Malaria parasites undergo two rounding-up transformations in their life cycle: the ookinete-to-oocyst transformation in the mosquito midgut, and the sporozoite-to-EEF (exo-erythrocytic form) differentiation in the host hepatocyte. Both events are characterized by the loss of polarity, implying that cytoskeletal reorganization is involved. In other eukaryotes, regulation of the actin skeleton is fundamental to subcellular remodeling. Although filamentous actin is well known to be involved in the motility of api-complexan parasites, its participation in their morphological regulation is still largely unexplored. Here we describe the fundamental role of Actin depolymerization factor 2 (ADF2), a vector-stage-specific ADF isoform, in morphological changes accompanying the parasite life cycle. Among protozoan parasites, Plasmodium is unique in having two actin and two ADF genes. Disruption of the ADF2 gene in the rodent malaria parasite P. berghei had no effect on ookinete development or its subsequent invasion of the midgut. However, both the ookinete-to-oocyst and sporozoite-to-EEF transformations showed significant defects. These results indicated that Plasmodium ADF2 plays a pivotal role in transformation in the malaria parasite life cycle. (C) 2010 Elsevier Inc. All rights reserved.