1. Disease-associated missense mutations in the EVH1 domain disrupt intrinsic WASp function causing dysregulated actin dynamics and impaired dendritic cell migration.
- Author
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Worth AJ, Metelo J, Bouma G, Moulding D, Fritzsche M, Vernay B, Charras G, Cory GO, Thrasher AJ, and Burns SO
- Subjects
- Actins metabolism, Animals, Biopolymers, Carrier Proteins metabolism, Cell Movement, Cells, Cultured, Cytoskeletal Proteins, Dendritic Cells metabolism, Humans, Mice, Mice, Inbred C57BL, Mice, Knockout, Mice, Transgenic, Phosphorylation, Polymerization, Protein Interaction Mapping, Protein Processing, Post-Translational, Protein Stability, Protein Structure, Tertiary, Pseudopodia pathology, Recombinant Fusion Proteins physiology, Sequence Deletion, Specific Pathogen-Free Organisms, Wiskott-Aldrich Syndrome Protein chemistry, Wiskott-Aldrich Syndrome Protein deficiency, Wiskott-Aldrich Syndrome Protein metabolism, Wiskott-Aldrich Syndrome Protein physiology, Dendritic Cells pathology, Mutation, Missense, Wiskott-Aldrich Syndrome Protein genetics
- Abstract
Wiskott Aldrich syndrome (WAS), an X-linked immunodeficiency, results from loss-of-function mutations in the human hematopoietic cytoskeletal regulator gene WAS. Many missense mutations in the Ena Vasp homology1 (EVH1) domain preserve low-level WAS protein (WASp) expression and confer a milder clinical phenotype. Although disrupted binding to WASp-interacting protein (WIP) leads to enhanced WASp degradation in vivo, the intrinsic function of EVH1-mutated WASp is poorly understood. In the present study, we show that, despite mediating enhanced actin polymerization compared with wild-type WASp in vitro, EVH1 missense mutated proteins did not support full biologic function in cells, even when levels were restored by forced overexpression. Podosome assembly was aberrant and associated with dysregulated lamellipodia formation and impaired persistence of migration. At sites of residual podosome-associated actin polymerization, localization of EVH1-mutated proteins was preserved even after deletion of the entire domain, implying that WIP-WASp complex formation is not absolutely required for WASp localization. However, retention of mutant proteins in podosomes was significantly impaired and associated with reduced levels of WASp tyrosine phosphorylation. Our results indicate that the EVH1 domain is important not only for WASp stability, but also for intrinsic biologic activity in vivo.
- Published
- 2013
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