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Emerging evidence links genes within human-specific segmental duplications (HSDs) to traits and diseases unique to our species. Strikingly, despite being nearly identical by sequence (>98.5%), paralogous HSD genes are differentially expressed across human cell and tissue types, though the underlying mechanisms have not been examined. We compared cross-tissue mRNA levels of 75 HSD genes from 30 families between humans and chimpanzees and found expression patterns consistent with relaxed selection on or neofunctionalization of derived paralogs. In general, ancestral paralogs exhibited greatest expression conservation with chimpanzee orthologs, though exceptions suggest certain derived paralogs may retain or supplant ancestral functions. Concordantly, analysis of long-read isoform sequencing data sets from diverse human tissues and cell lines found that about half of derived paralogs exhibited globally lower expression. To understand mechanisms underlying these differences, we leveraged data from human lymphoblastoid cell lines (LCLs) and found no relationship between paralogous expression divergence and post-transcriptional regulation, sequence divergence, or copy-number variation. Considering cis-regulation, we reanalyzed ENCODE data and recovered hundreds of previously unidentified candidate CREs in HSDs. We also generated large-insert ChIP-sequencing data for active chromatin features in an LCL to better distinguish paralogous regions. Some duplicated CREs were sufficient to drive differential reporter activity, suggesting they may contribute to divergent cis-regulation of paralogous genes. This work provides evidence that cis-regulatory divergence contributes to novel expression patterns of recent gene duplicates in humans. more...

Background Transposable element (TE) sequences are classified into families based on the reconstructed history of replication, and into subfamilies based on more fine-grained features that are often intended to capture family history. We evaluate the reliability of annotation with common subfamilies by assessing the extent to which subfamily annotation is reproducible in replicate copies created by segmental duplications in the human genome, and in homologous copies shared by human and chimpanzee. Results We find that standard methods annotate over 10% of replicates as belonging to different subfamilies, despite the fact that they are expected to be annotated as belonging to the same subfamily. Point mutations and homologous recombination appear to be responsible for some of this discordant annotation (particularly in the young Alu family), but are unlikely to fully explain the annotation unreliability. Conclusions The surprisingly high level of disagreement in subfamily annotation of homologous sequences highlights a need for further research into definition of TE subfamilies, methods for representing subfamily annotation confidence of TE instances, and approaches to better utilizing such nuanced annotation data in downstream analysis. more...

Genome-wide analysis methods, such as array comparative genomic hybridization (CGH) and whole-genome sequencing (WGS), have greatly advanced the identification of structural variants (SVs) in the human genome. However, even with standard high-throughput sequencing techniques, complex rearrangements with multiple breakpoints are often difficult to resolve, and predicting their effects on gene expression and phenotype remains a challenge. Here, we address these problems by using high-throughput chromosome conformation capture (Hi-C) generated from cultured cells of nine individuals with developmental disorders (DDs). Three individuals had previously been identified as harboring duplications at the SOX9 locus and six had been identified with translocations. Hi-C resolved the positions of the duplications and was instructive in interpreting their distinct pathogenic effects, including the formation of new topologically associating domains (neo-TADs). Hi-C was very sensitive in detecting translocations, and it revealed previously unrecognized complex rearrangements at the breakpoints. In several cases, we observed the formation of fused-TADs promoting ectopic enhancer-promoter interactions that were likely to be involved in the disease pathology. In summary, we show that Hi-C is a sensible method for the detection of complex SVs in a clinical setting. The results help interpret the possible pathogenic effects of the SVs in individuals with DDs. more...

The human genome contains hundreds of low-copy repeats (LCRs) that are challenging to analyze using short-read sequencing technologies due to extensive copy number variation and ambiguity in read mapping. Copy number and sequence variants in more than 150 duplicated genes that overlap LCRs have been implicated in monogenic and complex human diseases. We describe a computational tool, Parascopy, for estimating the aggregate and paralog-specific copy number of duplicated genes using whole-genome sequencing (WGS). Parascopy is an efficient method that jointly analyzes reads mapped to different repeat copies without the need for global realignment. It leverages multiple samples to mitigate sequencing bias and to identify reliable paralogous sequence variants (PSVs) that differentiate repeat copies. Analysis of WGS data for 2504 individuals from diverse populations showed that Parascopy is robust to sequencing bias, has higher accuracy compared to existing methods and enables prioritization of pathogenic copy number changes in duplicated genes. more...

Significance Extrachromosomal circular DNA (eccDNA) plays a role in human diseases such as cancer, but little is known about the impact of eccDNA in healthy human biology. Since eccDNA is a tiny fraction of nuclear DNA, artificial amplification has been employed to increase eccDNA amounts, resulting in the loss of native compositions. We developed an approach to enrich eccDNA populations at the native state (naïve small circular DNA, nscDNA) and investigated their origins in the human genome. We found that, in human sperm, the vast majority of nscDNA came from high-copy genomic regions, including the most variable regions between individuals. Because eccDNA can be incorporated back into chromosomes, eccDNA may promote human genetic variation., Extrachromosomal circular DNA (eccDNA) originates from linear chromosomal DNA in various human tissues under physiological and disease conditions. The genomic origins of eccDNA have largely been investigated using in vitro–amplified DNA. However, in vitro amplification obscures quantitative information by skewing the total population stoichiometry. In addition, the analyses have focused on eccDNA stemming from single-copy genomic regions, leaving eccDNA from multicopy regions unexamined. To address these issues, we isolated eccDNA without in vitro amplification (naïve small circular DNA, nscDNA) and assessed the populations quantitatively by integrated genomic, molecular, and cytogenetic approaches. nscDNA of up to tens of kilobases were successfully enriched by our approach and were predominantly derived from multicopy genomic regions including segmental duplications (SDs). SDs, which account for 5% of the human genome and are hotspots for copy number variations, were significantly overrepresented in sperm nscDNA, with three times more sequencing reads derived from SDs than from the entire single-copy regions. SDs were also overrepresented in mouse sperm nscDNA, which we estimated to comprise 0.2% of nuclear DNA. Considering that eccDNA can be integrated into chromosomes, germline-derived nscDNA may be a mediator of genome diversity. more...

Segmental duplications or low copy repeats (LCRs) constitute duplicated regions interspersed in the human genome, currently neglected in standard analyses due to their extreme complexity. Recent functional studies have indicated the potential of genes within LCRs in synaptogenesis, neuronal migration, and neocortical expansion in the human lineage. One of the regions with the highest proportion of duplicated sequence is the 22q11.2 locus, carrying eight LCRs (LCR22-A until LCR22-H), and rearrangements between them cause the 22q11.2 deletion syndrome. The LCR22-A block was recently reported to be hypervariable in the human population. It remains unknown whether this variability also exists in non-human primates, since research is strongly hampered by the presence of sequence gaps in the human and non-human primate reference genomes. To chart the LCR22 haplotypes and the associated inter- and intra-species variability, we de novo assembled the region in non-human primates by a combination of optical mapping techniques. A minimal and likely ancient haplotype is present in the chimpanzee, bonobo, and rhesus monkey without intra-species variation. In addition, the optical maps identified assembly errors and closed gaps in the orthologous chromosome 22 reference sequences. These findings indicate the LCR22 expansion to be unique to the human population, which might indicate involvement of the region in human evolution and adaptation. Those maps will enable LCR22-specific functional studies and investigate potential associations with the phenotypic variability in the 22q11.2 deletion syndrome. ispartof: FRONTIERS IN GENETICS vol:12 ispartof: location:Switzerland status: published more...

Segmental duplications (SDs) are a class of long, repetitive DNA elements whose paralogs share a high level of sequence similarity with each other. SDs mediate chromosomal rearrangements that lead to structural variation in the general population as well as genomic disorders associated with multiple congenital anomalies, including the 7q11.23 (Williams–Beuren Syndrome, WBS), 15q13.3, and 16p12.2 microdeletion syndromes. Population-level characterization of SDs has generally been lacking because most techniques used for analyzing these complex regions are both labor and cost intensive. In this study, we have used a high-throughput technique to genotype complex structural variation with a single molecule, long-range optical mapping approach. We characterized SDs and identified novel structural variants (SVs) at 7q11.23, 15q13.3, and 16p12.2 using optical mapping data from 154 phenotypically normal individuals from 26 populations comprising five super-populations. We detected several novel SVs for each locus, some of which had significantly different prevalence between populations. Additionally, we localized the microdeletion breakpoints to specific paralogous duplicons located within complex SDs in two patients with WBS, one patient with 15q13.3, and one patient with 16p12.2 microdeletion syndromes. The population-level data presented here highlights the extreme diversity of large and complex SVs within SD-containing regions. The approach we outline will greatly facilitate the investigation of the role of inter-SD structural variation as a driver of chromosomal rearrangements and genomic disorders. more...

The ability to characterize repetitive regions of the human genome is limited by the read lengths of short-read sequencing technologies. Although long-read sequencing technologies such as Pacific Biosciences (PacBio) and Oxford Nanopore Technologies can potentially overcome this limitation, long segmental duplications with high sequence identity pose challenges for long-read mapping. We describe a probabilistic method, DuploMap, designed to improve the accuracy of long-read mapping in segmental duplications. It analyzes reads mapped to segmental duplications using existing long-read aligners and leverages paralogous sequence variants (PSVs)—sequence differences between paralogous sequences—to distinguish between multiple alignment locations. On simulated datasets, DuploMap increased the percentage of correctly mapped reads with high confidence for multiple long-read aligners including Minimap2 (74.3–90.6%) and BLASR (82.9–90.7%) while maintaining high precision. Across multiple whole-genome long-read datasets, DuploMap aligned an additional 8–21% of the reads in segmental duplications with high confidence relative to Minimap2. Using DuploMap-aligned PacBio circular consensus sequencing reads, an additional 8.9 Mb of DNA sequence was mappable, variant calling achieved a higher F1 score and 14 713 additional variants supported by linked-read data were identified. Finally, we demonstrate that a significant fraction of PSVs in segmental duplications overlaps with variants and adversely impacts short-read variant calling. more...

Ever since the availability of genomes from Neanderthals, Denisovans, and ancient humans, the field of evolutionary genomics has been searching for protein-coding variants that may hold clues to how our species evolved over the last ∼600,000 years. In this study, we identify such variants in the human-specific NOTCH2NL gene family, which were recently identified as possible contributors to the evolutionary expansion of the human brain. We find evidence for the existence of unique protein-coding NOTCH2NL variants in Neanderthals and Denisovans which could affect their ability to activate Notch signaling. Furthermore, in the Neanderthal and Denisovan genomes, we find unusual NOTCH2NL configurations, not found in any of the modern human genomes analyzed. Finally, genetic analysis of archaic and modern humans reveals ongoing adaptive evolution of modern human NOTCH2NL genes, identifying three structural variants acting complementary to drive our genome to produce a lower dosage of NOTCH2NL protein. Because copy-number variations of the 1q21.1 locus, encompassing NOTCH2NL genes, are associated with severe neurological disorders, this seemingly contradicting drive toward low levels of NOTCH2NL protein indicates that the optimal dosage of NOTCH2NL may have not yet been settled in the human population. more...

The 22q11.2 deletion syndrome (22q11.2DS) results from non-allelic homologous recombination between low-copy repeats termed LCR22. About 60%-70% of individuals with the typical 3 megabase (Mb) deletion from LCR22A-D have congenital heart disease, mostly of the conotruncal type (CTD), whereas others have normal cardiac anatomy. In this study, we tested whether variants in the hemizygous LCR22A-D region are associated with risk for CTDs on the basis of the sequence of the 22q11.2 region from 1,053 22q11.2DS individuals. We found a significant association (FDR p < 0.05) of the CTD subset with 62 common variants in a single linkage disequilibrium (LD) block in a 350 kb interval harboring CRKL. A total of 45 of the 62 variants were associated with increased risk for CTDs (odds ratio [OR) ranges: 1.64-4.75). Associations of four variants were replicated in a meta-analysis of three genome-wide association studies of CTDs in affected individuals without 22q11.2DS. One of the replicated variants, rs178252, is located in an open chromatin region and resides in the double-elite enhancer, GH22J020947, that is predicted to regulate CRKL (CRK-like proto-oncogene, cytoplasmic adaptor) expression. Approximately 23% of patients with nested LCR22C-D deletions have CTDs, and inactivation of Crkl in mice causes CTDs, thus implicating this gene as a modifier. Rs178252 and rs6004160 are expression quantitative trait loci (eQTLs) of CRKL. Furthermore, set-based tests identified an enhancer that is predicted to target CRKL and is significantly associated with CTD risk (GH22J020946, sequence kernal association test (SKAT) p = 7.21 × 10-5) in the 22q11.2DS cohort. These findings suggest that variance in CTD penetrance in the 22q11.2DS population can be explained in part by variants affecting CRKL expression. ispartof: AMERICAN JOURNAL OF HUMAN GENETICS vol:106 issue:1 pages:26-40 ispartof: location:United States status: published more...

The sequence and assembly of human genomes using long-read sequencing technologies has revolutionized our understanding of structural variation and genome organization. We compared the accuracy, continuity, and gene annotation of genome assemblies generated from either high-fidelity (HiFi) or continuous long-read (CLR) datasets from the same complete hydatidiform mole human genome. We find that the HiFi sequence data assemble an additional 10% of duplicated regions and more accurately represent the structure of tandem repeats, as validated with orthogonal analyses. As a result, an additional 5 Mbp of pericentromeric sequences are recovered in the HiFi assembly, resulting in a 2.5-fold increase in the NG50 within 1 Mbp of the centromere (HiFi 480.6 kbp, CLR 191.5 kbp). Additionally, the HiFi genome assembly was generated in significantly less time with fewer computational resources than the CLR assembly. Although the HiFi assembly has significantly improved continuity and accuracy in many complex regions of the genome, it still falls short of the assembly of centromeric DNA and the largest regions of segmental duplication using existing assemblers. Despite these shortcomings, our results suggest that HiFi may be the most effective stand-alone technology for de novo assembly of human genomes. more...

Motivation Several algorithms have been developed that use high-throughput sequencing technology to characterize structural variations (SVs). Most of the existing approaches focus on detecting relatively simple types of SVs such as insertions, deletions and short inversions. In fact, complex SVs are of crucial importance and several have been associated with genomic disorders. To better understand the contribution of complex SVs to human disease, we need new algorithms to accurately discover and genotype such variants. Additionally, due to similar sequencing signatures, inverted duplications or gene conversion events that include inverted segmental duplications are often characterized as simple inversions, likewise, duplications and gene conversions in direct orientation may be called as simple deletions. Therefore, there is still a need for accurate algorithms to fully characterize complex SVs and thus improve calling accuracy of more simple variants. Results We developed novel algorithms to accurately characterize tandem, direct and inverted interspersed segmental duplications using short read whole genome sequencing datasets. We integrated these methods to our TARDIS tool, which is now capable of detecting various types of SVs using multiple sequence signatures such as read pair, read depth and split read. We evaluated the prediction performance of our algorithms through several experiments using both simulated and real datasets. In the simulation experiments, using a 30× coverage TARDIS achieved 96% sensitivity with only 4% false discovery rate. For experiments that involve real data, we used two haploid genomes (CHM1 and CHM13) and one human genome (NA12878) from the Illumina Platinum Genomes set. Comparison of our results with orthogonal PacBio call sets from the same genomes revealed higher accuracy for TARDIS than state-of-the-art methods. Furthermore, we showed a surprisingly low false discovery rate of our approach for discovery of tandem, direct and inverted interspersed segmental duplications prediction on CHM1 ( Availability and implementation TARDIS source code is available at https://github.com/BilkentCompGen/tardis, and a corresponding Docker image is available at https://hub.docker.com/r/alkanlab/tardis/. Supplementary information Supplementary data are available at Bioinformatics online. more...

Large structural variants (SVs) in the human genome are difficult to detect and study by conventional sequencing technologies. With long-range genome analysis platforms, such as optical mapping, one can identify large SVs (>2 kb) across the genome in one experiment. Analyzing optical genome maps of 154 individuals from the 26 populations sequenced in the 1000 Genomes Project, we find that phylogenetic population patterns of large SVs are similar to those of single nucleotide variations in 86% of the human genome, while ~2% of the genome has high structural complexity. We are able to characterize SVs in many intractable regions of the genome, including segmental duplications and subtelomeric, pericentromeric, and acrocentric areas. In addition, we discover ~60 Mb of non-redundant genome content missing in the reference genome sequence assembly. Our results highlight the need for a comprehensive set of alternate haplotypes from different populations to represent SV patterns in the genome., Large structural variants (SV) are understudied in human genetics research because of the difficulty to detect them in the routinely generated short-read sequencing data. Here, the authors generate optical genome maps of 154 individuals from 26 populations that allow comprehensive examination of large SVs. more...

Identifying genomic features that differ between individuals and cells can help uncover the functional variants that drive phenotypes and disease susceptibilities. For this, single-cell studies are paramount, as it becomes increasingly clear that the contribution of rare but functional cellular subpopulations is important for disease prognosis, management, and progression. Until now, studying these associations has been challenged by our inability to map structural rearrangements accurately and comprehensively. To overcome this, we coupled single-cell sequencing of DNA template strands (Strand-seq) with custom analysis software to rapidly discover, map, and genotype genomic rearrangements at high resolution. This allowed us to explore the distribution and frequency of inversions in a heterogeneous cell population, identify several polymorphic domains in complex regions of the genome, and locate rare alleles in the reference assembly. We then mapped the entire genomic complement of inversions within two unrelated individuals to characterize their distinct inversion profiles and built a nonredundant global reference of structural rearrangements in the human genome. The work described here provides a powerful new framework to study structural variation and genomic heterogeneity in single-cell samples, whether from individuals for population studies or tissue types for biomarker discovery. more...

With the aid of novel and powerful molecular biology techniques, recent years have witnessed a dramatic increase in the number of studies reporting the involvement of complex structural variants in several genomic disorders. In fact, with the discovery of Copy Number Variants (CNVs) and other forms of unbalanced structural variation, much attention has been directed to the detection and characterization of such rearrangements, as well as the identification of the mechanisms involved in their formation. However, it has long been appreciated that chromosomes can undergo other forms of structural changes - balanced rearrangements - that do not involve quantitative variation of genetic material. Indeed, a particular subtype of balanced rearrangement - inversions - was recently found to be far more common than had been predicted from traditional cytogenetics. Chromosomal inversions alter the orientation of a specific genomic sequence and, unless involving breaks in coding or regulatory regions (and, disregarding complex trans effects, in their close vicinity), appear to be phenotypically silent. Such a surprising finding, which is difficult to reconcile with the classical interpretation of inversions as a mechanism causing subfertility (and ultimately reproductive isolation), motivated a new series of theoretical and empirical studies dedicated to understand their role in human genome evolution and to explore their possible association to complex genetic disorders. With this review, we attempt to describe the latest methodological improvements to inversions detection at a genome wide level, while exploring some of the possible implications of inversion rearrangements on the evolution of the human genome. more...

Copy number variation represents a major source of genetic divergence, yet the evolutionary dynamics of genic copy number variation in natural populations during differentiation and adaptation remain unclear. We applied a read depth approach to genome resequencing data to detect copy number variants (CNVs) ≥1 kb in wild-caught mice belonging to four populations of Mus musculus domesticus. We complemented the bioinformatics analyses with experimental validation using droplet digital PCR. The specific focus of our analysis is CNVs that include complete genes, as these CNVs could be expected to contribute most directly to evolutionary divergence. In total, 1863 transcription units appear to be completely encompassed within CNVs in at least one individual when compared to the reference assembly. Further, 179 of these CNVs show population-specific copy number differences, and 325 are subject to complete deletion in multiple individuals. Among the most copy-number variable genes are three highly conserved genes that encode the splicing factor CWC22, the spindle protein SFI1, and the Holliday junction recognition protein HJURP. These genes exhibit population-specific expansion patterns that suggest involvement in local adaptations. We found that genes that overlap with large segmental duplications are generally more copy-number variable. These genes encode proteins that are relevant for environmental and behavioral interactions, such as vomeronasal and olfactory receptors, as well as major urinary proteins and several proteins of unknown function. The overall analysis shows that genic CNVs contribute more to population differentiation in mice than in humans and may promote and speed up population divergence. more...

A previously detected copy number polymorphism (Ep CNP) in patients affected with neuroectodermal tumors led us to investigate its frequency and length in the normal population. For this purpose, a program called Sequence Allocator was developed and applied for the construction of an array that consisted of unique and duplicated fragments, allowing the assessment of copy number variation within regions of segmental duplications. The average resolution of this array was 11 kb and we determined the size of the Ep CNP to be 290 kb. Analysis of normal controls identified 7.7 and 7.1% gains in peripheral blood and lymphoblastoid cell line (LCL) DNA, respectively, while deletions were found only in the LCL group (7.1%). This array platform allows the detection of DNA copy number variation within regions of pronounced genomic complexity, which constitutes an improvement over available technologies. more...

Recurrent deletions of chromosome 15q13.3 associate with intellectual disability, schizophrenia, autism and epilepsy. To gain insight into the instability of this region, we sequenced it in affected individuals, normal individuals and nonhuman primates. We discovered five structural configurations of the human chromosome 15q13.3 region ranging in size from 2 to 3 Mb. These configurations arose recently (∼0.5-0.9 million years ago) as a result of human-specific expansions of segmental duplications and two independent inversion events. All inversion breakpoints map near GOLGA8 core duplicons-a ∼14-kb primate-specific chromosome 15 repeat that became organized into larger palindromic structures. GOLGA8-flanked palindromes also demarcate the breakpoints of recurrent 15q13.3 microdeletions, the expansion of chromosome 15 segmental duplications in the human lineage and independent structural changes in apes. The significant clustering (P = 0.002) of breakpoints provides mechanistic evidence for the role of this core duplicon and its palindromic architecture in promoting the evolutionary and disease-related instability of chromosome 15. more...

Background Although the reference human genome sequence was declared finished in 2003, some regions of the genome remain incomplete due to their complex architecture. One such region, 1q21.1-q21.2, is of increasing interest due to its relevance to human disease and evolution. Elucidation of the exact variants behind these associations has been hampered by the repetitive nature of the region and its incomplete assembly. This region also contains 238 of the 270 human DUF1220 protein domains, which are implicated in human brain evolution and neurodevelopment. Additionally, examinations of this protein domain have been challenging due to the incomplete 1q21 build. To address these problems, a single-haplotype hydatidiform mole BAC library (CHORI-17) was used to produce the first complete sequence of the 1q21.1-q21.2 region. Results We found and addressed several inaccuracies in the GRCh37sequence of the 1q21 region on large and small scales, including genomic rearrangements and inversions, and incorrect gene copy number estimates and assemblies. The DUF1220-encoding NBPF genes required the most corrections, with 3 genes removed, 2 genes reassigned to the 1p11.2 region, 8 genes requiring assembly corrections for DUF1220 domains (~91 DUF1220 domains were misassigned), and multiple instances of nucleotide changes that reassigned the domain to a different DUF1220 subtype. These corrections resulted in an overall increase in DUF1220 copy number, yielding a haploid total of 289 copies. Approximately 20 of these new DUF1220 copies were the result of a segmental duplication from 1q21.2 to 1p11.2 that included two NBPF genes. Interestingly, this duplication may have been the catalyst for the evolutionarily important human lineage-specific chromosome 1 pericentric inversion. Conclusions Through the hydatidiform mole genome sequencing effort, the 1q21.1-q21.2 region is complete and misassemblies involving inter- and intra-region duplications have been resolved. The availability of this single haploid sequence path will aid in the investigation of many genetic diseases linked to 1q21, including several associated with DUF1220 copy number variations. Finally, the corrected sequence identified a recent segmental duplication that added 20 additional DUF1220 copies to the human genome, and may have facilitated the chromosome 1 pericentric inversion that is among the most notable human-specific genomic landmarks. more...

The short arms of the five acrocentric human chromosomes harbor sequences that direct the assembly and function of the nucleolus, one of the key functional domains of the nucleus, yet they are absent from the current human genome assembly. Here we describe the genomic architecture of these human nucleolar organizers. Sequences distal and proximal to ribosomal gene arrays are conserved among the acrocentric chromosomes, suggesting they are sites of frequent recombination. Although previously believed to be heterochromatic, characterization of these two flanking regions reveals that they share a complex genomic architecture similar to other euchromatic regions of the genome, but they have distinct genomic characteristics. Proximal sequences are almost entirely segmentally duplicated, similar to the regions bordering centromeres. In contrast, the distal sequence is predominantly unique to the acrocentric short arms and is dominated by a very large inverted repeat. We show that the distal element is localized to the periphery of the nucleolus, where it appears to anchor the ribosomal gene repeats. This, combined with its complex chromatin structure and transcriptional activity, suggests that this region is involved in nucleolar organization. Our results provide a platform for investigating the role of NORs in nucleolar formation and function, and open the door for determining the role of these regions in the well-known empirical association of nucleoli with pathology. more...

Contains fulltext : 80657.pdf (Publisher’s version ) (Closed access) BACKGROUND: Recurrent 15q13.3 microdeletions were recently identified with identical proximal (BP4) and distal (BP5) breakpoints and associated with mild to moderate mental retardation and epilepsy. METHODS: To assess further the clinical implications of this novel 15q13.3 microdeletion syndrome, 18 new probands with a deletion were molecularly and clinically characterised. In addition, we evaluated the characteristics of a family with a more proximal deletion between BP3 and BP4. Finally, four patients with a duplication in the BP3-BP4-BP5 region were included in this study to ascertain the clinical significance of duplications in this region. RESULTS: The 15q13.3 microdeletion in our series was associated with a highly variable intra- and inter-familial phenotype. At least 11 of the 18 deletions identified were inherited. Moreover, 7 of 10 siblings from four different families also had this deletion: one had a mild developmental delay, four had only learning problems during childhood, but functioned well in daily life as adults, whereas the other two had no learning problems at all. In contrast to previous findings, seizures were not a common feature in our series (only 2 of 17 living probands). Three patients with deletions had cardiac defects and deletion of the KLF13 gene, located in the critical region, may contribute to these abnormalities. The limited data from the single family with the more proximal BP3-BP4 deletion suggest this deletion may have little clinical significance. Patients with duplications of the BP3-BP4-BP5 region did not share a recognisable phenotype, but psychiatric disease was noted in 2 of 4 patients. CONCLUSIONS: Overall, our findings broaden the phenotypic spectrum associated with 15q13.3 deletions and suggest that, in some individuals, deletion of 15q13.3 is not sufficient to cause disease. The existence of microdeletion syndromes, associated with an unpredictable and variable phenotypic outcome, will pose the clinician with diagnostic difficulties and challenge the commonly used paradigm in the diagnostic setting that aberrations inherited from a phenotypically normal parent are usually without clinical consequences. more...

The presence of chromosome-specific low-copy repeats (LCRs) predisposes chromosome 22 to deletions and duplications. The current diagnostic procedure for detecting aberrations at 22q11.2 is chromosomal analysis coupled with fluorescence in situ hybridization (FISH) or PCR-based multiplex ligation dependent probe amplification (MLPA). However, there are copy number variations (CNVs) in 22q11.2 that are only detected by high-resolution platforms such as array comparative genomic hybridization (aCGH). We report on development of a high-definition MLPA (MLPA-HD) 22q11 kit that detects copy number changes at 37 loci on the long arm of chromosome 22. These include the 3-Mb region commonly deleted in DiGeorge/velocardiofacial syndrome (DGS/VCFS), the cat eye syndrome (CES) region, and more distal regions in 22q11 that have recently been shown to be deleted. We have used this MLPA-HD probe set to analyze 363 previously well, characterized samples with a variety of different rearrangements at 22q11 and demonstrate that it can detect copy number alterations with high sensitivity and specificity. In addition to detection of the common recurrent deletions associated with DGS/VCFS, variant and novel chromosome 22 aberrations have been detected. These include duplications within as well as deletions distal to this region. Further, the MLPA-HD detects deletion endpoint differences between patients with the common 3-Mb deletion. The MLPA-HD kit is proposed as a cost effective alternative to the currently available detection methods for individuals with features of the 22q11 aberrations. In patients with the relevant phenotypic characteristics, this MLPA-HD probe set could replace FISH for the clinical diagnosis of 22q11.2 deletions and duplications. more...

PUBLISHED, Background Duplications and deletions in the human genome can cause disease or predispose persons to disease. Advances in technologies to detect these changes allow for the routine identification of submicroscopic imbalances in large numbers of patients. Methods We tested for the presence of microdeletions and microduplications at a specific region of chromosome 1q21.1 in two groups of patients with unexplained mental retardation, autism, or congenital anomalies and in unaffected persons. Results We identified 25 persons with a recurrent 1.35-Mb deletion within 1q21.1 from screening 5218 patients. The microdeletions had arisen de novo in eight patients, were inherited from a mildly affected parent in three patients, were inherited from an apparently unaffected parent in six patients, and were of unknown inheritance in eight patients. The deletion was absent in a series of 4737 control persons (P=1.1x10?7). We found considerable variability in the level of phenotypic expression of the microdeletion; phenotypes included mild-to-moderate mental retardation, microcephaly, cardiac abnormalities, and cataracts. The reciprocal duplication was enriched in nine children with mental retardation or autism spectrum disorder and other variable features (P=0.02). We identified three deletions and three duplications of the 1q21.1 region in an independent sample of 788 patients with mental retardation and congenital anomalies. Conclusions We have identified recurrent molecular lesions that elude syndromic classification and whose disease manifestations must be considered in a broader context of development as opposed to being assigned to a specific disease. Clinical diagnosis in patients with these lesions may be most readily achieved on the basis of genotype rather than phenotype., Supported in part by grants from the National Institutes of Health (NIH) (HD043569, to Dr. Eichler), the South Carolina Department of Disabilities and Special Needs (to Drs. Skinner, Stevenson, and Schwartz), the Wellcome Trust (061183, to Dr. Tassabehji), the Andre & Cyprien Foundation and the University Hospitals of Geneva (to Drs. Antonarakis, Bena, and Gallati), and the European Union (EU) (project 219250, to Dr. Sharp; AnEUploidy project 037627, to Drs. Leeuw, Armengol, Antonarakis, Estivill, Veltman, and de Vries). The Irish Autism Study was funded by the Wellcome Trust and the Health Research Board (a grant to Drs. Gallagher and Gill). Dr. Poot was supported by a grant from the Dutch Foundation for Brain Research (Hersenstichting grant 2008(1) 34); Drs. Regan and Knight, by the Oxford Partnership Comprehensive Biomedical Research Centre; Dr. Willatt, by the Cambridge Biomedical Research Centre, with funding from the United Kingdom Department of Health's National Institute for Health Research Biomedical Research Centres funding scheme; Drs. Huang and Maloney, as part of the National Genetics Reference Laboratory (Wessex) by the United Kingdom Department of Health; Ms. Buysse, as a research assistant of the Research Foundation?Flanders (FWO?Vlaanderen); and Dr. Eichler, as an investigator of the Howard Hughes Medical Institute. more...

Mouse and rat genomic sequences permit us to obtain a global view of evolutionary rearrangements that have occurred between the two species and to define hallmarks that might underlie these events. We present a comparative study of the sequence assemblies of mouse and rat genomes and report an enrichment of rodent-specific segmental duplications in regions where synteny is not preserved. We show that segmental duplications present higher rates of molecular evolution and that genes in rearranged regions have evolved faster than those located elsewhere. Previous studies have shown that synteny breakpoints between the mouse and the human genomes are enriched in human segmental duplications, suggesting a causative connection between such structures and evolutionary rearrangements. Our work provides further evidence to support the role of segmental duplications in chromosomal rearrangements in the evolution of the architecture of mammalian chromosomes and in the speciation processes that separate the mouse and the rat. more...

An intron is an extended genomic feature whose function requires multiple constrained positions—donor and acceptor splice sites, a branch point, a polypyrimidine tract and suitable splicing enhancers—that may be distributed over hundreds or thousands of nucleotides. New introns are therefore unlikely to emerge by incremental accumulation of functional sub-elements. Here we demonstrate that a functional intron can be created de novo in a single step by a segmental genomic duplication. This experiment recapitulates in vivo the birth of an intron that arose in the ancestral jawed vertebrate lineage nearly half-a-billion years ago., The appearance of a new intron that splits an exon without disrupting the corresponding peptide sequence is a rare event in vertebrate genomes. Hellsten et al. demonstrate that, under certain circumstances, a functional intron can be produced in a single step by segmental genomic duplication. more...