Myosin-Va regulates exocytosis through the submicromolar Ca2+-dependent binding of syntaxin-1A.
Myosin-Va is an actin-based processive motor that conveys intracellular cargoes. Synaptic vesicles are one of the most important cargoes for myosin-Va, but the role of mammalian myosin-Va in secretion is less clear than for its yeast homologue, Myo2p. In the current studies, we show that myosin-Va on synaptic vesicles interacts with syntaxin-1A, a t-SNARE involved in exocytosis, at or above 0.3 microM Ca2+. Interference with formation of the syntaxin-1A-myosin-Va complex reduces the exocytotic frequency in chromaffin cells. Surprisingly, the syntaxin-1A-binding site was not in the tail of myosin-Va but rather in the neck, a region that contains calmodulin-binding IQ-motifs. Furthermore, we found that syntaxin-1A binding by myosin-Va in the presence of Ca2+ depends on the release of calmodulin from the myosin-Va neck, allowing syntaxin-1A to occupy the vacant IQ-motif. Using an anti-myosin-Va neck antibody, which blocks this binding, we demonstrated that the step most important for the antibody's inhibitory activity is the late sustained phase, which is involved in supplying readily releasable vesicles. Our results demonstrate that the interaction between myosin-Va and syntaxin-1A is involved in exocytosis and suggest that the myosin-Va neck contributes not only to the large step size but also to the regulation of exocytosis by Ca2+.
Watanabe
Michitoshi
M
Division of Molecular and Cellular Biology, Graduate School of Medical and Dental Sciences, Niigata University, Niigata, Niigata 951-8510, Japan.
Nomura
Kazushige
K
Ohyama
Akihiro
A
Ishikawa
Ryoki
R
Komiya
Yoshiaki
Y
Hosaka
Kohei
K
Yamauchi
Emiko
E
Taniguchi
Hisaaki
H
Sasakawa
Nobuyuki
N
Kumakura
Konosuke
K
Ushiki
Tatsuo
T
Sato
Osamu
O
Ikebe
Mitsuo
M
Igarashi
Michihiro
M
D000595:Amino Acid Sequence; D000818:Animals; D001921:Brain / Q000378:metabolism / Q000648:ultrastructure; D002118:Calcium / Q000502:physiology*; D002478:Cells, Cultured; D019439:Chromaffin Cells / Q000378:metabolism; D005089:Exocytosis / Q000502:physiology*; D018625:Microscopy, Atomic Force; D008969:Molecular Sequence Data; D018995:Myosin Heavy Chains / Q000502:physiology* / Q000648:ultrastructure; D024701:Myosin Type V / Q000502:physiology* / Q000648:ultrastructure; D011485:Protein Binding; D051381:Rats; D013572:Synaptic Vesicles / Q000378:metabolism; D050827:Syntaxin 1 / Q000378:metabolism* / Q000648:ultrastructure
C513081:Myo5a protein, rat; D050827:Syntaxin 1; D024701:Myosin Type V; D018995:Myosin Heavy Chains; D002118:Calcium