Bhalla Akhil, Tucker Ward C, Chapman Edwin R
Department of Physiology, University of Wisconsin, Madison, WI 53706, USA.
Mol Biol Cell. 2005 Oct;16(10):4755-64. doi: 10.1091/mbc.e05-04-0277. Epub 2005 Aug 10.
Ca2+-triggered exocytosis of synaptic vesicles is controlled by the Ca2+-binding protein synaptotagmin (syt) I. Fifteen additional isoforms of syt have been identified. Here, we compared the abilities of three syt isoforms (I, VII, and IX) to regulate soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE)-mediated membrane fusion in vitro in response to divalent cations. We found that different isoforms of syt couple distinct ranges of Ca2+, Ba2+, and Sr2+ to membrane fusion; syt VII was approximately 400-fold more sensitive to Ca2+ than was syt I. Omission of phosphatidylserine (PS) from both populations of liposomes completely abrogated the ability of all three isoforms of syt to stimulate fusion. Mutations that selectively inhibit syt.target-SNARE (t-SNARE) interactions reduced syt stimulation of fusion. Using Sr2+ and Ba2+, we found that binding of syt to PS and t-SNAREs can be dissociated from activation of fusion, uncovering posteffector-binding functions for syt. Our data demonstrate that different syt isoforms are specialized to sense different ranges of divalent cations and that PS is an essential effector of Ca2+.syt action.
突触小泡的钙离子触发胞吐作用由钙离子结合蛋白突触结合蛋白（syt）I控制。已鉴定出另外15种syt亚型。在此，我们比较了三种syt亚型（I、VII和IX）在体外响应二价阳离子时调节可溶性N - 乙基马来酰亚胺敏感因子附着蛋白受体（SNARE）介导的膜融合的能力。我们发现，不同的syt亚型将不同范围的Ca2 +、Ba2 +和Sr2 +与膜融合偶联起来；syt VII对Ca2 +的敏感性比syt I高约400倍。从两组脂质体中去除磷脂酰丝氨酸（PS）完全消除了所有三种syt亚型刺激融合的能力。选择性抑制syt - 靶标SNARE（t - SNARE）相互作用的突变降低了syt对融合的刺激作用。使用Sr2 +和Ba2 +，我们发现syt与PS和t - SNAREs的结合可以与融合激活解离，揭示了syt的效应器后结合功能。我们的数据表明，不同的syt亚型专门用于感知不同范围的二价阳离子，并且PS是Ca2 + - syt作用的必需效应器。
