misregulation of downstream genes by the absence of WRN protein contributes to the variable premature aging phenotypes of WS. There were no CpG sites showing significant differences in DNA methylation changes with age between WS patients and controls. Genes with both WS‐ and age‐related methylation changes exhibited a constant offset of methylation between WRN‐mutant patients and controls across the entire analyzed age range. WS‐specific epigenetic signatures occur early in life and do not simply reflect an acceleration of normal epigenetic aging processes.
Die Pierre-Robin-Sequenz ist eine angeborene kraniofaziale Fehlbildung, bei der häufig eine Triade von Symptomen, bestehend aus mandibulärer Mikrognathie/Retrognathie, Glossoptose und einer Gaumenspalte, beobachtet werden kann. Aufgrund der Heterogenität der PRS und der häufigen Vergesellschaftung mit Syndromen, konnten Ätiologie und Pathogenese der PRS bisher nur unzureichend geklärt werden. Für einen Teil der Patienten mit isolierter PRS konnte eine familiäre Häufung von PRS-Fällen nachgewiesen werden, was auf eine erbliche Komponente als krankheitsauslösenden Faktor hinweist. In diesem Zusammenhang konnten bei Patienten mit isolierter PRS gehäuft genetische Veränderungen mit einer Entfernung von über 1Mb zentromerisch (5´) von SOX9 auf dem Chromosom 17 detektiert werden. Es wird vermutet, dass diese genetischen Aberrationen am SOX9 Lokus eine gewebsspezifische Fehlregulation von SOX9 während der Embryonalentwicklung auslösen und somit ursächlich für die Entstehung von PRS sein können.
Das Ziel dieser Arbeit war es, eine Würzburger Patientenkohorte mit isolierter PRS zu gewinnen und Informationen über die phänotypischen Merkmale der Studienteilnehmer auszuwerten. Im Anschluss sollte die Patienten-DNS mittels molekulargenetischen Analysemethoden auf potenziell krankheitsauslösende genetische Aberrationen am SOX9 Lokus untersucht werden.
Zunächst konnte eine Kohorte mit sieben PRS-Patienten erstellt und Informationen über die phänotypischen Krankheitsmerkmale erfasst und ausgewertet werden. Anschließend wurden bei den Studienteilnehmern eine Array-CGH, eine quantitative Echtzeit-Polymerase-Kettenreaktion und im Bereich von drei konservierten, potenziell regulatorischen Elementen des SOX9 Lokus eine Sanger Sequenzierung durchgeführt. Die Array-CGH ergab zunächst bei einem Patienten zwei große Deletionen im regulativen Umfeld des SOX9 Lokus, welche im Weiteren nicht durch qPCR bestätigt werden konnten. Letztendlich konnten durch die Sanger Sequenzierung 22 Varianten detektiert werden, wovon für drei Einzelnukleotid-Polymorphismen eine prädisponierende Wirkung diskutierbar und für zwei Einzelnukleotid-Varianten eine ursächlich pathogene Wirkung nicht auszuschließen ist.
Background
Estimation of incidence in rare diseases is often challenging due to unspecific and incomplete coding and recording systems. Patient- and health care provider-driven data collections are held with different organizations behind firewalls to protect the privacy of patients. They tend to be fragmented, incomplete and their aggregation leads to further inaccuracies, as the duplicated records cannot easily be identified. We here report about a novel approach to evaluate the incidences of Duchenne muscular dystrophy (DMD) and spinal muscular atrophy (SMA) in Germany.
Methods
We performed a retrospective epidemiological study collecting data from patients with dystrophinopathies (DMD and Becker muscular dystrophy) and SMA born between 1995 and 2018. We invited all neuromuscular centers, genetic institutes and the patient registries for DMD and SMA in Germany to participate in the data collection. A novel web-based application for data entry was developed converting patient identifying information into a hash code. Duplicate entries were reliably allocated to the distinct patient.
Results
We collected 5409 data entries in our web-based database representing 1955 distinct patients with dystrophinopathies and 1287 patients with SMA. 55.0% of distinct patients were found in one of the 3 data sources only, while 32.0% were found in 2, and 13.0% in all 3 data sources. The highest number of SMA patients was reported by genetic testing laboratories, while for DMD the highest number was reported by the clinical specialist centers. After the removal of duplicate records, the highest yearly incidence for DMD was calculated as 2.57:10,000 in 2001 and the highest incidence for SMA as 1.36:10,000 in 2014.
Conclusion
With our novel approach (compliant with data protection regulations), we were able to identify unique patient records and estimate the incidence of DMD and SMA in Germany combining and de-duplicating data from patient registries, genetic institutes, and clinical care centers. Although we combined three different data sources, an unknown number of patients might not have been reported by any of these sources. Therefore, our results reflect the minimal incidence of these diseases.
Background
Inherited pathogenic variants in BRCA1 and BRCA2 are the most common causes of hereditary breast and ovarian cancer (HBOC). The risk of developing breast cancer by age 80 in women carrying a BRCA1 pathogenic variant is 72%. The lifetime risk varies between families and even within affected individuals of the same family. The cause of this variability is largely unknown, but it is hypothesized that additional genetic factors contribute to differences in age at onset (AAO). Here we investigated whether truncating and rare missense variants in genes of different DNA-repair pathways contribute to this phenomenon.
Methods
We used extreme phenotype sampling to recruit 133 BRCA1-positive patients with either early breast cancer onset, below 35 (early AAO cohort) or cancer-free by age 60 (controls). Next Generation Sequencing (NGS) was used to screen for variants in 311 genes involved in different DNA-repair pathways.
Results
Patients with an early AAO (73 women) had developed breast cancer at a median age of 27 years (interquartile range (IQR); 25.00–27.00 years). A total of 3703 variants were detected in all patients and 43 of those (1.2%) were truncating variants. The truncating variants were found in 26 women of the early AAO group (35.6%; 95%-CI 24.7 - 47.7%) compared to 16 women of controls (26.7%; 95%-CI 16.1 to 39.7%). When adjusted for environmental factors and family history, the odds ratio indicated an increased breast cancer risk for those carrying an additional truncating DNA-repair variant to BRCA1 mutation (OR: 3.1; 95%-CI 0.92 to 11.5; p-value = 0.07), although it did not reach the conventionally acceptable significance level of 0.05.
Conclusions
To our knowledge this is the first time that the combined effect of truncating variants in DNA-repair genes on AAO in patients with hereditary breast cancer is investigated. Our results indicate that co-occurring truncating variants might be associated with an earlier onset of breast cancer in BRCA1-positive patients. Larger cohorts are needed to confirm these results.
Genome-wide association studies (GWAS) have identified more than 170 breast cancer susceptibility loci. Here we hypothesize that some risk-associated variants might act in non-breast tissues, specifically adipose tissue and immune cells from blood and spleen. Using expression quantitative trait loci (eQTL) reported in these tissues, we identify 26 previously unreported, likely target genes of overall breast cancer risk variants, and 17 for estrogen receptor (ER)-negative breast cancer, several with a known immune function. We determine the directional effect of gene expression on disease risk measured based on single and multiple eQTL. In addition, using a gene-based test of association that considers eQTL from multiple tissues, we identify seven (and four) regions with variants associated with overall (and ER-negative) breast cancer risk, which were not reported in previous GWAS. Further investigation of the function of the implicated genes in breast and immune cells may provide insights into the etiology of breast cancer.