| Abstract |
PCH family proteins are fundamentally important proteins, linking membrane curvature events with cytoskeletal reorganisation. One group, the MEGAPs (also called srGAPs and WRPs) contain RhoGAP domains in addition to the F-BAR domain. We disrupted MEGAP1 and MEGAP2 in Dictyostelium both singly and in combination. We found a strong cytoskeletal phenotype in MEGAP1(-) cells and a subtle phototaxis defect in MEGAP2(-) slugs. MEGAP1(-)/2(-) cells have an overabundance of filopodia and slug motility and function are affected. The most dramatic changes, however, are on contractile vacuoles. MEGAP1(-)/2(-) cells empty their contractile vacuoles less efficiently than normal and consequently have three times the usual number. GFP-tagged MEGAP1 localises to tubules of the contractile vacuole network and when vacuoles start to empty they recruit cytosolic GFP-MEGAP1. Mutants in the Saccharomyces homologues RGD1 and RGD2 also show abnormal vacuoles, implying that this role is conserved. Thus, MEGAP is an important regulator of the contractile vacuole network, and we propose that tubulation of the contractile vacuole by MEGAP1 represents a novel mechanism for driving vacuole emptying.
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