Structure of the light chain-binding domain of myosin V

Myosin V is a double-headed molecular motor involved in organelle transport. Two distinctive features of this motor, processivity and the ability to take extended linear steps of ≈36 nm along the actin helical track, depend on its unusually long light chain-binding domain (LCBD). The LCBD of myosin V consists of six tandem IQ motifs, which constitute the binding sites for calmodulin (CaM) and CaM-like light chains. Here, we report the 2-Å resolution crystal structure of myosin light chain 1 (Mlc1p) bound to the IQ2–IQ3 fragment of Myo2p, a myosin V from Saccharomyces cerevisiae. This structure, combined with FRET distance measurements between probes in various CaM–IQ complexes, comparative sequence analysis, and the previously determined structures of Mlc1p-IQ2 and Mlc1p-IQ4, allowed building a model of the LCBD of myosin V. The IQs of myosin V are distributed into three pairs. There appear to be specific cooperative interactions between light chains within each IQ pair, but little or no interaction between pairs, providing flexibility at their junctions. The second and third IQ pairs each present a light chain, whether CaM or a CaM-related molecule, bound in a noncanonical extended conformation in which the N-lobe does not interact with the IQ motif. The resulting free N-lobes may engage in protein–protein interactions. The extended conformation is characteristic of the single IQ of myosin VI and is common throughout the myosin superfamily. The model points to a prominent role of the LCBD in the function, regulation, and molecular interactions of myosin V

Annexin A2-dependent polymerization of actin mediates endosome biogenesis

Early endosomes give rise to multivesicular intermediates during transport toward late endosomes. Much progress has been made in understanding the sorting of receptors into these intermediates, but the mechanisms responsible for their biogenesis remain unclear. Here, we report that F-actin is necessary for transport beyond early endosomes and endosome formation. We found that endosomes captured by actin cables were essentially stationary, but early endosomes also exhibited patches of F-actin and facilitated selective F-actin nucleation and polymerization. Our data show that nucleation of actin patches by early endosomes is strictly dependent on annexin A2, a protein involved in early-to-late endosome transport. It also requires the actin nucleation factor Spire1 and involves Arp2/3, which is needed for filament branching. We conclude that actin patches are nucleated on early endosomes via annexin A2 and Spire1, and that these patches control endosome biogenesis, presumably by driving the membrane remodeling process