Abstract EU has reclassified the sika deer ( Cervus nippon ) as an undesirable invasive species based on reports that hybridization with the indigenous red deer ( C. elaphus ) may produce fertile offspring. Since sika-derived DNA previosuly introduced into the red deer population (introgression) cannot be removed, the crucial question is whether new (F1) hybridisation occur. To address this, we analysed the chromosomes in 56 sika and 22 red deer. All red deer had a chromosome number 2n=68. In contrast, the chromosome number in sika ranged from 64 to 67, due to the variable presence of two sika-specific Robertsonian translocations (ROB1,ROB2). In the free-ranging sika population in Jutland, >90% of the sika deer were homozygote for at least one of these ROBs, excluding that they could be F1-hybrids. Moreover, ROB2 was in Hardy-Weinberg equilibrium, further supporting the absence of gene flow between the two species. In contrast, ROB1 was in Hardy-Weinberg disequilibrium, suggesting negative fitness of heterozygotes, including potential F1-hybrids. In Jægersborg Deer Park, the eight examined sika deer had the same genotype (absence of ROB1, homozygosity of ROB2), supporting that it is a founder population which may have been isolated for ∼100 years. Again, none of these can be F1-hybrids due to the homozygosity of ROB2. We conclude that F1-hybridisation between sika and red deer either does not occur or occur very rarely in Denmark. The study establish the Danish sika-populations as unique models for adressing important biological questions: What underlies the absence of hybridisation? Why are ROBs frequent in sika deer but not in the closely related red deer? How fast do new species/subspecies develop in isolated founder populations? Which factors determine, that some ROBs have little heterozygous effects, whereas others are selected against, with implications for the role of ROBs as genetic barriers promoting speciation, and for fertility problems in some human ROB carriers.

Topologically associating domains (TADs) are generally considered as a homogeneous basic units of genome folding, which is critical for transcriptional regulation. However, recent studies indicate that both the TAD domain structures and the boundaries between them are not as homogeneous as originally recognized. Here, we address the heterogeneity of the TAD boundaries in the human genome at a large scale, which varies between active and inactive chromatin and across cell lines and tissues. To address this, based on the well-annotated TAD boundaries extracted from multiple cell lines and tissues, we examine their nucleotide content, resulting in two main clusters, one GC-rich and one AT-rich, which are mainly distributed in active and inactive chromatin, respectively. Also, they contain different types of repetitive sequences and have different epigenetic patterns, with more CTCF binding motifs in the GC-rich cluster. Hence, our observations of the TAD boundary content provide novel insights into TAD genomic architecture. In addition, we find that cell- or tissue-specific boundaries are less evolutionarily conserved than other boundaries. We highlight the importance of TAD boundary diversity in different functional contexts and discuss the importance of the different types of repetitive sequences and epigenetic patterns in the two main types of boundaries.

A fundamental understanding of genome organization relies on accurately annotating topologically associating domains (TADs) and their boundaries. This is crucial for understanding how cis-regulatory elements regulate gene expression. To go beyond calling TADs and boundaries from Hi-C data, several machine learning-based methods have been proposed to go the step further and predict TAD boundaries from genomic sequences. As the growing evidence of TADs and their boundaries, TADs have been proved exhibiting diverse properties, such as differences in replication timing and epigenetic patterns. However, existing methods do not take this heterogeneity into account. To address this, we propose a method called TADBpred for TAD boundary prediction in a large genomic context in humans. TADBpred focuses on TAD boundaries in active and inactive chromatin across cell-lines and tissues, which are GC-rich and AT-rich, respectively. By integrating genomic elements and sequence composition, we designed two models for GC-rich and AT-rich boundaries, respectively. When testing the performance on respective independent held-out datasets, we obtain AUC scores of 0.91 and 0.80. Our results indicate that TADBpred excels in TAD boundary prediction. Additionally, feature importance analysis highlights the essential features for different classes of TAD boundaries, thereby enhancing our understanding of these TAD boundaries.

The host genetics of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have previously been studied based on cases from the earlier waves of the pandemic in 2020 and 2021, identifying 51 genomic loci associated with infection and/or severity. SARS-CoV-2 has shown rapid sequence evolution, increasing transmissibility, particularly for Omicron variants, which raises the question of whether this affected the host genetic factors. We performed a genome-wide association study of SARS-CoV-2 infection with Omicron variants, including more than 150,000 cases from four cohorts. We identified 13 genome-wide significant loci, of which only five were previously described as associated with SARS-CoV-2 infection. The strongest signal was a single nucleotide polymorphism in an intron of ST6GAL1, a gene affecting immune development and function, connected to three other associated loci (harboring MUC1, MUC5AC and MUC16) through O-glycan biosynthesis. Our study provides robust evidence for individual genetic variation related to glycosylation, translating into susceptibility to SARS-CoV-2 infections with Omicron variants. Genome-wide analyses identify 13 loci associated with susceptibility to infection with SARS-CoV-2 Omicron variants, including variation in ST6GAL1, previously associated with influenza susceptibility, and other genes involved in glycosylation processes.

Females inactivate one of their two X-chromosomes in each cell before implantation, at a time where any factor that affect the number of these primordial cells may lead to deviation from the usual random choice. Thus, non-random (skewed) X-inactivation may occur due to early stochastic effects in the limited primordial pool size. The primodial pool size can be calculated from the variance of the ratio between the unmethylated and methylated CAG-repeats within exon 1 of the androgen receptor gene (AR) corresponding to the active and inactive X-chromosome (X-inactivation (XI)-ratio), respectively. Since maternal smoking during pregnancy is associated with a general growth inhibition and a reduced number of fetal gonadal cells, we have tested the XI-ratios in tissues obtained from fetuses of smoking (n = 8) and non-smoking (n = 10) mothers in connection with legal termination of the pregnancy. A tighter distribution of XI-ratios was seen in tissues from fetuses in the smoking group. Combined with more skewed X-inactivation (>=80 %) and a higher mean XI-ratio in the fetal samples from pregnancies of non-smoking mothers, this support a larger pool of primordial cells in fetuses exposed to smoking, suggesting that smoking during pregnancy delays fetal X inactivation.

BACKGROUND:Primary congenital glaucoma (PCG) is a rare genetic disorder affecting the ocular drainage system, accounting for only 0.01-0.04% blindness related cases. However, its prevalence varies significantly in ethnicities, being higher in populations that practice consanguinity, such as Pakistan where approximately 70% of marriages are consanguineous. This study aimed to investigate the genetic cause of PCG in a large Pakistani family with autosomal recessive inheritance. METHODS:A large multigenerational family having multiple consanguineous marriages resulting in fifteen affected individuals was recruited for the current study. All relevant clinical information was collected and venous blood drawn for further genetic analysis. The family was subjected to direct sequencing of CYP1B1 which is the most plausible candidate of PCG. The resulting candidate variant was further confirmed using BanII restriction enzyme analysis. RESULTS:The sequence analysis revealed a novel indel (c.862delinsCC) in exon 2 of the CYP1B1 gene, resulting in a frameshift mutation (p.Ala288Profs*39) thereby creating a premature stop codon 39 amino acids downstream. BanII restriction enzyme analysis further confirmed this putative null mutation co-segregating with the disease trait in all the family members of the pedigree. CONCLUSION:The novel indel, putative null mutation causes PCG related disease phenotypes. This genetic variant has a high penetrance but shows variable expressivity among the affected members of the family. This putative null mutation enhances the mutation spectrum of CYP1B1 globally and from Pakistan in particular.

We reanalyzed through a cytogenomics approach a case published 20 years ago, describing a girl with developmental delay and epilepsy. Karyotype and FISH analysis showed a de novo 2.3 Mb terminal inverted-duplication at 8q24.3. The interpretation was inconsistent with the absence of a more distal deletion as expected for distal inverted duplications, and it was inconceivable to highlight rearrangements smaller than 5–10 Mb at that time. Chromosomal microarray (CMA), optical genome mapping (OGM), and short-read whole genome sequencing (srWGS) identified a complex configuration at 8q24.3, which resembles events like chromoanasynthesis or DUP-TRP/INV-DUP (duplication-triplication/inverted-duplication), both characterized by clustered duplications and triplications, some of which are inverted. In the EBV-line genes located in the amplified regions were overexpressed. Despite a more precise definition of the imbalance, we were unable to provide a clear-cut explanation for the proband’s clinical features.

Silver-Russell Syndrome (SRS) is a genetic disorder characterized by intrauterine and postnatal growth restriction. Most cases are caused by an imprinting error either with hypomethylation of the Imprinted Control Region 1 at 11p15.5, or maternal uniparental disomy of chromosome 7. Approximately 40% of the cases have unknown etiology, thus distinct mechanisms have been described in association with the syndrome. Here, we present a case of monozygotic twin sisters with a clinical diagnosis of SRS, mild intellectual disability and epilepsy who carry a balanced translocation between chromosomes 3 and 12 that interrupts the NAALADL2 and HMGA2 genes, respectively. Disruption of HMGA2, a gene previously described as causative of SRS, confirms the initial diagnosis. NAALADL2 gene has been recently proposed as a candidate for intellectual disability and could partially contribute to our patient's phenotype.

Background: The growing body of knowledge on the human genome and its variants points towards the significance of genetic factors in oral health and disease. Since the dental curricula have historically prioritized clinically oriented subjects, this focus has resulted in insufficient coverage of genetics. To leverage this knowledge in patient care, dental education must equip students with an understanding of the principles of genetics. Method: We have established “Genetic Educators Network in Dentistry” (GEN-Dent) to identify common concerns regarding genetics in dental education and work for a greater emphasis on genetics in future dental programs to make sure that professionals in dentistry are well-prepared to navigate the complexities of the evolving “human genome era”. Results: Here, GEN-Dent proposes specific learning goals for medical genetics in dentistry and provides supporting teaching material addressing each learning goal. The five life-like case studies exemplify different dental conditions and introduce important concepts of genetics, inspiring other educators. Conclusions: Opportunities in Scandinavian countries can be an advantage in increasing global awareness of the importance of genetics in dentistry. The integration of genetics into dental education not only aims to improve patient care but also seeks to inspire a new generation of basic scientists with clinical backgrounds in dentistry. We expect that using life-like patient cases will significantly motivate dental students when learning medical genetics.

Background Mesomelia-Synostoses Syndrome (MSS)(OMIM 600,383) is a rare autosomal dominant disorder characterized by mesomelic limb shortening, acral synostoses and multiple congenital malformations which is described as a contiguous deletion syndrome involving the two genes SULF1 and SLCO5A1. The study of apparently balanced chromosomal rearrangements (BCRs) is a cytogenetic strategy used to identify candidate genes associated with Mendelian diseases or abnormal phenotypes. With the improved development of genomic technologies, new methods refine this search, allowing better delineation of breakpoints as well as more accurate genotype-phenotype correlation. Case presentation We present a boy with a global development deficit, delayed speech development and an ASD (Asperger) family history, with an apparently balanced "de novo" reciprocal translocation [t(1;8)(p32.2;q13)dn]. The cytogenetic molecular study identified a likely pathogenic deletion of 21 kb in the 15q12 region, while mate pair sequencing identified gene-truncations at both the 1p32.2 and 8q13 translocation breakpoints. Conclusions The identification of a pathogenic alteration on 15q12 involving GABRA5 was likely the main cause of the ASD-phenotype. Importantly, the chr8 translocation breakpoint truncating SLCO5A1 exclude SLCO5A1 as a candidate for MSS, leaving SULF1 as the primary candidate. However, the deletions observed in MSS remove a topological associated domain (TAD) boundary separating SULF1 and SLCO5A1. Hence, Mesomelia-Synostoses syndrome is either caused by haploinsufficiency of SULF1 or ectopic enhancer effects where skeletal/chrondrogenic SULF1 enhancers drive excopic expression of developmental genes in adjacent TADs including PRDM14, NCOA2 and/or EYA1.

Clonal hematopoiesis of indeterminate potential (CHIP) refers to the nonmalignant clonal expansion of blood stem cells that carry somatic mutations in myeloid cancer-associated genes.1 The main driver of CHIP progression is age, but other factors like smoking or chemotherapy exposure can also have significant effects.2 CHIP has been associated with an increased risk of hematological malignancies and a range of other age-related conditions, including severe infections and death.1, 3 The basis for this link is suggested to be an altered inflammatory response, which could occur as a result of mutations that impair the regulation of proinflammatory factor secretion from myeloid cells.4-6 Over 3 years have passed since the World Health Organization declared the coronavirus disease 2019 (COVID-19) a global pandemic. Caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), 773 million infections and 6.9 million deaths have been reported worldwide as of January 2024.7 The clinical presentation of SARS-CoV-2 infection varies widely, ranging from asymptomatic to more severe responses that result in acute respiratory distress syndrome, intensive care unit (ICU) admission, or death.8 The hyperinflammatory response that characterizes the severe form of COVID-19,9 as well as the significantly higher risk of older, more comorbid males to suffer from it,10 closely resembles the CHIP phenotype and thus has raised the question of whether CHIP might influence COVID-19 severity. Several studies have addressed this,11-17 but their widely variable study designs, phenotype definitions, and patient cohorts have generated mostly conflicting results. The current matched case–control study included 470 Danish individuals with a positive SARS-CoV-2 polymerase chain reaction (PCR) test between March 1, 2020 and December 1, 2021. They were selected to be over 60 years of age at the time of the PCR test and to have no record of a hematological malignancy previously associated with CHIP (ICD10:C81-90/92-96) in the Danish National Patient Registry (DNPR). Five individuals with chronic lymphoid leukemia (ICD10:C91.1) were identified, but still included in the selection, as this type of cancer has not been associated with a higher CHIP prevalence.18 As part of the Danish COVID-19 Genetic Consortium,19 all participants had an available biobank sample for genetic analysis. Scientific ethics permission for this study was obtained from the Danish National and Capital Region Ethics Committees (NVK-2003947 and H-20026501). Written consent was required from all living individuals. Individuals who died due to COVID-19 before January 6, 2021 were included without consent, as approved by the regional Ethics Committee. More information about the Consortium and the ethical approvals can be found in Supporting Information S2: Methods. We assessed the association between CHIP and COVID-19 severity in our population using univariable and multivariable logistic regressions. We adjusted the models by including a selection of lifestyle and clinical covariates known to be associated with both CHIP progression and severe COVID-1910: body mass index (BMI), smoking status (active smoking vs. smoking in the past or never), diabetes, cardiovascular disease, lung disease, cancer, and prior chemo- and/or radiotherapy treatment. These data were extracted from hospital-administered diagnoses and treatments in the DNPR, self-reported questionnaires,19 patient records from hospital admissions, the national intensive care unit database, and the COVIMUN study database. More details on participant information retrieval and missing data handling are given in Supporting Information S2: Methods and in Supporting Information S1: Figures 1–3. Statistical analyses were performed using R.20 Covariate effects can be found in Supporting Information S1: Figures 4 and 5. To identify CHIP mutations, we performed deep targeted sequencing of peripheral whole blood for 31 genes recurrently mutated in myeloid cancer, using a custom capture-based panel from Twist Biosciences. We limited our analysis to variants with a variant allele fraction (VAF) ≥ 2%. More details regarding CHIP typing are provided in the Supporting Information S2: Methods. In the primary analysis, we compared 235 individuals who had been hospitalized within 14 days of the positive SARS-CoV-2 PCR test, to 235 with no hospitalization. Hospitalized cases and nonhospitalized controls were matched one-to-one on sex, age at the time of the positive SARS-CoV-2 PCR test (±12 months), and sample age (±14 months), defined as the number of months between the blood sample collection and the positive SARS-CoV-2 PCR test (Figure 1A and Supporting Information S2: Methods). The matching by age and sex was performed due to the known strong associations with both CHIP prevalence and COVID-19 severity. The matching by blood sample age aimed at eliminating any bias that could have arisen from the expansion of CHIP over time, as blood samples were collected over several years at the biobanks. We chose to further adjust the regressions for blood sample age to account for any residual effects of the variable; sex and age were considered sufficiently matched for additional adjustments to be necessary (Supporting Information S1: Figures 6 and 7). We identified a total of 187 mutations in 143 out of 470 study participants (30.4%). The most frequently mutated genes were DNMT3A, TET2, ASXL1, PPM1D, and TP53 (Figure 1B). CHIP was observed in 81 (34.5%) of the 235 hospitalized cases, and 62 (26.4%) of the 235 nonhospitalized controls. Multiple mutations were found in 22 (9.4%) cases and eight (3.4%) controls. Clones with a VAF ≥ 10% were detected in 32 (13.6%) cases and 18 (7.7%) controls. CHIP mutations within DNA repair genes (PPM1D and/or TP53) were seen in 13 (5.5%) cases and eight (3.4%) controls. Mutations within PPM1D, previously found to be associated with severe COVID-19 in the large study by Kessler et al.,11 were present in 10 (4.3%) cases and three (1.3%) controls. In the covariate-adjusted model (Figure 1C), presenting multiple CHIP clones—compared to one or none—or large (VAF ≥ 10%) mutation(s)—compared to small (VAF < 10%) or none—was statistically significantly associated with COVID-19 hospitalization. The effects of carrying any CHIP clone, having mutations in DNA repair genes (PPM1D and/or TP53), or in PPM1D specifically were not statistically significant. However, all effect size estimates were positive, suggestive of an overrepresentation of the different CHIP phenotypes among hospitalized COVID-19 patients. We also performed a secondary analysis on the subset of hospitalized participants, where we compared 123 patients who had received a general admission to 112 who had been further admitted into the ICU within 14 days of the positive test (28 patients) or had died at the hospital within 30 days of the test (84 patients; Figure 2A,B). Importantly, participants' sex, age at the time of the positive SARS-CoV-2 PCR test, and sample age were no longer matched between the two outcome groups in this modified setup, so we included them as covariates in the adjusted regressions. We did not find any statistically significant associations between carrying CHIP, multiple clones, large (VAF ≥ 10%) mutation(s), or mutation(s) in a DNA repair gene, and an ICU admission or in-hospital death, compared to a general admission (Figure 2C). There was only a borderline significant overrepresentation of mutations in PPM1D among participants with an ICU admission or an in-hospital death. In summary, we observed that the risk of COVID-19 hospitalization increased with the presence of multiple or large (VAF ≥ 10%) CHIP clone(s) in this study of 470 Danish individuals, PCR-confirmed positive for SARS-CoV-2 while 60–89 years old. These results are consistent with those of the large study by Kessler et al.,11 but not of the smaller one by Zhou et al.13 In the subset of 235 COVID-19 hospitalized cases, we did not find CHIP to be a risk factor for ICU admission or in-hospital death, in contrast to general admission, which is also consistent with findings by Duployez et al.,14 Hameister et al.,15 Petzer et al.,16 Miller et al.,12 and Del Pozo-Valero et al.17 However, these results should be interpreted with caution as our study was designed as a matched case–control study between hospitalized and nonhospitalized COVID-19. The previously observed association between PPM1D CHIP and severe COVID-19 by Kessler et al.11 was only suggestively replicated in our data and may grant further investigation. The main limitations of our study include the collection of blood samples across several years, which could bias our measurement of CHIP but is mitigated through matching; the use of self-reported and hospital-recorded lifestyle and clinical information, which can have different accuracies; and our inability to differentiate individuals with past smoking habits from those who have never smoked. Overall, the findings presented add to our understanding of the effect of CHIP on inflammation and infectious diseases, and emphasize the importance of disease prophylaxis such as vaccinations in CHIP carriers. We are deeply grateful to all of the patients for their participation. Morten Tulstrup and Kirsten Grønbæk conceived and designed the study. Sofie Bliddal, Ioanna Nissen, Sisse Rye Ostrowski, and Karina Meden Sørensen secured participant consent and samples from biobanks. Jakob Schmidt Jespersen handled library preparation and DNA sequencing. Morten Tulstrup handled variant calling and filtering. Celia Burgos Sequeros, Karina Banasik, Sofie Bliddal, Omid Rezahosseini, and Morten Tulstrup planned the statistical analyses. Ioanna Nissen and Ole Birger Vestager Pedersen were in charge of ethical and data permissions and secured data from the Danish registers. Sofie Bliddal collected data from patients' records. Celia Burgos Sequeros analyzed the data and wrote the first draft of the manuscript. Celia Burgos Sequeros, Morten Tulstrup, and Kirsten Grønbæk interpreted the data. All co-authors contributed to the sample or data collection and critically revised the manuscript. Kirsten Grønbæk has received research funding and/or consultancy fees from Janssen Pharma and GSK. Carsten Utoft Niemann has received research funding and/or consultancy fees from Abbvie, AstraZeneca, Janssen, Octapharma, Beigene, Genmab, CSL Behring, Takeda, Lilly, and MSD. Anne-Mette Lebech has received unrestricted research grant from Gilead and consultancy fees from GSK, MSD, and Pfizer. This study was supported by a research grant from the European Hematology Association (EHA COVID-19 in Hematology Research Grant 2020). In addition, the study is part of the Danish Research Center for Precision Medicine in Blood Cancers (Danish Cancer Society Grant R223-A13071). A grant from Sygesikring Danmark (2020-0178) was given to the Danish COVID-19 Genetic Consortium. Celia Burgos Sequeros, Karina Banasik, and Søren Brunak acknowledge the Novo Nordisk Foundation (Grant numbers: NNF17OC0027594 and NNF14CC0001). This work was also supported by a grant from the Ministry of Higher Education and Science, Copenhagen, Denmark (0238-00006B). This research has been conducted using the Danish National Biobank resource. UFR's research salary was sponsored by Kirsten and Freddy Johansen's Fund. The data that support the findings of this study are available on request from the corresponding author. The data are not publicly available due to privacy or ethical restrictions. Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.

ABSTRACT Balanced chromosomal rearrangements (BCRs), including inversions, translocations, and insertions, reorganize large sections of the genome and contribute substantial risk for developmental disorders (DDs). However, the rarity and lack of systematic screening for BCRs in the population has precluded unbiased analyses of the genomic features and mechanisms associated with risk for DDs versus normal developmental outcomes. Here, we sequenced and analyzed 1,420 BCR breakpoints across 710 individuals, including 406 DD cases and the first large-scale collection of 304 control BCR carriers. We found that BCRs were not more likely to disrupt genes in DD cases than controls, but were seven-fold more likely to disrupt genes associated with dominant DDs (21.3% of cases vs. 3.4% of controls; P = 1.60×10 −12 ). Moreover, BCRs that did not disrupt a known DD gene were significantly enriched for breakpoints that altered topologically associated domains (TADs) containing dominant DD genes in cases compared to controls (odds ratio [OR] = 1.43, P = 0.036). We discovered six TADs enriched for noncoding BCRs (false discovery rate < 0.1) that contained known DD genes ( MEF2C, FOXG1, SOX9, BCL11A, BCL11B , and SATB2 ) and represent candidate pathogenic long-range positional effect (LRPE) loci. These six TADs were collectively disrupted in 7.4% of the DD cohort. Phased Hi-C analyses of five cases with noncoding BCR breakpoints localized to one of these putative LRPEs, the 5q14.3 TAD encompassing MEF2C , confirmed extensive disruption to local 3D chromatin structures and reduced frequency of contact between the MEF2C promoter and annotated enhancers. We further identified six genomic features enriched in TADs preferentially disrupted by noncoding BCRs in DD cases versus controls and used these features to build a model to predict TADs at risk for LRPEs across the genome. These results emphasize the potential impact of noncoding structural variants to cause LRPEs in unsolved DD cases, as well as the complex interaction of features associated with predicting three-dimensional chromatin structures intolerant to disruption.

Purpose Heterozygous variants in PRRT2 are mostly associated with benign phenotypes, being the major genetic cause of benign familial infantile seizures (BFIS), as well as in paroxysmal disorders. We report two children from unrelated families with BFIS that evolved to encephalopathy related to status epilepticus during sleep (ESES). Methods and results Two probands presented with focal motor seizures at 3 months of age, with a limited course. Both children presented, at around 5 years of age, with centro-temporal interictal epileptiform discharges with a source in the frontal operculum, markedly activated by sleep, and associated with stagnation on neuropsychological development. Whole-exome sequencing and co-segregation analysis revealed a frameshift mutation c.649dupC in the proline-rich transmembrane protein 2 (PRRT2) in both probands and all affected family members. Conclusion The mechanism leading to epilepsy and the phenotypic variability of PRRT2 variants remain poorly understood. However, its wide cortical and subcortical expression, in particular in the thalamus, could partially explain both the focal EEG pattern and the evolution to ESES. No variants in the PRRT2 gene have been previously reported in patients with ESES. Due to the rarity of this phenotype, other possible causative cofactors are likely contributing to the more severe course of BFIS in our probands.

Structural variants are a common cause of disease and contribute to a large extent to inter-individual variability, but their detection and interpretation remain a challenge. Here, we investigate 11 individuals with complex genomic rearrangements including germline chromothripsis by combining short- and long-read genome sequencing (GS) with Hi-C. Large-scale genomic rearrangements are identified in Hi-C interaction maps, allowing for an independent assessment of breakpoint calls derived from the GS methods, resulting in >300 genomic junctions. Based on a comprehensive breakpoint detection and Hi-C, we achieve a reconstruction of whole rearranged chromosomes. Integrating information on the three-dimensional organization of chromatin, we observe that breakpoints occur more frequently than expected in lamina-associated domains (LADs) and that a majority reshuffle topologically associating domains (TADs). By applying phased RNA-seq, we observe an enrichment of genes showing allelic imbalanced expression (AIG) within 100 kb around the breakpoints. Interestingly, the AIGs hit by a breakpoint (19/22) display both up- and downregulation, thereby suggesting different mechanisms at play, such as gene disruption and rearrangements of regulatory information. However, the majority of interpretable genes located 200 kb around a breakpoint do not show significant expression changes. Thus, there is an overall robustness in the genome towards large-scale chromosome rearrangements.

Mohr-Tranebjærg syndrome is an X-linked syndrome characterized by sensorineural hearing impairment in childhood, followed by progressive neurodegeneration leading to a broad phenotypic spectrum. Genetically MTS is caused by pathogenic variants in the TIMM8A gene, including gene deletions and larger contiguous gene deletions. Some of the latter involve the neighboring gene BTK , resulting in agammaglobulinemia. By next‐generation mate‐pair sequencing we have mapped the chromosomal deletion breakpoints of one MTS case and three XLA-MTS cases and used breakpoint-spanning PCR to fine map the breakpoints by Sanger sequencing. Two of the XLA-MTS cases presented with large deletions (63.5 and 27.2 kb), and the junctional regions were characterized by long stretches of microhomology, indicating that the events have emerged through homologous recombination. Conversely, the MTS case exhibited a small 2 bp region of microhomology, and the regions were not characterized by extensive microhomology. The third XLA-MTS case had a more complex breakpoint, including a 59 bp inverted insertion, thus at least four breakpoints were involved in this event. In conclusion, mate-pair library generation combined with next-generation sequencing is an efficient method for breakpoint identification, also in regions characterized by repetitive elements.

HiC proximity-based sequencing has revealed that genomes are organized into megabase-sized topological associating domains (TADs) as units for cis-regulatory interactions. In a joint analysis across consortia studies led by the International Breakpoint Mapping Consortium (IBMC) and the Developmental Genome Anatomy Project (DGAP), we have used systematic mapping of 1,420 breakpoints from simple two-way balanced constitutional chromosomal rearrangements (BCRs) from both affected and healthy carriers to establish a first morbid map of this 3D-genome organization. Only ∼20% of the morbidity of BCRs can be explained by direct truncation of a known or plausible AD disease gene. Hence, dysregulatory long range position effects (LRPE) may be more common than previously assumed, and we define genomic features and ranked lists of TADs that are at risk for LRPE. Moreover, our data support that both breakpoints in simple two-way BCRs may frequently contribute to the associated morbidity. In a significant subset of BCRs, dual loss and gain of putative regulatory elements exchanges between two high-risk TADs (enhancer swapping) may lead to novel, unpredictable developmental disorders.