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The FEBS Journal Oct 2017Intrinsically disordered proteins (IDPs) play key roles in signaling and regulation. Many IDPs undergo folding upon binding to their targets. We have proposed that...
Intrinsically disordered proteins (IDPs) play key roles in signaling and regulation. Many IDPs undergo folding upon binding to their targets. We have proposed that coupled folding and binding of IDPs generally follow a dock-and-coalesce mechanism, whereby a segment of the IDP, through diffusion, docks to its cognate subsite and, subsequently, the remaining segments coalesce around their subsites. Here, by a combination of experiment and computation, we determined the precise form of dock-and-coalesce operating in the association between the intrinsically disordered GTPase-binding domain (GBD) of the Wiskott-Aldrich Syndrome protein and the Cdc42 GTPase. The association rate constants (k ) were measured by stopped-flow fluorescence under various solvent conditions. k reached 10 m ·s at physiological ionic strength and had a strong salt dependence, suggesting that an electrostatically enhanced, diffusion-controlled docking step may be rate limiting. Our computation, based on the transient-complex theory, identified the N-terminal basic region of the GBD as the docking segment. However, several other changes in solvent conditions provided strong evidence that the coalescing step also contributed to determining the magnitude of k . Addition of glucose and trifluoroethanol and an increase in temperature all produced experimental k values much higher than expected from the effects on the docking rate alone. Conversely, addition of urea led to k values much lower than expected if only the docking rate was affected. These results all pointed to k being approximately two-thirds of the docking rate constant under physiological solvent conditions.
Topics: Humans; Intrinsically Disordered Proteins; Protein Binding; Wiskott-Aldrich Syndrome Protein; cdc42 GTP-Binding Protein
PubMed: 28805312
DOI: 10.1111/febs.14197 -
Frontiers in Immunology 2021Wiskott-Aldrich Syndrome, WAS/WAVE, is a rare, X-linked immune-deficiency disease caused by mutations in the gene, which together with its homolog, N-, regulates actin...
Wiskott-Aldrich Syndrome, WAS/WAVE, is a rare, X-linked immune-deficiency disease caused by mutations in the gene, which together with its homolog, N-, regulates actin cytoskeleton remodeling and cell motility. WAS patients suffer from microthrombocytopenia, characterized by a diminished number and size of platelets, though the underlying mechanism is not fully understood. Here, we identified FLI1 as a direct transcriptional regulator of and its binding partner . Depletion of either or in human erythroleukemic cells accelerated cell proliferation, suggesting tumor suppressor function of both genes in leukemia. Depletion of also led to a significant reduction in the percentage of CD41 and CD61 positive cells, which mark committed megakaryocytes. RNAseq analysis revealed common changes in megakaryocytic gene expression following FLI1 or WASP knockdown. However, in contrast to FLI1, WASP depletion did not alter expression of late-stage platelet-inducing genes. N-WASP was not regulated by FLI1, yet its silencing also reduced the percentage of CD41+ and CD61+ megakaryocytes. Moreover, combined knockdown of WASP and N-WASP further suppressed megakaryocyte differentiation, indicating a major cooperation of these related genes in controlling megakaryocytic cell fate. However, unlike WASP/WIP, N-WASP loss suppressed leukemic cell proliferation. WASP, WIP and N-WASP depletion led to induction of FLI1 expression, mediated by GATA1, and this may mitigate the severity of platelet deficiency in WAS patients. Together, these results uncover a crucial role for FLI1 in megakaryocyte differentiation, implicating this transcription factor in regulating microthrombocytopenia associated with Wiskott-Aldrich syndrome.
Topics: Animals; Base Sequence; Biomarkers; Cell Line; Chromatin Immunoprecipitation Sequencing; Cytoskeletal Proteins; Disease Models, Animal; Disease Susceptibility; Gene Expression Regulation; Humans; Intracellular Signaling Peptides and Proteins; Leukemia; Mice; Mice, Knockout; Promoter Regions, Genetic; Proto-Oncogene Protein c-fli-1; Signal Transduction; Thrombopoiesis; Wiskott-Aldrich Syndrome; Wiskott-Aldrich Syndrome Protein
PubMed: 33717090
DOI: 10.3389/fimmu.2021.607836 -
Frontiers in Immunology 2022Wiskott-Aldrich Syndrome (WAS) is characterized by recurrent infections, thrombocytopenia, and eczema. Here, we show that WASp-deficient mice on a BALB/c background have...
Wiskott-Aldrich Syndrome (WAS) is characterized by recurrent infections, thrombocytopenia, and eczema. Here, we show that WASp-deficient mice on a BALB/c background have dysregulated cutaneous immune homeostasis with increased leukocyte accumulation in the skin, 1 week after birth. Increased cutaneous inflammation was associated with epithelial abnormalities, namely, altered keratinization, abnormal epidermal tight junctional morphology and increased trans-epidermal water loss; consistent with epidermal barrier dysfunction. Immune and physical barrier disruption was accompanied by progressive skin dysbiosis, highlighting the functional significance of the disrupted cutaneous homeostasis. Interestingly, the dysregulated immunity in the skin preceded the systemic elevation in IgE and lymphocytic infiltration of the colonic lamina propria associated with WASp deficiency. Mechanistically, the enhanced immune cell accumulation in the skin was lymphocyte dependent. Elevated levels of both Type 2 (IL-4, IL-5) and Type 17 (IL-17, IL-22, IL-23) cytokines were present in the skin, as well as the 'itch' factor IL-31. Unexpectedly, the canonical WAS-associated cytokine IL-4 did not play a role in the immune dysfunction. Instead, IL-17 was critical for skin immune infiltration and elevation of both Type 2 and Type 17 cytokines. Our findings reveal a previously unrecognized IL-17-dependent breakdown in immune homeostasis and cutaneous barrier integrity in the absence of WASp, targeting of which may provide new therapeutic possibilities for the treatment of skin pathologies in WAS patients.
Topics: Animals; Cytokines; Homeostasis; Interleukin-17; Interleukin-4; Mice; Mice, Knockout; Wiskott-Aldrich Syndrome; Wiskott-Aldrich Syndrome Protein
PubMed: 35265075
DOI: 10.3389/fimmu.2022.817427 -
Experimental and Therapeutic Medicine Jan 2023Wiskott-Aldrich syndrome (WAS) is a rare X-linked primary immunodeficiency characterized by microthrombocytopenia, eczema, recurrent infection and increased incidence of...
Wiskott-Aldrich syndrome (WAS) is a rare X-linked primary immunodeficiency characterized by microthrombocytopenia, eczema, recurrent infection and increased incidence of autoimmune disorders and malignancy. WAS is caused by mutations in the gene, which is expressed exclusively in hematopoietic cells; the spleen serves an important role in hematopoiesis and red blood cell clearance. However, to the best of our knowledge, detailed comparative analysis of the spleen between WASp-knockout (WAS-KO) and wild-type (WT) mice, particularly at the transcriptomic level, have not been reported. The present study investigated the differences in the transcriptomes of spleen tissue of 10-week-old WAS-KO mice. Comparison of the gene expression profiles of WAS-KO and WT mice revealed 1,964 differentially expressed genes (DEGs). Among these genes, 996 DEGs were upregulated and 968 were downregulated in WAS-KO mice. To determine the functions of DEGs, Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analyses were performed for significantly upregulated and downregulated DEGs. The results showed that the levels of cell senescence and apoptosis-associated genes were increased, antigen processing and presentation mechanisms involved in the immune response were damaged and signal transduction processes were impaired in the spleen of WAS-KO mice. Thus, gene deletion may lead to anemia and hemolysis-associated disease, primarily due to increased osmotic fragility of red blood cells, low hemoglobin and increased bilirubin levels and serum ferritin. These results indicated that senescence and apoptosis of blood cells also play an important role in the occurrence of WAS. Therefore, the present findings provide a theoretical basis for further study to improve the treatment of WAS.
PubMed: 36605531
DOI: 10.3892/etm.2022.11763 -
Nature Communications Nov 2017Dysregulation of autophagy and inflammasome activity contributes to the development of auto-inflammatory diseases. Emerging evidence highlights the importance of the...
Dysregulation of autophagy and inflammasome activity contributes to the development of auto-inflammatory diseases. Emerging evidence highlights the importance of the actin cytoskeleton in modulating inflammatory responses. Here we show that deficiency of Wiskott-Aldrich syndrome protein (WASp), which signals to the actin cytoskeleton, modulates autophagy and inflammasome function. In a model of sterile inflammation utilizing TLR4 ligation followed by ATP or nigericin treatment, inflammasome activation is enhanced in monocytes from WAS patients and in WAS-knockout mouse dendritic cells. In ex vivo models of enteropathogenic Escherichia coli and Shigella flexneri infection, WASp deficiency causes defective bacterial clearance, excessive inflammasome activation and host cell death that are associated with dysregulated septin cage-like formation, impaired autophagic p62/LC3 recruitment and defective formation of canonical autophagosomes. Taken together, we propose that dysregulation of autophagy and inflammasome activities contribute to the autoinflammatory manifestations of WAS, thereby identifying potential targets for therapeutic intervention.
Topics: Actin Cytoskeleton; Animals; Autophagy; Bacterial Load; Cell Line, Tumor; Dendritic Cells; Enteropathogenic Escherichia coli; Humans; Immunity, Innate; Inflammasomes; Interferon Type I; Mice; Mice, Inbred C57BL; Mice, Knockout; Monocytes; NLR Family, Pyrin Domain-Containing 3 Protein; Nigericin; Septins; Shigella flexneri; THP-1 Cells; Toll-Like Receptor 4; Wiskott-Aldrich Syndrome; Wiskott-Aldrich Syndrome Protein
PubMed: 29146903
DOI: 10.1038/s41467-017-01676-0 -
FASEB Journal : Official Publication of... Jul 2016TGF-β1 induces an increase in paracellular permeability and actin stress fiber formation in lung microvascular endothelial and alveolar epithelial cells via small Rho...
TGF-β1 induces an increase in paracellular permeability and actin stress fiber formation in lung microvascular endothelial and alveolar epithelial cells via small Rho GTPase. The molecular mechanism involved is not fully understood. Neuronal Wiskott-Aldrich syndrome protein (N-WASP) has an essential role in actin structure dynamics. We hypothesized that N-WASP plays a critical role in these TGF-β1-induced responses. In these cell monolayers, we demonstrated that N-WASP down-regulation by short hairpin RNA prevented TGF-β1-mediated disruption of the cortical actin structure, actin stress filament formation, and increased permeability. Furthermore, N-WASP down-regulation blocked TGF-β1 activation mediated by IL-1β in alveolar epithelial cells, which requires actin stress fiber formation. Control short hairpin RNA had no effect on these TGF-β1-induced responses. TGF-β1-induced phosphorylation of Y256 of N-WASP via activation of small Rho GTPase and focal adhesion kinase mediates TGF-β1-induced paracellular permeability and actin cytoskeleton dynamics. In vivo, compared with controls, N-WASP down-regulation increases survival and prevents lung edema in mice induced by bleomycin exposure-a lung injury model in which TGF-β1 plays a critical role. Our data indicate that N-WASP plays a crucial role in the development of TGF-β1-mediated acute lung injury by promoting pulmonary edema via regulation of actin cytoskeleton dynamics.-Wagener, B. M., Hu, M., Zheng, A., Zhao, X., Che, P., Brandon, A., Anjum, N., Snapper, S., Creighton, J., Guan, J.-L., Han, Q., Cai, G.-Q., Han, X., Pittet, J.-F., Ding, Q. Neuronal Wiskott-Aldrich syndrome protein regulates TGF-β1-mediated lung vascular permeability.
Topics: Animals; Bleomycin; Capillary Permeability; Cells, Cultured; Endothelial Cells; Gene Expression Regulation; Lung; Lung Injury; Mice; Neurons; Rats; Transforming Growth Factor beta1; Wiskott-Aldrich Syndrome Protein, Neuronal
PubMed: 27025963
DOI: 10.1096/fj.201600102R -
British Journal of Haematology Jan 2021Wiskott-Aldrich syndrome (WAS) is a life-threatening primary immunodeficiency associated with bleeding of variable severity due to thrombocytopenia. Correction of the... (Observational Study)
Observational Study
Wiskott-Aldrich syndrome (WAS) is a life-threatening primary immunodeficiency associated with bleeding of variable severity due to thrombocytopenia. Correction of the thrombocytopenia is of paramount importance for most WAS patients. We report a retrospective analysis of the safety and efficacy of romiplostim treatment in reducing thrombocytopenia and bleeding tendency in 67 children (median age 1·3 years) with genetically confirmed WAS, followed in eight months (range, 1-12 months). Complete or partial primary responses regarding platelet counts were observed in 22 (33%) and 18 (27%) subjects, respectively. Yet, even in the non-responder group, the risk of haemorrhagic events decreased significantly, to 21%, after the first month of treatment. The responses tended to be durable and stable over time, with no significant fluctuations in platelets counts. The results of this retrospective study of a large cohort of WAS patients demonstrates that romiplostim can be used to increase platelet counts and reduce the risks of life-threatening bleeding in WAS patients awaiting haematopoietic stem cell transplantation or forgoing the procedure for various reasons.
Topics: Adolescent; Child; Child, Preschool; Hemorrhage; Humans; Infant; Platelet Count; Receptors, Fc; Recombinant Fusion Proteins; Retrospective Studies; Thrombocytopenia; Thrombopoietin; Treatment Outcome; Wiskott-Aldrich Syndrome
PubMed: 33131064
DOI: 10.1111/bjh.17174 -
Nucleus (Austin, Tex.) 2015Actin and proteins that regulate its dynamics or interactions have well-established roles in the cytoplasm where they function as key components of the cytoskeleton to...
Actin and proteins that regulate its dynamics or interactions have well-established roles in the cytoplasm where they function as key components of the cytoskeleton to control diverse processes, including cellular infrastructure, cellular motility, cell signaling, and vesicle transport. Recent work has also uncovered roles for actin and its regulatory proteins in the nucleus, primarily in mechanisms governing gene expression. The Wiskott Aldrich Syndrome (WAS) family of proteins, comprising the WASP/N-WASP, SCAR/WAVE, WHAMM/JMY/WHAMY, and WASH subfamilies, function in the cytoplasm where they activate the Arp2/3 complex to form branched actin filaments. WAS proteins are present in the nucleus and have been implicated as transcriptional regulators. We found that Drosophila Wash, in addition to transcriptional effects, is involved in global nuclear architecture. Here we summarize the regulation and function of nuclear WAS proteins, and highlight how our work with Wash expands the possibilities for the functions of these proteins in the nucleus.
Topics: Animals; Cell Nucleus; Drosophila Proteins; Female; Lamins; Male; Vesicular Transport Proteins
PubMed: 26305109
DOI: 10.1080/19491034.2015.1086051 -
Turkish Journal of Haematology :... Nov 2020Wiskott-Aldrich syndrome (WAS) is an X-linked primary immune deficiency characterized by microthrombocytopenia, eczema, and recurrent infections. We aimed to evaluate...
OBJECTIVE
Wiskott-Aldrich syndrome (WAS) is an X-linked primary immune deficiency characterized by microthrombocytopenia, eczema, and recurrent infections. We aimed to evaluate the clinical features and outcomes of a WAS cohort.
MATERIALS AND METHODS
We retrospectively evaluated the clinical courses, immunological features, treatments, and outcomes in a total of 23 WAS patients together with data related to 11 transplanted cases among them between 1982 and 2019.
RESULTS
Before admission, 11 patients (48%) were misdiagnosed with immune thrombocytopenia. WAS scores were mostly 4 or 5. Eleven patients were transplanted and they had an overall survival rate of 100% during a median follow-up period of 8.5 years (range: 8 months to 20 years). Five patients who were not transplanted died at a median of 7 years (range: 2-26 years). Nontransplanted patients had high morbidity due to organ damage, mostly caused by autoimmunity, bleeding, and infections. Two novel mutations were also defined.
CONCLUSION
All male babies with microthrombocytopenia should be evaluated for WAS. Hematopoietic stem cell transplantation should be performed at the earliest age with the best possible donors.
Topics: Adolescent; Biomarkers; Child; Child, Preschool; Combined Modality Therapy; Diagnosis, Differential; Disease Management; Disease Susceptibility; Female; Hematopoietic Stem Cell Transplantation; Humans; Immunoglobulins, Intravenous; Infant; Infant, Newborn; Male; Patient Outcome Assessment; Phenotype; Prognosis; Reinfection; Symptom Assessment; Treatment Outcome; Wiskott-Aldrich Syndrome; Young Adult
PubMed: 32812413
DOI: 10.4274/tjh.galenos.2020.2020.0334 -
The Journal of Allergy and Clinical... Jan 2020Wiskott-Aldrich syndrome (WAS) is an X-linked primary immune deficiency disorder resulting from Wiskott-Aldrich syndrome protein (WASp) deficiency. Lymphocytes from...
BACKGROUND
Wiskott-Aldrich syndrome (WAS) is an X-linked primary immune deficiency disorder resulting from Wiskott-Aldrich syndrome protein (WASp) deficiency. Lymphocytes from patients with WAS manifest increased DNA damage and lymphopenia from cell death, yet how WASp influences DNA damage-linked cell survival is unknown. A recently described mechanism promoting cell survival after ionizing radiation (IR)-induced DNA damage involves fragmentation and dispersal of the Golgi apparatus, known as the Golgi-dispersal response (GDR), which uses the Golgi phosphoprotein 3 (GOLPH3)-DNA-dependent protein kinase (DNA-PK)-myosin XVIIIA-F-actin signaling pathway.
OBJECTIVE
We sought to define WASp's role in the DNA damage-induced GDR and its disruption as a contributor to the development of radiosensitivity-linked immunodeficiency in patients with WAS.
METHODS
In human T and B-cell culture systems, DNA damage-induced GDR elicited by IR or radiomimetic chemotherapy was monitored in the presence or absence of WASp or GOLPH3 alone or both together.
RESULTS
WASp deficiency completely prevents the development of IR-induced GDR in human T and B cells, despite the high DNA damage load. Loss of WASp impedes nuclear translocation of GOLPH3 and its colocalization with the DNA-dependent protein kinase catalytic subunit (DNA-PKcs). Surprisingly, however, depletion of GOLPH3 alone or depolymerization of F-actin in WASp-sufficient T cells still allows development of robust GDR, suggesting that WASp, but not GOLPH3, is essential for GDR and cell survival after IR-induced DNA-damage in human lymphocytes.
CONCLUSION
The study identifies WASp as a novel effector of the nucleus-to-Golgi cell-survival pathway triggered by IR-induced DNA damage in cells of the hematolymphoid lineage and proposes an impaired GDR as a new cause for development of a "radiosensitive" form of immune dysregulation in patients with WAS.
Topics: B-Lymphocytes; DNA Damage; DNA-Activated Protein Kinase; Golgi Apparatus; Humans; Membrane Proteins; Signal Transduction; T-Lymphocytes; Wiskott-Aldrich Syndrome; Wiskott-Aldrich Syndrome Protein Family
PubMed: 31604087
DOI: 10.1016/j.jaci.2019.09.026