Abstract
Arfs and Arf GTPase-activating proteins (ArfGAPs) are regulators of membrane trafficking and actin dynamics in mammalian cells. In this study, we identified a primordial Arf, ArfA, and two ArfGAPs (ACAP-A/B) containing BAR, PH, ArfGAP and Ankyrin repeat domains in the eukaryote Dictyostelium discoideum. In vitro, ArfA has similar nucleotide binding properties as mammalian Arfs and, with GTP bound, is a substrate for ACAP-A and B. We also investigated the physiological functions of ACAP-A/B by characterizing cells lacking both ACAP-A and B. Although ACAP-A/B knockout cells showed no defects in cell growth, migration or chemotaxis, they exhibited abnormal actin protrusions and ∼50% reduction in spore yield. We conclude that while ACAP-A/B have a conserved biochemical mechanism and effect on actin organization, their role in migration is not conserved. The absence of an effect on Dictyostelium migration may be due to a specific requirement for ACAPs in mesenchymal migration, which is observed in epithelial cancer cells where most studies of mammalian ArfGAPs were performed.
Highlights
Eukaryotic cells contain complex internal organelles enclosed within membranes and a cytoskeleton, which differentiate them from prokaryotic cells
ACAP-A/B Are Arf GTPase-activating proteins (ArfGAPs) Homologs in Dictyostelium To identify ArfGAPs in Dictyostelium, the ArfGAP domain of the human ASAP1 was used to search for homologous proteins in the Dictyostelium genome database
ACAP-A contains a long (,526 a.a.) carboxyl-terminal tail after the ankyrin repeats, which is characteristic of human ASAPs
Summary
Eukaryotic cells contain complex internal organelles enclosed within membranes and a cytoskeleton, which differentiate them from prokaryotic cells. These organelles provide compartmentalization for various metabolic activities. The cytoskeleton is a network of protein filaments extending throughout the cytoplasm to provide the structural framework of the cell. Remodeling of membranes and the actin cytoskeleton is essential for a variety of eukaryotic cell functions including protein secretion, maintenance of cell morphology and cell movement. Protein machineries that affect and regulate changes in membrane and actin structures have been identified and the coordination of the action of these proteins is being studied. Among them are the Arf family of GTP-binding proteins and the accessory proteins that regulate them, guanine nucleotide exchange factors (GEFs) and GTPase-activating proteins (GAPs) [1,2]
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