The Diaphanous-related formin dDia1 is required for highly directional phototaxis and formation of properly sized fruiting bodies in Dictyostelium

Diaphanous-related formins (DRFs) act as downstream effectors of Rho family GTPases and drive the formation and elongation of linear actin filaments in various cellular processes. Here we analyzed the DRF dDia1 from Dictyostelium cells. The biochemical characterization of recombinant dDia1-FH1FH2 by bulk polymerization assays and single filament TIRF microscopy revealed that dDia1 is a rather weak nucleator. Addition of any of the three Dictyostelium profilin isoforms, however, markedly accelerated formin-mediated actin filament barbed end elongation in TIRF assays. Interestingly, filament elongation was significantly faster in presence of DdPFN I (profilin I) when compared to the other two isoforms, suggesting selectivity of dDia1 for DdPFN I. Additionally, we frequently observed dissociation of the formin from growing barbed ends. These findings are consistent with dilution-induced depolymerization assays in presence of dDia1-FH1FH2 showing that dDia1 is a weak capper in comparison with heterodimeric capping protein. To study the physiological role of this formin, we created cell lines lacking dDia1 or overexpressing GFP-tagged dDia1. Of note, constitutively active dDia1 accumulated homogenously in the entire pseudopod suggesting that it controls microfilament architecture to regulate cell migration. Comparison of wild type and dDia1 -null cells in random cell migration and chemotaxis toward a cAMP gradient revealed no major differences. By contrast, phototaxis of dDia1-deficient cells during the multicellular stage was markedly impaired. While wild type slugs moved with high directionality toward the light source, the trails of dDia1 -null slugs displayed a characteristic V-shaped profile and deviated in angles between 50 and 60 from the path of the incident light. Possibly in conjunction with this defect, dDia1 -null cells also formed substantially smaller fruiting bodies. These findings demonstrate dDia1 to be critically involved in collective cell migration during terminal differentiation. (C) 2013 Elsevier GmbH. All rights reserved.