Aberrant splicing and defective mRNA production induced by somatic spliceosome mutations in myelodysplasia

Yusuke Shiozawa,Yusuke Sato,Satoru Miyano,Aiko Sato-Otsubo,Hiromichi Suzuki,Kenichi Yoshida,Kenichi Chiba,Yosaku Watatani,Mario Cazzola,Keisuke Kataoka,Hideki Makishima,Yuichi Shiraishi,Masashi Sanada,Luca Malcovati,Seishi Ogawa,Tetsuichi Yoshizato,Anna Gallì,Eva Hellström-Lindberg

doi:10.1038/s41467-018-06063-x

Abstract

Spliceosome mutations are frequently found in myelodysplasia. Splicing alterations induced by these mutations, their precise targets, and the effect at the transcript level have not been fully elucidated. Here we report transcriptomic analyses of 265 bone marrow samples from myelodysplasia patients, followed by a validation using CRISPR/Cas9-mediated gene editing and an assessment of nonsense-mediated decay susceptibility. Small but widespread reduction of intron-retaining isoforms is the most frequent splicing alteration in SF3B1-mutated samples. SF3B1 mutation is also associated with 3′ splice site alterations, leading to the most pronounced reduction of canonical transcripts. Target genes include tumor suppressors and genes of mitochondrial iron metabolism or heme biosynthesis. Alternative exon usage is predominant in SRSF2- and U2AF1-mutated samples. Usage of an EZH2 cryptic exon harboring a premature termination codon is increased in both SRSF2- and U2AF1-mutated samples. Our study reveals a landscape of splicing alterations and precise targets of various spliceosome mutations.

Highlights

Spliceosome mutations are frequently found in myelodysplasia
We reveal that SRSF2 and U2AF1 mutations have a common target of splicing alterations: EZH2
We studied 214 patients with myeloid neoplasms with myelodysplasia followed at the Department of Hematology Oncology, Fondazione IRCCS Policlinico San Matteo, Pavia, Italy

Summary

Introduction

Spliceosome mutations are frequently found in myelodysplasia. Splicing alterations induced by these mutations, their precise targets, and the effect at the transcript level have not been fully elucidated. Specific alternative splicing events observed in human cells were often absent in mouse models, in which binding site sequences of mutant SFs are not always conserved[19,20,23] This highlights the importance of identifying precise targets of aberrant RNA splicing in human hematopoietic cells. A frameshift or a premature termination codon (PTC) that is frequently introduced by mutant-SF-associated mis-splicing events is likely to have deleterious effects on gene function with a reduction in their canonical transcripts[10,11,12]. A reduction of canonical transcripts due to mutant-SF-induced mis-splicing events and their sensitivity to NMD have not been evaluated in a comprehensive manner To address these questions, we perform a comprehensive transcriptome analysis on a large cohort of patients with myelodysplasia. This study provides a landscape of abnormal splicing associated with different spliceosome mutations and their precise targets

Methods

Results

Conclusion