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Wash regulates lamin-dependent nuclear genome organization [Chromatin Accessibility]

ABSTRACT: The cytoplasmic functions of Wiskott-Aldrich Syndrome family (WASP) proteins are well known and include roles in cytoskeleton reorganization and membrane-cytoskeletal interactions important for membrane/vesicle trafficking, morphogenesis, immune response and signal transduction. Mis-regulation of these proteins is associated with immune deficiency and metastasis. Cytoplasmic WASP proteins act as effectors of Rho family GTPases and polymerize branched actin through the Arp2/3 complex. However, recent evidence has revealed that this classically cytoplasmic protein family also functions in the nucleus. Previously, we identified Drosophila washout (wash) as a new member of the WASP family with essential cytoplasmic roles in early development. Here we show that Wash is also present in the nucleus and plays a key role in nuclear organization via its interaction with Lamin Dm0 at the nuclear envelope. Wash and Lamin Dm0 occupy similar genomic regions that overlap with transcriptionally silent chromatin including constitutive heterochromatin. Strikingly, wash mutant and knockdown nuclei exhibit the same abnormal wrinkled morphology observed in diverse laminopathies, including the Hutchinson-Gilford progeria syndrome, and consistent with disruption of the nuclear organization of several sub-nuclear structures including cajal bodies and the chromocenter in salivary glands. We also found that Wash and Lamin knockdown disrupt chromatin accessibility of repressive compartments in agreement with an observed global redistribution of repressive histone modifications. Functional genetic approaches show wash mutants exhibit similar phenotypes to lamin Dm0 mutants, suggesting they participate in similar regulatory networks. Our results reveal a novel role for Wash in modulating nuclear organization via its interaction with the nuclear envelope protein Lamin Dm0. These findings highlight the functional complexity of WASP family proteins and provide new venues to understand their molecular roles in cell biology and disease. We evaluated the effect of Wash knockdown in S2R+ cells on chromatin accessibility using an M.SssI-based approach.

ORGANISM(S): Drosophila melanogaster

PROVIDER: GSE59852 | GEO | 2015/03/10

SECONDARY ACCESSION(S): PRJNA256474

REPOSITORIES: GEO

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Project description:The cytoplasmic functions of Wiskott-Aldrich Syndrome family (WASP) proteins are well known and include roles in cytoskeleton reorganization and membrane-cytoskeletal interactions important for membrane/vesicle trafficking, morphogenesis, immune response and signal transduction. Mis-regulation of these proteins is associated with immune deficiency and metastasis. Cytoplasmic WASP proteins act as effectors of Rho family GTPases and polymerize branched actin through the Arp2/3 complex. However, recent evidence has revealed that this classically cytoplasmic protein family also functions in the nucleus. Previously, we identified Drosophila washout (wash) as a new member of the WASP family with essential cytoplasmic roles in early development. Here we show that Wash is also present in the nucleus and plays a key role in nuclear organization via its interaction with Lamin Dm0 at the nuclear envelope. Wash and Lamin Dm0 occupy similar genomic regions that overlap with transcriptionally silent chromatin including constitutive heterochromatin. Strikingly, wash mutant and knockdown nuclei exhibit the same abnormal wrinkled morphology observed in diverse laminopathies, including the Hutchinson-Gilford progeria syndrome, and consistent with disruption of the nuclear organization of several sub-nuclear structures including cajal bodies and the chromocenter in salivary glands. We also found that Wash and Lamin knockdown disrupt chromatin accessibility of repressive compartments in agreement with an observed global redistribution of repressive histone modifications. Functional genetic approaches show wash mutants exhibit similar phenotypes to lamin Dm0 mutants, suggesting they participate in similar regulatory networks. Our results reveal a novel role for Wash in modulating nuclear organization via its interaction with the nuclear envelope protein Lamin Dm0. These findings highlight the functional complexity of WASP family proteins and provide new venues to understand their molecular roles in cell biology and disease. DamID chromatin profiling demostrate that Wash binds similar regions to those bound by Lamin Dm0, in particular transcriptional silent chromatin

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.

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Wash regulates lamin-dependent nuclear genome organization [Chromatin Accessibility]

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