Dataset Information

Precision analysis of mutant U2AF1 activity reveals deployment of stress granules in myeloid malignancies.

ABSTRACT: Splicing factor mutations are common among cancers, recently emerging as drivers of myeloid malignancies. U2AF1 carries hotspot mutations in its RNA-binding motifs; however, how they affect splicing and promote cancer remain unclear. The U2AF1/U2AF2 heterodimer is critical for 3' splice site (3'SS) definition. To specifically unmask changes in U2AF1 function in vivo, we developed a crosslinking and immunoprecipitation procedure that detects contacts between U2AF1 and the 3'SS AG at single-nucleotide resolution. Our data reveal that the U2AF1 S34F and Q157R mutants establish new 3'SS contacts at -3 and +1 nucleotides, respectively. These effects compromise U2AF2-RNA interactions, resulting predominantly in intron retention and exon exclusion. Integrating RNA binding, splicing, and turnover data, we predicted that U2AF1 mutations directly affect stress granule components, which was corroborated by single-cell RNA-seq. Remarkably, U2AF1-mutant cell lines and patient-derived MDS/AML blasts displayed a heightened stress granule response, pointing to a novel role for biomolecular condensates in adaptive oncogenic strategies.

SUBMITTER: Biancon G

PROVIDER: S-EPMC8988922 | biostudies-literature | 2022 Mar

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Precision analysis of mutant U2AF1 activity reveals deployment of stress granules in myeloid malignancies.

Biancon Giulia G Joshi Poorval P Zimmer Joshua T JT Hunck Torben T Gao Yimeng Y Lessard Mark D MD Courchaine Edward E Barentine Andrew E S AES Machyna Martin M Botti Valentina V Qin Ashley A Gbyli Rana R Patel Amisha A Song Yuanbin Y Kiefer Lea L Viero Gabriella G Neuenkirchen Nils N Lin Haifan H Bewersdorf Joerg J Simon Matthew D MD Neugebauer Karla M KM Tebaldi Toma T Halene Stephanie S

Molecular cell 20220301 6

Splicing factor mutations are common among cancers, recently emerging as drivers of myeloid malignancies. U2AF1 carries hotspot mutations in its RNA-binding motifs; however, how they affect splicing and promote cancer remain unclear. The U2AF1/U2AF2 heterodimer is critical for 3' splice site (3'SS) definition. To specifically unmask changes in U2AF1 function in vivo, we developed a crosslinking and immunoprecipitation procedure that detects contacts between U2AF1 and the 3'SS AG at single-nucleo ...[more]

PMID: 35303483

Similar Datasets

Project description:BackgroundMyelodysplastic syndromes (MDS) arise from somatic mutations acquired in hematopoietic stem and progenitor cells, causing cytopenias and predisposing to transformation into secondary acute myeloid leukemia (sAML). Recurrent mutations in spliceosome genes, including U2AF1, are attractive therapeutic targets as they are prevalent in MDS and sAML, arise early in neoplastic cells, and are generally absent from normal cells, including normal hematopoietic cells. MDS and sAML are susceptible to T cell-mediated killing, and thus engineered T-cell immunotherapies hold promise for their treatment. We hypothesized that targeting spliceosome mutation-derived neoantigens with transgenic T-cell receptor (TCR) T cells would selectively eradicate malignant cells in MDS and sAML.MethodsWe identified candidate neoantigen epitopes from recurrent protein-coding mutations in the spliceosome genes SRSF2 and U2AF1 using a multistep in silico process. Candidate epitopes predicted to bind human leukocyte antigen (HLA) class I, be processed and presented from the parent protein, and not to be subject to tolerance then underwent in vitro immunogenicity screening. CD8+ T cells recognizing immunogenic neoantigen epitopes were evaluated in in vitro assays to assess functional avidity, confirm the predicted HLA restriction, the potential for recognition of similar peptides, and the ability to kill neoplastic cells in an antigen-specific manner. Neoantigen-specific TCR were sequenced, cloned into lentiviral vectors, and transduced into third-party T cells after knock-out of endogenous TCR, then tested in vitro for specificity and ability to kill neoplastic myeloid cells presenting the neoantigen. The efficacy of neoantigen-specific T cells was evaluated in vivo in a murine cell line-derived xenograft model.ResultsWe identified two neoantigens created from a recurrent mutation in U2AF1, isolated CD8+ T cells specific for the neoantigens, and demonstrated that transferring their TCR to third-party CD8+ T cells is feasible and confers specificity for the U2AF1 neoantigens. Finally, we showed that these neoantigen-specific TCR-T cells do not recognize normal hematopoietic cells but efficiently kill malignant myeloid cells bearing the specific U2AF1 mutation, including primary cells, in vitro and in vivo.ConclusionsThese data serve as proof-of-concept for developing precision medicine approaches that use neoantigen-directed T-cell receptor-transduced T cells to treat MDS and sAML.

Dataset Information

Precision analysis of mutant U2AF1 activity reveals deployment of stress granules in myeloid malignancies.

Publications

Precision analysis of mutant U2AF1 activity reveals deployment of stress granules in myeloid malignancies.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets