Transcriptome sequencing reveals breast cancer rearrangement

In a paper to be published this week in the online edition of the Proceedings of the National Academy of Sciences (PNAS), an international research team introduced a high-throughput transcriptome sequencing to discover genome weight in breast cancer cell lines. The original theoretical proof of the row.

Researchers at the J.Craig Venter Institute, three branches of the Lugwig Cancer Institute, and the Memorial Sloan-Kettering Cancer in New York use Roche 454 Transcriptome sequencing to find genomic translocations in a highly rearranged breast cancer cell line. In the process, they discovered seven new genomic rearrangements - including five truncated proteins and two chimeric proteins, which are believed to affect at least nine genes.

The author of the article, Robert Strausberg, is also the acting director and team leader for JCVI Genomic Medicine. He told the reporter of GenomeWeb Daily News: "We are very excited about this." Strausberg praised high-throughput sequencing for providing researchers with a way to understand A new type of information for a transcriptome. He said: "This gives a detailed understanding of the dreams of so many transcripts in cells." Multiple studies confirm the effectiveness of large-scale genomic analysis in identifying mutations, rearrangements, and gene expression changes associated with multiple cancers. . For this latest study, Strausberg and his colleagues gathered information hidden in the transcriptome, a collection of transcripts in cells. Since the transcriptome reflects an active genome, it contains everything from different gene splicing forms to gene expression. But Strausberg and his team also wanted to learn about other information from the transcriptome - clues about genomic translocation events. To detect active gene products for gene rearrangement, they used Roche 454-FLX pyrosequencing to assess the transcriptome of the highly rearranged breast cancer cell line HCC1954.

The team generated 510,703 cDNA sequence reads from this cell line. Of these readings, more than 384,900 readings were paired with 9,221 RefSeq gene mRNAs. Excluding these sequences paired with RefSeq, the researchers attempted to align the remaining sequences with the human reference genome, leaving 47,370 sequences unpairable. The researchers then placed the remaining sequences into a computer analysis process to extract 496 potential chimeric transcripts containing at least two different genomic locus information. Approximately half of the transcripts represent rearrangements on the same chromosome, while the other half contains rearrangements between different chromosomes. The team selected 33 putative chimeras for subsequent studies and experimentally verified 13 chimeric cDNAs. Most of the variation detected by the panel is also present in the blood cell control line of the same individual.

This prompted researchers to use Long Range PCR, Sanger end sequencing, and fluorescence in situ hybridization to distinguish between trans-splicing events and true genomic rearrangements. The human genome scientist at Qi Zhao-JCVI, the lead author of the article, told GenomeWeb Daily News that although some of the rearrangements they found overlap with known changes in cell lines, others are new. For example, they found four inter-chromosomal translocations and one intrachromosomal rearrangement in the initial experiments. Next, the team went back to find chimeric transcripts that could be localized to more than one locus on the genome. In the process, they discovered and confirmed the rearrangement between the two chromosomes.

A total of seven rearrangements were identified in the study, and it is believed that at least nine different genes were affected. Five of the rearranged transcripts encode truncated proteins—including proteins with oncogenic functions. For example, the researchers detected truncations of MRE11A and NSD1. The protein encoded by the MRE11A gene is involved in the repair of double-strand breaks detected in several breast cancers, while the NSD1 gene encodes a possible transcriptional regulator, sometimes present in acute myeloids. Cell white
In blood disease.

The authors write: "Our results suggest that genomic translocation may be another mechanism of gene inactivation in breast cancer, and it may be as common as point mutations." In addition to this original theoretical study, Strausberg said the final The goal is to examine and understand cancer at a clinical level. He and his collaborators emphasized the need to edit and integrate different types of data—including genomic rearrangements and alternative splicing events—to increase the chances of getting better results from research. The authors conclude that: "The database provides the possibility to compare and contrast these changes in a variety of cancers, and it is important to identify common features in cancer, thus providing an opportunity to apply new interventions to effectively treat all cancers, and The patient's results can also be improved."

454 Life Sciences is one of Roche's Centers of Excellence in Applied Science, developing and commercializing the innovative 454 Sequencing System for ultra-high throughput DNA sequencing. Specific applications include de novo sequencing and resequencing of genomes, metagenomics, RNA analysis, and directed sequencing of DNA target regions. The 454 sequencing system features simple sample preparation and no bias, and the read length includes long read lengths and high accuracy. The technology of the 454 Sequencing System has spawned hundreds of peer-reviewed studies in various research areas, such as cancer and infectious disease research, drug screening, marine biology, anthropology, and paleontology.

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