GCAT|Panel, a comprehensive structural variant haplotype map of the Iberian population from high-coverage whole-genome sequencing.

The combined analysis of haplotype panels with phenotype clinical cohorts is a common approach to explore the genetic architecture of human diseases. However, genetic studies are mainly based on single nucleotide variants (SNVs) and small insertions and deletions (indels). Here, we contribute to fill this gap by generating a dense haplotype map focused on the identification, characterization, and phasing of structural variants (SVs). By integrating multiple variant identification methods and Logistic Regression Models (LRMs), we present a catalogue of 35 431 441 variants, including 89 178 SVs (≥50 bp), 30 325 064 SNVs and 5 017 199 indels, across 785 Illumina high coverage (30x) whole-genomes from the Iberian GCAT Cohort, containing a median of 3.52M SNVs, 606 336 indels and 6393 SVs per individual. The haplotype panel is able to impute up to 14 360 728 SNVs/indels and 23 179 SVs, showing a 2.7-fold increase for SVs compared with available genetic variation panels. The value of this panel for SVs analysis is shown through an imputed rare Alu element located in a new locus associated with Mononeuritis of lower limb, a rare neuromuscular disease. This study represents the first deep characterization of genetic variation within the Iberian population and the first operational haplotype panel to systematically include the SVs into genome-wide genetic studies.

Genomic and epigenomic insights into the origin, pathogenesis, and clinical behavior of mantle cell lymphoma subtypes.

Mantle cell lymphoma (MCL) is a mature B-cell neoplasm initially driven by CCND1 rearrangement with 2 molecular subtypes, conventional MCL (cMCL) and leukemic non-nodal MCL (nnMCL), that differ in their clinicobiological behavior. To identify the genetic and epigenetic alterations determining this diversity, we used whole-genome (n = 61) and exome (n = 21) sequencing (74% cMCL, 26% nnMCL) combined with transcriptome and DNA methylation profiles in the context of 5 MCL reference epigenomes. We identified that open and active chromatin at the major translocation cluster locus might facilitate the t(11;14)(q13;32), which modifies the 3-dimensional structure of the involved regions. This translocation is mainly acquired in precursor B cells mediated by recombination-activating genes in both MCL subtypes, whereas in 8% of cases the translocation occurs in mature B cells mediated by activation-induced cytidine deaminase. We identified novel recurrent MCL drivers, including CDKN1B, SAMHD1, BCOR, SYNE1, HNRNPH1, SMARCB1, and DAZAP1. Complex structural alterations emerge as a relevant early oncogenic mechanism in MCL, targeting key driver genes. Breakage-fusion-bridge cycles and translocations activated oncogenes (BMI1, MIR17HG, TERT, MYC, and MYCN), generating gene amplifications and remodeling regulatory regions. cMCL carried significant higher numbers of structural variants, copy number alterations, and driver changes than nnMCL, with exclusive alterations of ATM in cMCL, whereas TP53 and TERT alterations were slightly enriched in nnMCL. Several drivers had prognostic impact, but only TP53 and MYC aberrations added value independently of genomic complexity. An increasing genomic complexity, together with the presence of breakage-fusion-bridge cycles and high DNA methylation changes related to the proliferative cell history, defines patients with different clinical evolution.

Identification of novel type 2 diabetes candidate genes involved in the crosstalk between the mitochondrial and the insulin signaling systems.

Type 2 Diabetes (T2D) is a highly prevalent chronic metabolic disease with strong co-morbidity with obesity and cardiovascular diseases. There is growing evidence supporting the notion that a crosstalk between mitochondria and the insulin signaling cascade could be involved in the etiology of T2D and insulin resistance. In this study we investigated the molecular basis of this crosstalk by using systems biology approaches. We combined, filtered, and interrogated different types of functional interaction data, such as direct protein-protein interactions, co-expression analyses, and metabolic and signaling dependencies. As a result, we constructed the mitochondria-insulin (MITIN) network, which highlights 286 genes as candidate functional linkers between these two systems. The results of internal gene expression analysis of three independent experimental models of mitochondria and insulin signaling perturbations further support the connecting roles of these genes. In addition, we further assessed whether these genes are involved in the etiology of T2D using the genome-wide association study meta-analysis from the DIAGRAM consortium, involving 8,130 T2D cases and 38,987 controls. We found modest enrichment of genes associated with T2D amongst our linker genes (p = 0.0549), including three already validated T2D SNPs and 15 additional SNPs, which, when combined, were collectively associated to increased fasting glucose levels according to MAGIC genome wide meta-analysis (p = 8.12×10(-5)). This study highlights the potential of combining systems biology, experimental, and genome-wide association data mining for identifying novel genes and related variants that increase vulnerability to complex diseases.

ReLA, a local alignment search tool for the identification of distal and proximal gene regulatory regions and their conserved transcription factor binding sites.

MOTIVATION: The prediction and annotation of the genomic regions involved in gene expression has been largely explored. Most of the energy has been devoted to the development of approaches that detect transcription start sites, leaving the identification of regulatory regions and their functional transcription factor binding sites (TFBSs) largely unexplored and with important quantitative and qualitative methodological gaps. RESULTS: We have developed ReLA (for REgulatory region Local Alignment tool), a unique tool optimized with the Smith-Waterman algorithm that allows local searches of conserved TFBS clusters and the detection of regulatory regions proximal to genes and enhancer regions. ReLA's performance shows specificities of 81 and 50% when tested on experimentally validated proximal regulatory regions and enhancers, respectively.

Mutational topography reflects clinical neuroblastoma heterogeneity.

Neuroblastoma is a pediatric solid tumor characterized by strong clinical heterogeneity. Although clinical risk-defining genomic alterations exist in neuroblastomas, the mutational processes involved in their generation remain largely unclear. By examining the topography and mutational signatures derived from all variant classes, we identified co-occurring mutational footprints, which we termed mutational scenarios. We demonstrate that clinical neuroblastoma heterogeneity is associated with differences in the mutational processes driving these scenarios, linking risk-defining pathognomonic variants to distinct molecular processes. Whereas high-risk MYCN-amplified neuroblastomas were characterized by signs of replication slippage and stress, homologous recombination-associated signatures defined high-risk non-MYCN-amplified patients. Non-high-risk neuroblastomas were marked by footprints of chromosome mis-segregation and TOP1 mutational activity. Furthermore, analysis of subclonal mutations uncovered differential activity of these processes through neuroblastoma evolution. Thus, clinical heterogeneity of neuroblastoma patients can be linked to differences in the mutational processes that are active in their tumors.

The impact of non-additive genetic associations on age-related complex diseases.

Genome-wide association studies (GWAS) are not fully comprehensive, as current strategies typically test only the additive model, exclude the X chromosome, and use only one reference panel for genotype imputation. We implement an extensive GWAS strategy, GUIDANCE, which improves genotype imputation by using multiple reference panels and includes the analysis of the X chromosome and non-additive models to test for association. We apply this methodology to 62,281 subjects across 22 age-related diseases and identify 94 genome-wide associated loci, including 26 previously unreported. Moreover, we observe that 27.7% of the 94 loci are missed if we use standard imputation strategies with a single reference panel, such as HRC, and only test the additive model. Among the new findings, we identify three novel low-frequency recessive variants with odds ratios larger than 4, which need at least a three-fold larger sample size to be detected under the additive model. This study highlights the benefits of applying innovative strategies to better uncover the genetic architecture of complex diseases.

TIGER: The gene expression regulatory variation landscape of human pancreatic islets.

Genome-wide association studies (GWASs) identified hundreds of signals associated with type 2 diabetes (T2D). To gain insight into their underlying molecular mechanisms, we have created the translational human pancreatic islet genotype tissue-expression resource (TIGER), aggregating >500 human islet genomic datasets from five cohorts in the Horizon 2020 consortium T2DSystems. We impute genotypes using four reference panels and meta-analyze cohorts to improve the coverage of expression quantitative trait loci (eQTL) and develop a method to combine allele-specific expression across samples (cASE). We identify >1 million islet eQTLs, 53 of which colocalize with T2D signals. Among them, a low-frequency allele that reduces T2D risk by half increases CCND2 expression. We identify eight cASE colocalizations, among which we found a T2D-associated SLC30A8 variant. We make all data available through the TIGER portal (http://tiger.bsc.es), which represents a comprehensive human islet genomic data resource to elucidate how genetic variation affects islet function and translates into therapeutic insight and precision medicine for T2D.

Author Correction: PGBD5 promotes site-specific oncogenic mutations in human tumors.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

The proliferative history shapes the DNA methylome of B-cell tumors and predicts clinical outcome.

We report a systematic analysis of the DNA methylation variability in 1,595 samples of normal cell subpopulations and 14 tumor subtypes spanning the entire human B-cell lineage. Differential methylation among tumor entities relates to differences in cellular origin and to de novo epigenetic alterations, which allowed us to build an accurate machine learning-based diagnostic algorithm. We identify extensive patient-specific methylation variability in silenced chromatin associated with the proliferative history of normal and neoplastic B cells. Mitotic activity generally leaves both hyper- and hypomethylation imprints, but some B-cell neoplasms preferentially gain or lose DNA methylation. Subsequently, we construct a DNA methylation-based mitotic clock called epiCMIT, whose lapse magnitude represents a strong independent prognostic variable in B-cell tumors and is associated with particular driver genetic alterations. Our findings reveal DNA methylation as a holistic tracer of B-cell tumor developmental history, with implications in the differential diagnosis and prediction of clinical outcome.

Enhancer hijacking determines extrachromosomal circular MYCN amplicon architecture in neuroblastoma.

MYCN amplification drives one in six cases of neuroblastoma. The supernumerary gene copies are commonly found on highly rearranged, extrachromosomal circular DNA (ecDNA). The exact amplicon structure has not been described thus far and the functional relevance of its rearrangements is unknown. Here, we analyze the MYCN amplicon structure using short-read and Nanopore sequencing and its chromatin landscape using ChIP-seq, ATAC-seq and Hi-C. This reveals two distinct classes of amplicons which explain the regulatory requirements for MYCN overexpression. The first class always co-amplifies a proximal enhancer driven by the noradrenergic core regulatory circuit (CRC). The second class of MYCN amplicons is characterized by high structural complexity, lacks key local enhancers, and instead contains distal chromosomal fragments harboring CRC-driven enhancers. Thus, ectopic enhancer hijacking can compensate for the loss of local gene regulatory elements and explains a large component of the structural diversity observed in MYCN amplification.

Publisher Correction: Extrachromosomal circular DNA drives oncogenic genome remodeling in neuroblastoma.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Extrachromosomal circular DNA drives oncogenic genome remodeling in neuroblastoma.

Extrachromosomal circularization of DNA is an important genomic feature in cancer. However, the structure, composition and genome-wide frequency of extrachromosomal circular DNA have not yet been profiled extensively. Here, we combine genomic and transcriptomic approaches to describe the landscape of extrachromosomal circular DNA in neuroblastoma, a tumor arising in childhood from primitive cells of the sympathetic nervous system. Our analysis identifies and characterizes a wide catalog of somatically acquired and undescribed extrachromosomal circular DNAs. Moreover, we find that extrachromosomal circular DNAs are an unanticipated major source of somatic rearrangements, contributing to oncogenic remodeling through chimeric circularization and reintegration of circular DNA into the linear genome. Cancer-causing lesions can emerge out of circle-derived rearrangements and are associated with adverse clinical outcome. It is highly probable that circle-derived rearrangements represent an ongoing mutagenic process. Thus, extrachromosomal circular DNAs represent a multihit mutagenic process, with important functional and clinical implications for the origins of genomic remodeling in cancer.

Pan-cancer analysis of whole genomes identifies driver rearrangements promoted by LINE-1 retrotransposition.

About half of all cancers have somatic integrations of retrotransposons. Here, to characterize their role in oncogenesis, we analyzed the patterns and mechanisms of somatic retrotransposition in 2,954 cancer genomes from 38 histological cancer subtypes within the framework of the Pan-Cancer Analysis of Whole Genomes (PCAWG) project. We identified 19,166 somatically acquired retrotransposition events, which affected 35% of samples and spanned a range of event types. Long interspersed nuclear element (LINE-1; L1 hereafter) insertions emerged as the first most frequent type of somatic structural variation in esophageal adenocarcinoma, and the second most frequent in head-and-neck and colorectal cancers. Aberrant L1 integrations can delete megabase-scale regions of a chromosome, which sometimes leads to the removal of tumor-suppressor genes, and can induce complex translocations and large-scale duplications. Somatic retrotranspositions can also initiate breakage-fusion-bridge cycles, leading to high-level amplification of oncogenes. These observations illuminate a relevant role of L1 retrotransposition in remodeling the cancer genome, with potential implications for the development of human tumors.

Insight into genetic predisposition to chronic lymphocytic leukemia from integrative epigenomics.

Genome-wide association studies have provided evidence for inherited genetic predisposition to chronic lymphocytic leukemia (CLL). To gain insight into the mechanisms underlying CLL risk we analyze chromatin accessibility, active regulatory elements marked by H3K27ac, and DNA methylation at 42 risk loci in up to 486 primary CLLs. We identify that risk loci are significantly enriched for active chromatin in CLL with evidence of being CLL-specific or differentially regulated in normal B-cell development. We then use in situ promoter capture Hi-C, in conjunction with gene expression data to reveal likely target genes of the risk loci. Candidate target genes are enriched for pathways related to B-cell development such as MYC and BCL2 signalling. At 14 loci the analysis highlights 63 variants as the probable functional basis of CLL risk. By integrating genetic and epigenetic information our analysis reveals novel insights into the relationship between inherited predisposition and the regulatory chromatin landscape of CLL.

Re-analysis of public genetic data reveals a rare X-chromosomal variant associated with type 2 diabetes.

The reanalysis of existing GWAS data represents a powerful and cost-effective opportunity to gain insights into the genetics of complex diseases. By reanalyzing publicly available type 2 diabetes (T2D) genome-wide association studies (GWAS) data for 70,127 subjects, we identify seven novel associated regions, five driven by common variants (LYPLAL1, NEUROG3, CAMKK2, ABO, and GIP genes), one by a low-frequency (EHMT2), and one driven by a rare variant in chromosome Xq23, rs146662057, associated with a twofold increased risk for T2D in males. rs146662057 is located within an active enhancer associated with the expression of Angiotensin II Receptor type 2 gene (AGTR2), a modulator of insulin sensitivity, and exhibits allelic specific activity in muscle cells. Beyond providing insights into the genetics and pathophysiology of T2D, these results also underscore the value of reanalyzing publicly available data using novel genetic resources and analytical approaches.

Publisher Correction: Re-analysis of public genetic data reveals a rare X-chromosomal variant associated with type 2 diabetes.

In the originally published version of this Article, the affiliation details for Santi González, Jian'an Luan and Claudia Langenberg were inadvertently omitted. Santi González should have been affiliated with 'Barcelona Supercomputing Center (BSC), Joint BSC-CRG-IRB Research Program in Computational Biology, 08034 Barcelona, Spain', and Jian'an Luan and Claudia Langenberg should have been affiliated with 'MRC Epidemiology Unit, University of Cambridge School of Clinical Medicine, Cambridge Biomedical Campus, Cambridge CB2 0QQ, UK'. Furthermore, the abstract contained an error in the SNP ID for the rare variant in chromosome Xq23, which was incorrectly given as rs146662057 and should have been rs146662075. These errors have now been corrected in both the PDF and HTML versions of the Article.

The reference epigenome and regulatory chromatin landscape of chronic lymphocytic leukemia.

Chronic lymphocytic leukemia (CLL) is a frequent hematological neoplasm in which underlying epigenetic alterations are only partially understood. Here, we analyze the reference epigenome of seven primary CLLs and the regulatory chromatin landscape of 107 primary cases in the context of normal B cell differentiation. We identify that the CLL chromatin landscape is largely influenced by distinct dynamics during normal B cell maturation. Beyond this, we define extensive catalogues of regulatory elements de novo reprogrammed in CLL as a whole and in its major clinico-biological subtypes classified by IGHV somatic hypermutation levels. We uncover that IGHV-unmutated CLLs harbor more active and open chromatin than IGHV-mutated cases. Furthermore, we show that de novo active regions in CLL are enriched for NFAT, FOX and TCF/LEF transcription factor family binding sites. Although most genetic alterations are not associated with consistent epigenetic profiles, CLLs with MYD88 mutations and trisomy 12 show distinct chromatin configurations. Furthermore, we observe that non-coding mutations in IGHV-mutated CLLs are enriched in H3K27ac-associated regulatory elements outside accessible chromatin. Overall, this study provides an integrative portrait of the CLL epigenome, identifies extensive networks of altered regulatory elements and sheds light on the relationship between the genetic and epigenetic architecture of the disease.

PGBD5 promotes site-specific oncogenic mutations in human tumors.

Genomic rearrangements are a hallmark of human cancers. Here, we identify the piggyBac transposable element derived 5 (PGBD5) gene as encoding an active DNA transposase expressed in the majority of childhood solid tumors, including lethal rhabdoid tumors. Using assembly-based whole-genome DNA sequencing, we found previously undefined genomic rearrangements in human rhabdoid tumors. These rearrangements involved PGBD5-specific signal (PSS) sequences at their breakpoints and recurrently inactivated tumor-suppressor genes. PGBD5 was physically associated with genomic PSS sequences that were also sufficient to mediate PGBD5-induced DNA rearrangements in rhabdoid tumor cells. Ectopic expression of PGBD5 in primary immortalized human cells was sufficient to promote cell transformation in vivo. This activity required specific catalytic residues in the PGBD5 transposase domain as well as end-joining DNA repair and induced structural rearrangements with PSS breakpoints. These results define PGBD5 as an oncogenic mutator and provide a plausible mechanism for site-specific DNA rearrangements in childhood and adult solid tumors.