Comparison of the domain structures of the WASP and WAVE family proteins from different species. Color coding indicates conserved domains. The percentage amino acid similarity of WH1/EVH1 domains or WHD/SHD domains is shown below each domain. For species abbreviations, see the legend to Figure 2.

Multiple regulatory pathways for N-WASP and WAVE2 activation. (a) N-WASP is autoinhibited in a basal state through the interaction between the GBD/CRIB domain and the VCA region. PIP₂ and GTP-loaded Cdc42 bind to the B and GBD/CRIB domains, respectively, resulting in synergistic activation of N-WASP. Binding of SH3 domains to N-WASP can independently compete with the autoinhibitory interaction, and thus can activate N-WASP. SH3-domain-containing proteins that interact and potentially activate N-WASP include cortactin, WISH, Nck, Grb2, Crk, FBP17, CIP4, Toca1, Abi1, endophilin A, and sorting nexin 9 (not all shown on the diagram). Concurrently, the BAR-domain superfamily proteins bend the membrane. (b) WAVE proteins exist in cells as a heteropentameric protein complex as indicated. WAVE2 has been shown to translocate to the membrane via interactions with phosphatidylinositol-(3,4,5)-triphosphate (PIP₃) and IRSp53. The affinity of WAVE2 for IRSp53 is enhanced when GTP-loaded Rac binds to the RCB/MIM domain of IRSp53. IRSp53 is also able to enhance the ability of WAVE2 to stimulate Arp2/3-mediated actin polymerization [91]. This pathway via IRSp53 is an indirect activation by Rac, as it is suggested that Rac can activate the WAVE complex through direct interaction with Sra1. The direct pathway was shown in a recent paper but needs more experimental evidence to be widely accepted (hence marked by a question mark in the figure).

Evolutionary relationships between the WASP and WAVE family proteins. The phylogeny was inferred using the neighbor-joining method. ClustalW was used to align sequences and perform phylogenetic analysis. Any position containing gaps was excluded from the dataset. Trees were drawn by NJplot [89]. Bootstrap values were calculated over 1,000 iterations and values greater than 50% are shown as percentages next to branches. The bar in each figure indicates the proportion of amino acid differences. (a) The phylogenetic tree based on the alignment of combined sequences of V and C regions. WASP and WAVE sequences were retrieved from the NCBI protein database and the V/WH2 domain for each protein was identified by homology search over the Pfam-A database. C regions were identified according to the previously reported consensus sequence [29]. The sequence to be analyzed was generated by joining the identified V sequence and C sequence. (b) The phylogenic tree based on WH1/EVH1 domain alignment. WH1/EVH1 domains were identified by homology search over the PROSITE database. (c) The phylogenetic tree based on WHD/SHD domain alignment. WHD/SHD domains were identified following the consensus sequence described previously [90]. Species examined are Homo sapiens (Hs), Mus musculus (Mm), Danio rerio (Dr), Drosophila melanogaster (Dm), Caenorhabditis elegans (Ce), Saccharomyces cerevisiae (Sc), Dictyostelium discoideum (Dd) and Arabidopsis thaliana (At). Ensembl protein IDs for the zebrafish sequences used in the analysis are as follows: Dr WASP1, ENSDARP00000039217; Dr WASP2, ENSDARP00000007963; Dr N-WASPa, ENSDARP00000094295; Dr N-WASPb, ENSDARP00000005823; Dr WAVE1, ENSDARP00000079387; Dr WAVE2, ENSDARP00000093195; Dr WAVE3a, ENSDARP00000077123; Dr WAVE3b, ENSDARP00000085962. Two other homologous genes for WAVE were identified in the zebrafish genome, but could not be assigned to homologs of mammalian WAVE1/2/3, so they were omitted from the analysis. These proteins are ENSDARP00000047935 and ENSDARP00000102646.