cDNA-detector: detection and removal of cDNA contamination in DNA sequencing libraries

BACKGROUND: Exogenous cDNA introduced into an experimental system, either intentionally or accidentally, can appear as added read coverage over that gene in next-generation sequencing libraries derived from this system. If not properly recognized and managed, this cross-contamination with exogenous signal can lead to incorrect interpretation of research results. Yet, this problem is not routinely addressed in current sequence processing pipelines. RESULTS: We present cDNA-detector, a computational tool to identify and remove exogenous cDNA contamination in DNA sequencing experiments. We demonstrate that cDNA-detector can identify cDNAs quickly and accurately from alignment files. A source inference step attempts to separate endogenous cDNAs (retrocopied genes) from potential cloned, exogenous cDNAs. cDNA-detector provides a mechanism to decontaminate the alignment from detected cDNAs. Simulation studies show that cDNA-detector is highly sensitive and specific, outperforming existing tools. We apply cDNA-detector to several highly-cited public databases (TCGA, ENCODE, NCBI SRA) and show that contaminant genes appear in sequencing experiments where they lead to incorrect coverage peak calls. CONCLUSIONS: cDNA-detector is a user-friendly and accurate tool to detect and remove cDNA detection in NGS libraries. This two-step design reduces the risk of true variant removal since it allows for manual review of candidates. We find that contamination with intentionally and accidentally introduced cDNAs is an underappreciated problem even in widely-used consortium datasets, where it can lead to spurious results. Our findings highlight the importance of sensitive detection and removal of contaminant cDNA from NGS libraries before downstream analysis

EWS-FLI1 Utilizes Divergent Chromatin Remodeling Mechanisms to Directly Activate or Repress Enhancer Elements in Ewing Sarcoma

SummaryThe aberrant transcription factor EWS-FLI1 drives Ewing sarcoma, but its molecular function is not completely understood. We find that EWS-FLI1 reprograms gene regulatory circuits in Ewing sarcoma by directly inducing or repressing enhancers. At GGAA repeat elements, which lack evolutionary conservation and regulatory potential in other cell types, EWS-FLI1 multimers induce chromatin opening and create de novo enhancers that physically interact with target promoters. Conversely, EWS-FLI1 inactivates conserved enhancers containing canonical ETS motifs by displacing wild-type ETS transcription factors. These divergent chromatin-remodeling patterns repress tumor suppressors and mesenchymal lineage regulators while activating oncogenes and potential therapeutic targets, such as the kinase VRK1. Our findings demonstrate how EWS-FLI1 establishes an oncogenic regulatory program governing both tumor survival and differentiation

Wilms Tumor Chromatin Profiles Highlight Stem Cell Properties and a Renal Developmental Network

Wilms tumor is the most common pediatric kidney cancer. To identify transcriptional and epigenetic mechanisms that drive this disease, we compared genome-wide chromatin profiles of Wilms tumors, embryonic stem cells (ESCs), and normal kidney. Wilms tumors prominently exhibit large active chromatin domains previously observed in ESCs. In the cancer, these domains frequently correspond to genes that are critical for kidney development and expressed in the renal stem cell compartment. Wilms cells also express “embryonic” chromatin regulators and maintain stem cell-like p16 silencing. Finally, Wilms and ESCs both exhibit “bivalent” chromatin modifications at silent promoters that may be poised for activation. In Wilms tumor, bivalent promoters correlate to genes expressed in specific kidney compartments and point to a kidney-specific differentiation program arrested at an early-progenitor stage. We suggest that Wilms cells share a transcriptional and epigenetic landscape with a normal renal stem cell, which is inherently susceptible to transformation and may represent a cell of origin for this disease

Evolution of pathogenicity and sexual reproduction in eight Candida genomes

Candida species are the most common cause of opportunistic fungal infection worldwide. Here we report the genome sequences of six Candida species and compare these and related pathogens and non-pathogens. There are significant expansions of cell wall, secreted and transporter gene families in pathogenic species, suggesting adaptations associated with virulence. Large genomic tracts are homozygous in three diploid species, possibly resulting from recent recombination events. Surprisingly, key components of the mating and meiosis pathways are missing from several species. These include major differences at the mating-type loci (MTL); Lodderomyces elongisporus lacks MTL, and components of the a1/2 cell identity determinant were lost in other species, raising questions about how mating and cell types are controlled. Analysis of the CUG leucine-to-serine genetic-code change reveals that 99% of ancestral CUG codons were erased and new ones arose elsewhere. Lastly, we revise the Candida albicans gene catalogue, identifying many new genes.publishe

A deep learning system accurately classifies primary and metastatic cancers using passenger mutation patterns.

In cancer, the primary tumour's organ of origin and histopathology are the strongest determinants of its clinical behaviour, but in 3% of cases a patient presents with a metastatic tumour and no obvious primary. Here, as part of the ICGC/TCGA Pan-Cancer Analysis of Whole Genomes (PCAWG) Consortium, we train a deep learning classifier to predict cancer type based on patterns of somatic passenger mutations detected in whole genome sequencing (WGS) of 2606 tumours representing 24 common cancer types produced by the PCAWG Consortium. Our classifier achieves an accuracy of 91% on held-out tumor samples and 88% and 83% respectively on independent primary and metastatic samples, roughly double the accuracy of trained pathologists when presented with a metastatic tumour without knowledge of the primary. Surprisingly, adding information on driver mutations reduced accuracy. Our results have clinical applicability, underscore how patterns of somatic passenger mutations encode the state of the cell of origin, and can inform future strategies to detect the source of circulating tumour DNA

Sex differences in oncogenic mutational processes

Sex differences have been observed in multiple facets of cancer epidemiology, treatment and biology, and in most cancers outside the sex organs. Efforts to link these clinical differences to specific molecular features have focused on somatic mutations within the coding regions of the genome. Here we report a pan-cancer analysis of sex differences in whole genomes of 1983 tumours of 28 subtypes as part of the ICGC/TCGA Pan-Cancer Analysis of Whole Genomes (PCAWG) Consortium. We both confirm the results of exome studies, and also uncover previously undescribed sex differences. These include sex-biases in coding and non-coding cancer drivers, mutation prevalence and strikingly, in mutational signatures related to underlying mutational processes. These results underline the pervasiveness of molecular sex differences and strengthen the call for increased consideration of sex in molecular cancer research

Reconstructing and Reprogramming the Tumor-Propagating Potential of Glioblastoma Stem-like Cells

Developmental fate decisions are dictated by master transcription factors (TFs) that interact with cis-regulatory elements to direct transcriptional programs. Certain malignant tumors may also depend on cellular hierarchies reminiscent of normal development but superimposed on underlying genetic aberrations. In glioblastoma (GBM), a subset of stem-like tumor-propagating cells (TPCs) appears to drive tumor progression and underlie therapeutic resistance, yet remain poorly understood. Here, we identify a core set of neurodevelopmental TFs (POU3F2, SOX2, SALL2, OLIG2) essential for GBM propagation. These TFs coordinately bind and activate TPC-specific regulatory elements, and are sufficient to fully reprogram differentiated GBM cells to ‘induced’ TPCs, recapitulating the epigenetic landscape and phenotype of native TPCs. We reconstruct a network model that highlights critical interactions and identifies novel therapeutic targets for eliminating TPCs. Our study establishes the epigenetic basis of a developmental hierarchy in GBM, provides detailed insight into underlying gene regulatory programs, and suggests attendant therapeutic strategies

Pan-cancer analysis of whole genomes

Cancer is driven by genetic change, and the advent of massively parallel sequencing has enabled systematic documentation of this variation at the whole-genome scale(1-3). Here we report the integrative analysis of 2,658 whole-cancer genomes and their matching normal tissues across 38 tumour types from the Pan-Cancer Analysis of Whole Genomes (PCAWG) Consortium of the International Cancer Genome Consortium (ICGC) and The Cancer Genome Atlas (TCGA). We describe the generation of the PCAWG resource, facilitated by international data sharing using compute clouds. On average, cancer genomes contained 4-5 driver mutations when combining coding and non-coding genomic elements; however, in around 5% of cases no drivers were identified, suggesting that cancer driver discovery is not yet complete. Chromothripsis, in which many clustered structural variants arise in a single catastrophic event, is frequently an early event in tumour evolution; in acral melanoma, for example, these events precede most somatic point mutations and affect several cancer-associated genes simultaneously. Cancers with abnormal telomere maintenance often originate from tissues with low replicative activity and show several mechanisms of preventing telomere attrition to critical levels. Common and rare germline variants affect patterns of somatic mutation, including point mutations, structural variants and somatic retrotransposition. A collection of papers from the PCAWG Consortium describes non-coding mutations that drive cancer beyond those in the TERT promoter(4); identifies new signatures of mutational processes that cause base substitutions, small insertions and deletions and structural variation(5,6); analyses timings and patterns of tumour evolution(7); describes the diverse transcriptional consequences of somatic mutation on splicing, expression levels, fusion genes and promoter activity(8,9); and evaluates a range of more-specialized features of cancer genomes(8,10-18).Peer reviewe

Whole-genome characterization of lung adenocarcinomas lacking the RTK/RAS/RAF pathway

RTK/RAS/RAF pathway alterations (RPAs) are a hallmark of lung adenocarcinoma (LUAD). In this study, we use whole-genome sequencing (WGS) of 85 cases found to be RPA(−) by previous studies from The Cancer Genome Atlas (TCGA) to characterize the minority of LUADs lacking apparent alterations in this pathway. We show that WGS analysis uncovers RPA(+) in 28 (33%) of the 85 samples. Among the remaining 57 cases, we observe focal deletions targeting the promoter or transcription start site of STK11 (n = 7) or KEAP1 (n = 3), and promoter mutations associated with the increased expression of ILF2 (n = 6). We also identify complex structural variations associated with high-level copy number amplifications. Moreover, an enrichment of focal deletions is found in TP53 mutant cases. Our results indicate that RPA(−) cases demonstrate tumor suppressor deletions and genome instability, but lack unique or recurrent genetic lesions compensating for the lack of RPAs. Larger WGS studies of RPA(−) cases are required to understand this important LUAD subset. © 2021 The AuthorsCarrot-Zhang et al. perform whole-genome characterization of lung adenocarcinomas (LUADs) lacking RTK/RAS/RAF pathway alterations (RPAs) and identify mutations or structural variants in both coding and non-coding spaces that define a unique entity of RPA(−) LUADs and potentially explain the underlying biology of this disease