Dataset Information

Wiskott-Aldrich syndrome protein forms nuclear condensates and regulates alternative splicing.

ABSTRACT: The diverse functions of WASP, the deficiency of which causes Wiskott-Aldrich syndrome (WAS), remain poorly defined. We generated three isogenic WAS models using patient induced pluripotent stem cells and genome editing. These models recapitulated WAS phenotypes and revealed that WASP deficiency causes an upregulation of numerous RNA splicing factors and widespread altered splicing. Loss of WASP binding to splicing factor gene promoters frequently leads to aberrant epigenetic activation. WASP interacts with dozens of nuclear speckle constituents and constrains SRSF2 mobility. Using an optogenetic system, we showed that WASP forms phase-separated condensates that encompasses SRSF2, nascent RNA and active Pol II. The role of WASP in gene body condensates is corroborated by ChIPseq and RIPseq. Together our data reveal that WASP is a nexus regulator of RNA splicing that controls the transcription of splicing factors epigenetically and the dynamics of the splicing machinery through liquid-liquid phase separation.

SUBMITTER: Yuan B

PROVIDER: S-EPMC9233711 | biostudies-literature | 2022 Jun

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Wiskott-Aldrich syndrome protein forms nuclear condensates and regulates alternative splicing.

Yuan Baolei B Zhou Xuan X Suzuki Keiichiro K Ramos-Mandujano Gerardo G Wang Mengge M Tehseen Muhammad M Cortés-Medina Lorena V LV Moresco James J JJ Dunn Sarah S Hernandez-Benitez Reyna R Hishida Tomoaki T Kim Na Young NY Andijani Manal M MM Bi Chongwei C Ku Manching M Takahashi Yuta Y Xu Jinna J Qiu Jinsong J Huang Ling L Benner Christopher C Aizawa Emi E Qu Jing J Liu Guang-Hui GH Li Zhongwei Z Yi Fei F Ghosheh Yanal Y Shao Changwei C Shokhirev Maxim M Comoli Patrizia P Frassoni Francesco F Yates John R JR Fu Xiang-Dong XD Esteban Concepcion Rodriguez CR Hamdan Samir S Izpisua Belmonte Juan Carlos JC Li Mo M

Nature communications 20220625 1

The diverse functions of WASP, the deficiency of which causes Wiskott-Aldrich syndrome (WAS), remain poorly defined. We generated three isogenic WAS models using patient induced pluripotent stem cells and genome editing. These models recapitulated WAS phenotypes and revealed that WASP deficiency causes an upregulation of numerous RNA splicing factors and widespread altered splicing. Loss of WASP binding to splicing factor gene promoters frequently leads to aberrant epigenetic activation. WASP in ...[more]

PMID: 35752626

Similar Datasets

Project description:Wiskott-Aldrich syndrome (WAS), which is caused by mutations in the gene encoding WASP, manifests in a wide range of hematologic and immune dysfunctions and predisposition to cancer development 1. WASP is most known as a cytoplasmic effector of actin cytoskeleton rearrangement. However, defective actin polymerization cannot explain many aspects of WAS pathogenesis. Incomplete knowledge of WASP function precludes in-depth understanding of the underlying mechanisms of WAS, and therefore hampers the development of effective therapies. Here we generated induced pluripotent stem cells (iPSCs) from WAS patients (WAS-iPSC) and corresponding isogenic iPSCs wherein the mutations were corrected by targeted genome editing. Hematopoietic cells differentiated from WAS-iPSCs not only recapitulated known disease phenotypes, but also revealed novel nuclear functions of WASP. WASP deficiency causes differential expression of genes involved in many aspects of immune cell function, most prominently cell proliferation. It also leads to a large number of alternative splicing events that are highly enriched in cell cycle regulators. Proteomic analyses revealed that WASP physically interacted with nuclear body components, nuclear structural proteins, chromatin modifying complexes, and many RNA-binding proteins including multiple splicing factors. We show that WASP physically interacts with SRSF2 and is important for nuclear speckle organization. WASP regulates cell cycle progression by physically interacting with EZH2 and thereby influencing epigenetic silencing of its target gene CDKN2A (aka p16). Together, these observations unveil novel functions of WASP in regulation of RNA splicing and cell cycle. The binding of WASP to SRSF2 and EZH2 and the misregulation of these two frequently mutated genes in hematologic malignancies2 in WAS cells provide a possible mechanism for the frequent occurance of malignancy in WAS patients. Furthermore, WAS-associated splicing and proliferation abnormalities could serve as diagnosis tools for patients at risk for cancer and may be exploited as therapeutic targets.

Project description:Wiskott-Aldrich syndrome (WAS) is characterized by X-linked thrombocytopenia, eczema, immunodeficiency, recurrent infections and increased risk of autoimmunity and malignancies. WAS is caused by mutations in the WAS gene, which encodes the exclusively hematopoietic WAS protein (WASp) that is classically characterized as aÎ½ actin nucleator. However, disruption of F-actin polymerization by WAS mutations can not account for many aspects of WAS pathogenesis. Ignorance of other functions of WASP precludes in-depth understanding of the pathogenic effects of mutant WASP, and therefore hampers development of effective therapy. Here we generated induced pluripotent stem cells (iPSCs) from WAS patients (WAS-iPSC) bearing different mutations and corresponding isogenic iPSCs in which the pathogenic mutations had been corrected by targeted genome editing. Hematopoietic cells differentiated from WAS-iPSCs not only recapitulated known disease phenotypes, but also revealed novel defects of WASP deficient cells. WASP co-localized with nuclear pores, nucleoli, nuclear speckles and PML bodies by immunocytochemistry and/or serial block face scanning microscopy (SBF-SEM). MudPIT (multi-dimensional protein identification technology) analysis revealed that WASP physically interacted with nuclear body components, nuclear structural proteins, chromatin modifying complexes, and many RNA-binding proteins including major components of the spliceosome. Next-generation sequencing captured a dramatic global change of alternative splicing in WAS patient cells. WAS mutation impacted splicing of multiple genes frequently mutated in myelodysplastic syndrome and other cancers. RNA sequencing showed that WAS-iPSC derived immune cells misregulated many cell cycle regulators, tumor suppressors, immune function genes and splicing factors, and activated gene networks that drive cancer development and inflammatory diseases. Together these data uncovered previously unappreciated functions of the WASP and provided a mechanistic understanding of the pathogenesis of malignancy and autoimmunity in the most severe form of WAS. These new knowledge could help develop targeted therapy for WAS in the future. Human WAS-iPSCs (p.Phe35*) and gene corrected WAS-iPSCs (cWAS-iPSC) were differentiated into macrophages. WAS patient derived B-lymphocyte line ID00003 (p.Glu133Lys) and a wile-type B-lymphocyte line GM11518 were cultured using standard protocol. Total RNAs were extracted and been analyzed by RASL-seq.

Project description:Wiskott-Aldrich syndrome (WAS) is characterized by X-linked thrombocytopenia, eczema, immunodeficiency, recurrent infections and increased risk of autoimmunity and malignancies. WAS is caused by mutations in the WAS gene, which encodes the exclusively hematopoietic WAS protein (WASp) that is classically characterized as aν actin nucleator. However, disruption of F-actin polymerization by WAS mutations can not account for many aspects of WAS pathogenesis. Ignorance of other functions of WASP precludes in-depth understanding of the pathogenic effects of mutant WASP, and therefore hampers development of effective therapy. Here we generated induced pluripotent stem cells (iPSCs) from WAS patients (WAS-iPSC) bearing different mutations and corresponding isogenic iPSCs in which the pathogenic mutations had been corrected by targeted genome editing. Hematopoietic cells differentiated from WAS-iPSCs not only recapitulated known disease phenotypes, but also revealed novel defects of WASP deficient cells. WASP co-localized with nuclear pores, nucleoli, nuclear speckles and PML bodies by immunocytochemistry and/or serial block face scanning microscopy (SBF-SEM). MudPIT (multi-dimensional protein identification technology) analysis revealed that WASP physically interacted with nuclear body components, nuclear structural proteins, chromatin modifying complexes, and many RNA-binding proteins including major components of the spliceosome. Next-generation sequencing captured a dramatic global change of alternative splicing in WAS patient cells. WAS mutation impacted splicing of multiple genes frequently mutated in myelodysplastic syndrome and other cancers. RNA sequencing showed that WAS-iPSC derived immune cells misregulated many cell cycle regulators, tumor suppressors, immune function genes and splicing factors, and activated gene networks that drive cancer development and inflammatory diseases. Together these data uncovered previously unappreciated functions of the WASP and provided a mechanistic understanding of the pathogenesis of malignancy and autoimmunity in the most severe form of WAS. These new knowledge could help develop targeted therapy for WAS in the future.

Dataset Information

Wiskott-Aldrich syndrome protein forms nuclear condensates and regulates alternative splicing.

Publications

Wiskott-Aldrich syndrome protein forms nuclear condensates and regulates alternative splicing.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets