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Hydroxyurea induces de novo copy number variants in human cells.

ABSTRACT: Copy number variants (CNVs) are widely distributed throughout the human genome, where they contribute to genetic variation and phenotypic diversity. Spontaneous CNVs are also a major cause of genetic and developmental disorders and arise frequently in cancer cells. As with all mutation classes, genetic and environmental factors almost certainly increase the risk for new and deleterious CNVs. However, despite the importance of CNVs, there is limited understanding of these precipitating risk factors and the mechanisms responsible for a large percentage of CNVs. Here we report that low doses of hydroxyurea, an inhibitor of ribonucleotide reductase and an important drug in the treatment of sickle cell disease and other diseases induces a high frequency of de novo CNVs in cultured human cells that resemble pathogenic and aphidicolin-induced CNVs in size and breakpoint structure. These CNVs are distributed throughout the genome, with some hotspots of de novo CNV formation. Sequencing revealed that CNV breakpoint junctions are characterized by short microhomologies, blunt ends, and short insertions. These data provide direct experimental support for models of replication-error origins of CNVs and suggest that any agent or condition that leads to replication stress has the potential to induce deleterious CNVs. In addition, they point to a need for further study of the genomic consequences of the therapeutic use of hydroxyurea.

SUBMITTER: Arlt MF

PROVIDER: S-EPMC3198378 | biostudies-literature | 2011 Oct

REPOSITORIES: biostudies-literature

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Hydroxyurea induces de novo copy number variants in human cells.

Arlt Martin F MF Ozdemir Alev Cagla AC Birkeland Shanda R SR Wilson Thomas E TE Glover Thomas W TW

Proceedings of the National Academy of Sciences of the United States of America 20111010 42

Copy number variants (CNVs) are widely distributed throughout the human genome, where they contribute to genetic variation and phenotypic diversity. Spontaneous CNVs are also a major cause of genetic and developmental disorders and arise frequently in cancer cells. As with all mutation classes, genetic and environmental factors almost certainly increase the risk for new and deleterious CNVs. However, despite the importance of CNVs, there is limited understanding of these precipitating risk facto ...[more]

PMID: 21987784

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Project description:BackgroundExon-targeted microarrays can detect small (<1000 bp) intragenic copy number variants (CNVs), including those that affect only a single exon. This genome-wide high-sensitivity approach increases the molecular diagnosis for conditions with known disease-associated genes, enables better genotype-phenotype correlations, and facilitates variant allele detection allowing novel disease gene discovery.MethodsWe retrospectively analyzed data from 63,127 patients referred for clinical chromosomal microarray analysis (CMA) at Baylor Genetics laboratories, including 46,755 individuals tested using exon-targeted arrays, from 2007 to 2017. Small CNVs harboring a single gene or two to five non-disease-associated genes were identified; the genes involved were evaluated for a potential disease association.ResultsIn this clinical population, among rare CNVs involving any single gene reported in 7200 patients (11%), we identified 145 de novo autosomal CNVs (117 losses and 28 intragenic gains), 257 X-linked deletion CNVs in males, and 1049 inherited autosomal CNVs (878 losses and 171 intragenic gains); 111 known disease genes were potentially disrupted by de novo autosomal or X-linked (in males) single-gene CNVs. Ninety-one genes, either recently proposed as candidate disease genes or not yet associated with diseases, were disrupted by 147 single-gene CNVs, including 37 de novo deletions and ten de novo intragenic duplications on autosomes and 100 X-linked CNVs in males. Clinical features in individuals with de novo or X-linked CNVs encompassing at most five genes (224 bp to 1.6 Mb in size) were compared to those in individuals with larger-sized deletions (up to 5 Mb in size) in the internal CMA database or loss-of-function single nucleotide variants (SNVs) detected by clinical or research whole-exome sequencing (WES). This enabled the identification of recently published genes (BPTF, NONO, PSMD12, TANGO2, and TRIP12), novel candidate disease genes (ARGLU1 and STK3), and further confirmation of disease association for two recently proposed disease genes (MEIS2 and PTCHD1). Notably, exon-targeted CMA detected several pathogenic single-exon CNVs missed by clinical WES analyses.ConclusionsTogether, these data document the efficacy of exon-targeted CMA for detection of genic and exonic CNVs, complementing and extending WES in clinical diagnostics, and the potential for discovery of novel disease genes by genome-wide assay.

Project description:BackgroundIn studies of case-parent trios, we define copy number variants (CNVs) in the offspring that differ from the parental copy numbers as de novo and of interest for their potential functional role in disease. Among the leading array-based methods for discovery of de novo CNVs in case-parent trios is the joint hidden Markov model (HMM) implemented in the PennCNV software. However, the computational demands of the joint HMM are substantial and the extent to which false positive identifications occur in case-parent trios has not been well described. We evaluate these issues in a study of oral cleft case-parent trios.ResultsOur analysis of the oral cleft trios reveals that genomic waves represent a substantial source of false positive identifications in the joint HMM, despite a wave-correction implementation in PennCNV. In addition, the noise of low-level summaries of relative copy number (log R ratios) is strongly associated with batch and correlated with the frequency of de novo CNV calls. Exploiting the trio design, we propose a univariate statistic for relative copy number referred to as the minimum distance that can reduce technical variation from probe effects and genomic waves. We use circular binary segmentation to segment the minimum distance and maximum a posteriori estimation to infer de novo CNVs from the segmented genome. Compared to PennCNV on simulated data, MinimumDistance identifies fewer false positives on average and is comparable to PennCNV with respect to false negatives. Genomic waves contribute to discordance of PennCNV and MinimumDistance for high coverage de novo calls, while highly concordant calls on chromosome 22 were validated by quantitative PCR. Computationally, MinimumDistance provides a nearly 8-fold increase in speed relative to the joint HMM in a study of oral cleft trios.ConclusionsOur results indicate that batch effects and genomic waves are important considerations for case-parent studies of de novo CNV, and that the minimum distance is an effective statistic for reducing technical variation contributing to false de novo discoveries. Coupled with segmentation and maximum a posteriori estimation, our algorithm compares favorably to the joint HMM with MinimumDistance being much faster.

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Hydroxyurea induces de novo copy number variants in human cells.

Publications

Hydroxyurea induces de novo copy number variants in human cells.

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