Developmental expression ofDrosophilaWiskott‐Aldrich Syndrome family proteins

Abstract

Background Wiskott-Aldrich Syndrome (WASP) family proteins participate in many cellular processes involving rearrangements of the actin cytoskeleton. To the date, four WASP subfamily members have been described in Drosophila: Wash, WASp, SCAR, and Whamy. Wash, WASp, and SCAR are essential during early Drosophila development where they function in orchestrating cytoplasmic events including membrane-cytoskeleton interactions. A mutant for Whamy has not yet been reported. Results We generated monoclonal antibodies that are specific to Drosophila Wash, WASp, SCAR, and Whamy, and use these to describe their spatial and temporal localization patterns. Consistent with the importance of WASP family proteins in flies, we find that Wash, WASp, SCAR, and Whamy are dynamically expressed throughout oogenesis and embryogenesis. For example, we find that Wash accumulates at the oocyte cortex. WASp is highly expressed in the PNS, while SCAR is the most abundantly expressed in the CNS. Whamy exhibits an asymmetric subcellular localization that overlaps with mitochondria and is highly expressed in muscle. Conclusion All four WASP family members show specific expression patterns, some of which reflect their previously known roles and others revealing new potential functions. The monoclonal antibodies developed offer valuable new tools to investigate how WASP family proteins regulate actin cytoskeleton dynamics.

WASH Knockout T Cells Demonstrate Defective Receptor Trafficking, Proliferation, and Effector Function

“…Thus, we generated WASH conditional knockout mice (23) in order to determine the role of WASH in thymocyte development and peripheral T cell function. We found that total WASH knockout results in embryonic lethality around day E7.5 (T. S. Gomez and D. D. Billadeau, data not shown), which is consistent with studies in Drosophila, which have shown that WASH is expressed throughout embryogenesis and that WASH KO Drosophila are not viable (13,31). Therefore, we crossed WASH flox/ϩ mice with CD4 Cre mice to generate CD4 Cre WASH flox/flox conditional knockout (WASHout) mice, which allows for specific Cre-mediated excision of WASH in T cells in late thymic development (32).…”

Drosophila WASH is required for integrin-mediated cell adhesion, cell motility and lysosomal neutralization

“…Thus, redundant or compensatory pathways might mask WASH function in developing eggs. Previous expression data in wild-type flies revealed an accumulation at the oocyte cortex in addition to an overall cytoplasmic expression in all nurse and somatic follicle cells throughout oogenesis (Rodriguez-Mesa et al, 2012). The protein-Aaffinity-purified monoclonal antibody (mAb P3H3; Rodriguez-Mesa et al, 2012) used in this study specifically recognizes WASH in immunoblots (Fig.…”

Coordinated autoinhibition of F-BAR domain membrane binding and WASp activation by Nervous Wreck

“…The Arp2/3 complex was purchased from Cytoskeleton, Inc. Coprecipitation assays with GST-tagged proteins were conducted as described previously (16). Pellets and supernatants were denatured in Laemmli sample buffer and fractionated by SDS/PAGE, followed by immunoblotting with α-WASp antibody P5E1 (41).…”