RESUMEN
Cancer is driven by mutation. Worldwide, tobacco smoking is the principal lifestyle exposure that causes cancer, exerting carcinogenicity through >60 chemicals that bind and mutate DNA. Using massively parallel sequencing technology, we sequenced a small-cell lung cancer cell line, NCI-H209, to explore the mutational burden associated with tobacco smoking. A total of 22,910 somatic substitutions were identified, including 134 in coding exons. Multiple mutation signatures testify to the cocktail of carcinogens in tobacco smoke and their proclivities for particular bases and surrounding sequence context. Effects of transcription-coupled repair and a second, more general, expression-linked repair pathway were evident. We identified a tandem duplication that duplicates exons 3-8 of CHD7 in frame, and another two lines carrying PVT1-CHD7 fusion genes, indicating that CHD7 may be recurrently rearranged in this disease. These findings illustrate the potential for next-generation sequencing to provide unprecedented insights into mutational processes, cellular repair pathways and gene networks associated with cancer.
Asunto(s)
Neoplasias Pulmonares/etiología , Neoplasias Pulmonares/genética , Mutación/genética , Nicotiana/efectos adversos , Carcinoma Pulmonar de Células Pequeñas/etiología , Carcinoma Pulmonar de Células Pequeñas/genética , Fumar/efectos adversos , Carcinógenos/toxicidad , Línea Celular Tumoral , Variaciones en el Número de Copia de ADN/efectos de los fármacos , Variaciones en el Número de Copia de ADN/genética , Daño del ADN/genética , ADN Helicasas/genética , Análisis Mutacional de ADN , Reparación del ADN/genética , Proteínas de Unión al ADN/genética , Exones/genética , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Genoma Humano/efectos de los fármacos , Genoma Humano/genética , Humanos , Mutagénesis Insercional/efectos de los fármacos , Mutagénesis Insercional/genética , Mutación/efectos de los fármacos , Regiones Promotoras Genéticas/genética , Eliminación de Secuencia/genéticaRESUMEN
We describe the genome sequencing of an anonymous individual of African origin using a novel ligation-based sequencing assay that enables a unique form of error correction that improves the raw accuracy of the aligned reads to >99.9%, allowing us to accurately call SNPs with as few as two reads per allele. We collected several billion mate-paired reads yielding approximately 18x haploid coverage of aligned sequence and close to 300x clone coverage. Over 98% of the reference genome is covered with at least one uniquely placed read, and 99.65% is spanned by at least one uniquely placed mate-paired clone. We identify over 3.8 million SNPs, 19% of which are novel. Mate-paired data are used to physically resolve haplotype phases of nearly two-thirds of the genotypes obtained and produce phased segments of up to 215 kb. We detect 226,529 intra-read indels, 5590 indels between mate-paired reads, 91 inversions, and four gene fusions. We use a novel approach for detecting indels between mate-paired reads that are smaller than the standard deviation of the insert size of the library and discover deletions in common with those detected with our intra-read approach. Dozens of mutations previously described in OMIM and hundreds of nonsynonymous single-nucleotide and structural variants in genes previously implicated in disease are identified in this individual. There is more genetic variation in the human genome still to be uncovered, and we provide guidance for future surveys in populations and cancer biopsies.