Brain Myosin V Is a Synaptic Vesicle-associated Motor Protein: Evidence for a Ca2+-dependent Interaction with the Synaptobrevin–Synaptophysin Complex

Abstract

Brain myosin V is a member of a widely distributed class of unconventional myosins that may be of central importance to organelle trafficking in all eukaryotic cells. Molecular constituents that target this molecular motor to organelles have not been previously identified. Using a combination of immunopurification, extraction, cross-linking, and coprecipitation assays, we demonstrate that the tail domain of brain myosin V forms a stable complex with the synaptic vesicle membrane proteins, synaptobrevin II and synaptophysin. While myosin V was principally bound to synaptic vesicles during rest, this putative transport complex was promptly disassembled upon the depolarization-induced entry of Ca2+ into intact nerve endings. Coimmunoprecipitation assays further indicate that Ca2+ disrupts the in vitro binding of synaptobrevin II to synaptophysin in the presence but not in the absence of Mg2+. We conclude that hydrophilic forces reversibly couple the myosin V tail to a biochemically defined class of organelles in brain nerve terminals.

Myosin-Va Regulates Exocytosis through the Submicromolar Ca2+-dependent Binding of Syntaxin-1A

“…In the current studies, we found a novel interaction between myosin-Va, which is present on cortical synaptic vesicles (Prekeris and Terrian, 1997;Bridgman, 1999) and syntaxin-1A, a t-SNARE that participates in exocytosis (Duman and Forte, 2003;Li and Chin, 2003), in presence of micromolar levels of Ca 2ϩ . We also found that this unique interaction, linked to Ca 2ϩ -dependent release of CaM from the neck region of myosin-Va, is involved in Ca 2ϩ -regulated exocytosis.…”

“…In addition, Myo2p, a yeast homologue of myosin-Va, directs intracellular transport during secretion and budding through interactions with other proteins (Matsui, 2003). However, the roles of myosin-Va in secretion and Ca 2ϩ -regulated exocytosis are not as clear, probably because the myosin-Va-interacting molecules have not been identified in neurons (Reck-Peterson et al, 2000;Matsui, 2003).In the current studies, we found a novel interaction between myosin-Va, which is present on cortical synaptic vesicles (Prekeris and Terrian, 1997;Bridgman, 1999) and syntaxin-1A, a t-SNARE that participates in exocytosis (Duman and Forte, 2003;Li and Chin, 2003), in presence of micromolar levels of Ca 2ϩ . We also found that this unique interaction, linked to Ca 2ϩ -dependent release of CaM from the neck region of myosin-Va, is involved in Ca 2ϩ -regulated exocytosis.…”

“…Several lines of evidence indicate that synaptic vesicles, which undergo the Ca 2ϩ -regulated exocytosis, are one of the most important cargoes for myosin-Va (Prekeris and Terrian, 1997;Bridgman, 1999;Tilelli et al, 2003). In addition, Myo2p, a yeast homologue of myosin-Va, directs intracellular transport during secretion and budding through interactions with other proteins (Matsui, 2003).…”

Immunohistochemical localization of myosin va in the adult rat brain

“…Its association in brain with nerve terminals (Mani et al, 1994;Prekeris and Terrian, 1997), synaptic vesicles and synaptic vesicle associated proteins (Prekeris and Terrian, 1997;Costa et.al., 1999;Ohyama et al, 2001) as well as with organelle movement in axons (Evans et al, 1998;Tabb et al, 1998) suggest that myosin V has a functional role in organelle transport.…”

“…The specific dysfunction that leads to the convulsions and post-natal death in these animals is not known. However, recent evidence has implicated MVa in several neuronal processes, including its binding to synaptic vesicle associated proteins (Prekeris and Terrian, 1997;Costa et al, 1999;Ohyama et al, 2001), its association with organelle and vesicle movement within axons (Evans et al, 1998;Tabb et al, 1998) as well as its critical role in the localization of smooth endoplasmic reticulum in dendritic spines of Purkinje cells (Dekker-Ohno et al, 1996). The subcellular localization of MVa has been described in neuronal and glial cells by immunoprobes and shows a characteristic punctate labeling, frequently being concentrated at the neurite tips and growth cones, as well as at the perinuclear region .…”