title: Cancer-associated SF3B1 mutations affect alternative splicing by promoting alternative branchpoint usage.
abstract: Hotspot mutations in the spliceosome gene SF3B1 are reported in ∼20% of uveal melanomas. SF3B1 is involved in 3'-splice site (3'ss) recognition during RNA splicing; however, the molecular mechanisms of its mutation have remained unclear. Here we show, using RNA-Seq analyses of uveal melanoma, that the SF3B1(R625/K666) mutation results in deregulated splicing at a subset of junctions, mostly by the use of alternative 3'ss. Modelling the differential junctions in SF3B1(WT) and SF3B1(R625/K666) cell lines demonstrates that the deregulated splice pattern strictly depends on SF3B1 status and on the 3'ss-sequence context. SF3B1(WT) knockdown or overexpression do not reproduce the SF3B1(R625/K666) splice pattern, qualifying SF3B1(R625/K666) as change-of-function mutants. Mutagenesis of predicted branchpoints reveals that the SF3B1(R625/K666)-promoted splice pattern is a direct result of alternative branchpoint usage. Altogether, this study provides a better understanding of the mechanisms underlying splicing alterations induced by mutant SF3B1 in cancer, and reveals a role for alternative branchpoints in disease.
authors: Department of Genetics and Biology of Cancers, INSERM U830, Institut Curie, PSL Research University, Paris 75248, France.
authors: Samar
authors: S
authors: Alsafadi
authors: Department of Genetics and Biology of Cancers, INSERM U830, Institut Curie, PSL Research University, Paris 75248, France.
authors: Alexandre
authors: A
authors: Houy
authors: Department of Genetics and Biology of Cancers, INSERM U830, Institut Curie, PSL Research University, Paris 75248, France.
authors: Aude
authors: A
authors: Battistella
authors: Department of Genetics and Biology of Cancers, INSERM U830, Institut Curie, PSL Research University, Paris 75248, France.
authors: Tatiana
authors: T
authors: Popova
authors: Depatment of Developmental Biology and Genetics, CNRS UMR 3215/INSERM U934, Institut Curie, PSL Research University, Paris 75248, France.
authors: Michel
authors: M
authors: Wassef
authors: Translational Research Department, Institut Curie, PSL Research University, Paris 75248, France.
authors: Emilie
authors: E
authors: Henry
authors: Department of Genetics and Biology of Cancers, INSERM U830, Institut Curie, PSL Research University, Paris 75248, France.
authors: Franck
authors: http://orcid.org/0000-0003-4731-7817
authors: F
authors: Tirode
authors: Department of Molecular Bases of Human Diseases, CNRS UPR 1142, IGH-Institute of Human Genetics, Montpellier 34090, France.
authors: Angelos
authors: A
authors: Constantinou
authors: Department of Medical Oncology, Institut Curie, Paris 75248, France.
authors: Sophie
authors: S
authors: Piperno-Neumann
authors: Translational Research Department, Institut Curie, PSL Research University, Paris 75248, France.
authors: Sergio
authors: S
authors: Roman-Roman
authors: Department of Genotoxic stress and Cancer, CNRS UMR 3348, Institut Curie, PSL Research University, Orsay 91400, France.
authors: Martin
authors: M
authors: Dutertre
authors: Department of Genetics and Biology of Cancers, INSERM U830, Institut Curie, PSL Research University, Paris 75248, France.
authors: Marc-Henri
authors: MH
authors: Stern
mesh_terms: D017398:Alternative Splicing / Q000235:genetics*; D045744:Cell Line, Tumor; D057809:HEK293 Cells; D006801:Humans; D015151:Immunoblotting; D047468:Immunoprecipitation; D008545:Melanoma / Q000235:genetics*; D009154:Mutation; D010750:Phosphoproteins / Q000235:genetics*; D022821:RNA Splice Sites / Q000235:genetics*; D000072260:RNA Splicing Factors; D017413:Ribonucleoprotein, U2 Small Nuclear / Q000235:genetics*; D017422:Sequence Analysis, DNA; D017423:Sequence Analysis, RNA; D014604:Uveal Neoplasms / Q000235:genetics*; D000098943:Uveal Melanoma
chemical_list: D010750:Phosphoproteins; D022821:RNA Splice Sites; D000072260:RNA Splicing Factors; D017413:Ribonucleoprotein, U2 Small Nuclear; C112491:SF3B1 protein, human