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Identification of novel fusion genes in lung cancer using breakpoint assembly of transcriptome sequencing data.

Fernandez-Cuesta L, Sun R, Menon R, George J, Lorenz S, Meza-Zepeda LA, Peifer M, Plenker D, Heuckmann JM, Leenders F, Zander T, Dahmen I, Koker M, Schöttle J, Ullrich RT, Altmüller J, Becker C, Nürnberg P, Seidel H, Böhm D, Göke F, Ansén S, Russell PA, Wright GM, Wainer Z, Solomon B, Petersen I, Clement JH, Sänger J, Brustugun OT, Helland Å, Solberg S, Lund-Iversen M, Buettner R, Wolf J, Brambilla E, Vingron M, Perner S, Haas SA, Thomas RK - Genome Biol. (2015)

Bottom Line: Genomic translocation events frequently underlie cancer development through generation of gene fusions with oncogenic properties.Identification of such fusion transcripts by transcriptome sequencing might help to discover new potential therapeutic targets.We apply TRUP to RNA-seq data of different tumor types, and find it to be more sensitive than alternative tools in detecting chimeric transcripts, such as secondary rearrangements in EML4-ALK-positive lung tumors, or recurrent inactivating rearrangements affecting RASSF8.

View Article: PubMed Central - PubMed

ABSTRACT
Genomic translocation events frequently underlie cancer development through generation of gene fusions with oncogenic properties. Identification of such fusion transcripts by transcriptome sequencing might help to discover new potential therapeutic targets. We developed TRUP (Tumor-specimen suited RNA-seq Unified Pipeline) (https://github.com/ruping/TRUP), a computational approach that combines split-read and read-pair analysis with de novo assembly for the identification of chimeric transcripts in cancer specimens. We apply TRUP to RNA-seq data of different tumor types, and find it to be more sensitive than alternative tools in detecting chimeric transcripts, such as secondary rearrangements in EML4-ALK-positive lung tumors, or recurrent inactivating rearrangements affecting RASSF8.

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Recurrent inactivating rearrangements ofRASSF8in cancer. Identification of ASUN-RASSF8 in a lung adenocarcinoma tumor (upper panel) and RASSF8-MARS in the KPD osteosarcoma cell-line (lower panel). Schematic representation of the fusion transcripts and some of the transcriptome sequencing reads spanning the fusion point.
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Fig5: Recurrent inactivating rearrangements ofRASSF8in cancer. Identification of ASUN-RASSF8 in a lung adenocarcinoma tumor (upper panel) and RASSF8-MARS in the KPD osteosarcoma cell-line (lower panel). Schematic representation of the fusion transcripts and some of the transcriptome sequencing reads spanning the fusion point.

Mentions: We next applied the method to 17 additional primary lung adenocarcinoma specimens (Additional file 10). Of the 17 samples analyzed, one carried an inactivating chimeric RASSF8 transcript (Figure 5, upper panel; Additional file 11a). The sample was negative for EGFR or KRAS mutations and belonged to the adenocarcinoma of a current smoker (Additional file 10). RASSF8 is one of the four N-terminal RASSF proteins (RASSF7-10) that belong to the Ras-association-domain-containing family of proteins, which also include the classical RASSF proteins (RASSF1-6) that are known to act as tumor suppressors and are frequently epigenetically silenced in tumors [22]. In order to further investigate the role of RASSF8 in lung adenocarcinoma, we silenced RASSF8 expression in the lung cancer cell line, H1395, which expresses wild-type RASSF8. In comparison to the EGFP transfected cells, silencing of RASSF8 led to a significant increase of cell proliferation of more than 60% (P <0.0001) (Additional file 11b). RASSF8 was not completely silenced, as detected by western blotting (Additional file 11c), suggesting that low doses rather than complete loss of the RASSF8 protein is sufficient to induce cell proliferation. Furthermore, we identified an inactivating rearrangement of RASSF8 in the osteosarcoma cell line, KPD (Figure 5, lower panel; Additional file 11a). The breakpoint of this translocation event was also detectable when analyzing the copy number data (Additional file 12) suggesting that the rearrangement happened at the genomic level.Figure 5


Identification of novel fusion genes in lung cancer using breakpoint assembly of transcriptome sequencing data.

Fernandez-Cuesta L, Sun R, Menon R, George J, Lorenz S, Meza-Zepeda LA, Peifer M, Plenker D, Heuckmann JM, Leenders F, Zander T, Dahmen I, Koker M, Schöttle J, Ullrich RT, Altmüller J, Becker C, Nürnberg P, Seidel H, Böhm D, Göke F, Ansén S, Russell PA, Wright GM, Wainer Z, Solomon B, Petersen I, Clement JH, Sänger J, Brustugun OT, Helland Å, Solberg S, Lund-Iversen M, Buettner R, Wolf J, Brambilla E, Vingron M, Perner S, Haas SA, Thomas RK - Genome Biol. (2015)

Recurrent inactivating rearrangements ofRASSF8in cancer. Identification of ASUN-RASSF8 in a lung adenocarcinoma tumor (upper panel) and RASSF8-MARS in the KPD osteosarcoma cell-line (lower panel). Schematic representation of the fusion transcripts and some of the transcriptome sequencing reads spanning the fusion point.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4300615&req=5

Fig5: Recurrent inactivating rearrangements ofRASSF8in cancer. Identification of ASUN-RASSF8 in a lung adenocarcinoma tumor (upper panel) and RASSF8-MARS in the KPD osteosarcoma cell-line (lower panel). Schematic representation of the fusion transcripts and some of the transcriptome sequencing reads spanning the fusion point.
Mentions: We next applied the method to 17 additional primary lung adenocarcinoma specimens (Additional file 10). Of the 17 samples analyzed, one carried an inactivating chimeric RASSF8 transcript (Figure 5, upper panel; Additional file 11a). The sample was negative for EGFR or KRAS mutations and belonged to the adenocarcinoma of a current smoker (Additional file 10). RASSF8 is one of the four N-terminal RASSF proteins (RASSF7-10) that belong to the Ras-association-domain-containing family of proteins, which also include the classical RASSF proteins (RASSF1-6) that are known to act as tumor suppressors and are frequently epigenetically silenced in tumors [22]. In order to further investigate the role of RASSF8 in lung adenocarcinoma, we silenced RASSF8 expression in the lung cancer cell line, H1395, which expresses wild-type RASSF8. In comparison to the EGFP transfected cells, silencing of RASSF8 led to a significant increase of cell proliferation of more than 60% (P <0.0001) (Additional file 11b). RASSF8 was not completely silenced, as detected by western blotting (Additional file 11c), suggesting that low doses rather than complete loss of the RASSF8 protein is sufficient to induce cell proliferation. Furthermore, we identified an inactivating rearrangement of RASSF8 in the osteosarcoma cell line, KPD (Figure 5, lower panel; Additional file 11a). The breakpoint of this translocation event was also detectable when analyzing the copy number data (Additional file 12) suggesting that the rearrangement happened at the genomic level.Figure 5

Bottom Line: Genomic translocation events frequently underlie cancer development through generation of gene fusions with oncogenic properties.Identification of such fusion transcripts by transcriptome sequencing might help to discover new potential therapeutic targets.We apply TRUP to RNA-seq data of different tumor types, and find it to be more sensitive than alternative tools in detecting chimeric transcripts, such as secondary rearrangements in EML4-ALK-positive lung tumors, or recurrent inactivating rearrangements affecting RASSF8.

View Article: PubMed Central - PubMed

ABSTRACT
Genomic translocation events frequently underlie cancer development through generation of gene fusions with oncogenic properties. Identification of such fusion transcripts by transcriptome sequencing might help to discover new potential therapeutic targets. We developed TRUP (Tumor-specimen suited RNA-seq Unified Pipeline) (https://github.com/ruping/TRUP), a computational approach that combines split-read and read-pair analysis with de novo assembly for the identification of chimeric transcripts in cancer specimens. We apply TRUP to RNA-seq data of different tumor types, and find it to be more sensitive than alternative tools in detecting chimeric transcripts, such as secondary rearrangements in EML4-ALK-positive lung tumors, or recurrent inactivating rearrangements affecting RASSF8.

Show MeSH
Related in: MedlinePlus