The crescent‐shaped BAR (Bin/Amphiphysin/Rvs‐homology) domain dimer is a versatile protein module that senses and generates positive membrane curvature. The BAR domain dimer of human endophilin‐A1, solved at 3.1 Å, has a unique structure consisting of a pair of helix–loop appendages sprouting out from the crescent. The appendage's short helices form a hydrophobic ridge, which runs across the concave surface at its center. Examining liposome binding and tubulation in vitro using purified BAR domain and its mutants indicated that the ridge penetrates into the membrane bilayer and enhances liposome tubulation. BAR domain‐expressing cells exhibited marked plasma membrane tubulation in vivo. Furthermore, a swinging‐arm mutant lost liposome tubulation activity yet retaining liposome binding. These data suggested that the rigid crescent dimer shape is crucial for the tubulation. We here propose that the BAR domain drives membrane curvature by coordinate action of the crescent's scaffold mechanism and the ridge's membrane insertion in addition to membrane binding via amino‐terminal amphipathic helix.

Insulin receptor tyrosine kinase substrate p53 (IRSp53) has been identified as an SH3 domain-containing adaptor that links Rac1 with a Wiskott-Aldrich syndrome family verprolin-homologous protein 2 (WAVE2) to induce lamellipodia or Cdc42 with Mena to induce filopodia. The recruitment of these SH3-binding partners by IRSp53 is thought to be crucial for F-actin rearrangements. Here, we show that the N-terminal predicted helical stretch of 250 amino acids of IRSp53 is an evolutionarily conserved F-actin bundling domain involved in filopodium formation. Five proteins including IRSp53 and missing in metastasis (MIM) protein share this unique domain and are highly conserved in vertebrates. We named the conserved domain IRSp53/MIM homology domain (IMD). The IMD has domain relatives in invertebrates but does not show obvious homology to any known actin interacting proteins. The IMD alone, derived from either IRSp53 or MIM, induced filopodia in HeLa cells and the formation of tightly packed parallel F-actin bundles in vitro. These results suggest that IRSp53 and MIM belong to a novel actin bundling protein family. Furthermore, we found that filopodium-inducing IMD activity in the full-length IRSp53 was regulated by active Cdc42 and Rac1. The SH3 domain was not necessary for IMD-induced filopodium formation. Our results indicate that IRSp53, when activated by small GTPases, participates in F-actin reorganization not only in an SH3-dependent manner but also in a manner dependent on the activity of the IMD.Insulin receptor tyrosine kinase substrate p53 (IRSp53), 1 also known as brain-specific angiogenesis inhibitor 1-associated protein 2, is a multifunctional adaptor protein enriched in the central nervous system (1-3). The protein contains a unique N-terminal 250-amino acid stretch, a half-Cdc42/Rac interactive binding (CRIB) motif, a proline-rich domain, a Src homology 3 (SH3) domain, and a WW domain-binding motif (WWB). IRSp53 is directly regulated by Rho family small GTPases Rac1 and Cdc42 and provides a molecular link between these GTPases and the actin cytoskeleton regulators WiskottAldrich syndrome protein (WASP) family verprolin homologous protein 2 (WAVE2) and mammalian enabled (Mena), which are involved in the formation of lamellipodia (4, 5) and filopodia (6, 7). Active Cdc42 binds to the half-CRIB motif (6, 7), whereas Rac1 binds to the unique N-terminal domain (8). The association of Rac1 or Cdc42 is proposed to liberate the C-terminal SH3 domain masked intramolecularly by its N terminus, thereby allowing the SH3 domain to interact with its binding partners (4, 7, 9). Thus, the SH3 domain is thought to be essential for IRSp53-mediated actin reorganization. However, the N-terminal half of IRSp53 lacking the SH3 domain was reported to induce neurite outgrowth in a neuroblastoma cell line (6) and filopodia in B16 melanoma cells (10), suggesting that IRSp53 promotes actin reorganization independently of SH3 domain-mediated intermolecular interactions.Recently, a novel monomeric actin-binding protein, missing in met...

During cell division, many animal cells transform into a spherical shape and assemble a contractile ring composed of actin filaments and myosin motors at the equator to separate the cell body into two. Although actomyosin regulatory proteins are spatio-temporally controlled during cytokinesis, the direct contribution of cell shape and actomyosin activity to the contractile ring assembly remains unclear. Here, we demonstrated in vitro that actin polymerization inside cell-sized spherical droplets induced the spontaneous formation of single ring-shaped actin bundles in the presence of bundling factors. Despite a lack of spatial regulatory signals, the rings always assembled at the equator to minimize the elastic energy of the bundles. Myosin promoted ring formation by the dynamic remodelling of actin networks, and an increase in the effective concentration of myosin triggered ring contraction. These results will help us understand how animal cells coordinate cell shape and actomyosin activities to direct cytokinesis.

CorrectionsBIOPHYSICS. For the article ''Load-dependent ADP binding to myosins V and VI: Implications for subunit coordination and function,'' by Yusuke Oguchi,

¡Vive la différence! In cardiac contraction, the reduction in sarcomere length—rather than length itself—determines contractile force.