Long-read sequencing von chronischer lymphatischer Leukämie und myelodysplastischem Syndrom mit und ohne SF3B1 Mutationen zur Identifikation von Mechanismen genomischer Instabilität, Therapieresistenz und deren Einfluss auf die Prognose.
Zusammenfassung der Projektergebnisse
Alternative splicing plays a critical role in generating transcriptome diversity and is frequently observed in cancer. Mutations in the splicing factor SF3B1 are particularly common in patients with chronic lymphocytic leukaemia (CLL) myelodysplastic syndromes (MDS) and other malignancies. To investigate the impact of SF3B1 mutations on alternative splicing, we used long-read sequencing to profile splicing events in a MDS and CLL patient cohort consisting of 19 CLL and 25 MDS patients, as well as cell lines with and without SF3B1 mutations. For the data analyses we developed a software package (IsoTools) for long-read transcriptome sequencing. The software was extensively tested with public and proprietary data. Furthermore, with IsoTools we participated at the LRGASP challenge involving diverse controlled data sets achieving excellent results and performance. The LRGASP challenge is currently accepted by Nature Methods. Our results revealed that SF3B1 mutations specifically alter the usage of 3' alternative splice sites, followed by a decrease in intron retention in SF3B1 mutated samples. With the long-read transcriptome sequencing data we were able to detect more than 60% of novel isoforms including alternative spliced isoforms of MAP3K7, BRD9, SEPTIN6, SETD4 and SF3B1 itself. We performed splicing reporter assays to confirm our findings. These results are now summarized in a manuscript and under review for publication.
Projektbezogene Publikationen (Auswahl)
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Altered DNA Methylation Profiles in SF3B1 Mutated CLL Patients. International Journal of Molecular Sciences, 22(17), 9337.
Pacholewska, Alicja; Grimm, Christina; Herling, Carmen D.; Lienhard, Matthias; Königs, Anja; Timmermann, Bernd; Altmüller, Janine; Mücke, Oliver; Reinhardt, Hans Christian; Plass, Christoph; Herwig, Ralf; Hallek, Michael & Schweiger, Michal R.
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Systematic assessment of long-read RNA-seq methods for transcript identification and quantification.
Pardo-Palacios, Francisco; Reese, Fairlie; Carbonell-Sala, Silvia; Diekhans, Mark; Liang, Cindy; Wang, Dingjie; Williams, Brian; Adams, Matthew; Behera, Amit; Lagarde, Julien; Li, Haoran; Prjibelski, Andrey; Balderrama-Gutierrez, Gabriela; Çelik, Muhammed Hasan; De María, Maite; Denslow, Nancy; Garcia-Reyero, Natàlia; Goetz, Stefan; Hunter, Margaret ... & Brooks, Angela N.
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IsoTools: a flexible workflow for long-read transcriptome sequencing analysis. Bioinformatics, 39(6).
Lienhard, Matthias; van den Beucken, Twan; Timmermann, Bernd; Hochradel, Myriam; Börno, Stefan; Caiment, Florian; Vingron, Martin & Herwig, Ralf
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Long-read transcriptome sequencing of CLL and MDS patients uncovers molecular effects of SF3B1 mutations. Genome Research, 34(11), 1832-1848.
Pacholewska, Alicja; Lienhard, Matthias; Brüggemann, Mirko; Hänel, Heike; Bilalli, Lorina; Königs, Anja; Heß, Felix; Becker, Kerstin; Köhrer, Karl; Kaiser, Jesko; Gohlke, Holger; Gattermann, Norbert; Hallek, Michael; Herling, Carmen D.; König, Julian; Grimm, Christina; Herwig, Ralf; Zarnack, Kathi & Schweiger, Michal R.
