Modelling of human Wiskott-Aldrich syndrome protein mutants in zebrafish larvae using in vivo live imaging

Abstract

Wiskott-Aldrich syndrome (WAS) and X-linked neutropenia (XLN) are immunodeficiencies in which the function of several haematopoietic cell lineages is perturbed as a result of mutations in the actin regulator WASp. From in vitro cell biology experiments, and biochemical and structural approaches, we know much about the functional domains of WASp and how WASp might regulate the dynamic actin cytoskeleton downstream of activators such as Cdc42, but in vivo experiments are much more challenging. In patients, there is a correlation between clinical disease and genotype, with severe reductions in WASp expression or function associating with complex multilineage immunodeficiency, whereas specific mutations that cause constitutive activation of WASp result in congenital neutropenia. Here, we take advantage of the genetic tractability and translucency of zebrafish larvae to first characterise how a null mutant in zfWASp influences the behaviour of neutrophils and macrophages in response to tissue damage and to clearance of infections. We then use this mutant background to study how leukocyte lineage-specific transgenic replacement with humanWASp variants (including normal wild type and point mutations that either fail to bind Cdc42 or cannot be phosphorylated, and a constitutively active mutant equivalent to that seen in XLN patients) alter the capacity for generation of neutrophils, their chemotactic response to wounds and the phagocytic clearance capacity of macrophages. This model provides a unique insight into WASp-related immunodeficiency at both a cellular and whole organism level. © 2013. Published by The Company of Biologists Ltd.