Grishanin Ruslan N, Kowalchyk Judith A, Klenchin Vadim A, Ann Kyougsook, Earles Cynthia A, Chapman Edwin R, Gerona Roy R L, Martin Thomas F J
Department of Biochemistry, University of Wisconsin, Madison, Wisconsin 53706, USA.
Neuron. 2004 Aug 19;43(4):551-62. doi: 10.1016/j.neuron.2004.07.028.
CAPS-1 is required for Ca2+-triggered fusion of dense-core vesicles with the plasma membrane, but its site of action and mechanism are unknown. We analyzed the kinetics of Ca2+-triggered exocytosis reconstituted in permeable PC12 cells. CAPS-1 increased the initial rate of Ca2+-triggered vesicle exocytosis by acting at a rate-limiting, Ca2+-dependent prefusion step. CAPS-1 activity depended upon prior ATP-dependent priming during which PIP2 synthesis occurs. CAPS-1 activity and binding to the plasma membrane depended upon PIP2. Ca2+ was ineffective in triggering vesicle fusion in the absence of CAPS-1 but instead promoted desensitization to CAPS-1 resulting from decreased plasma membrane PIP2. We conclude that CAPS-1 functions following ATP-dependent priming as a PIP2 binding protein to enhance Ca2+-dependent DCV exocytosis. Essential prefusion steps in dense-core vesicle exocytosis involve sequential ATP-dependent synthesis of PIP2 and the subsequent PIP2-dependent action of CAPS-1. Regulation of PIP2 levels and CAPS-1 activity would control the secretion of neuropeptides and monoaminergic transmitters.
CAPS-1是钙离子触发的致密核心囊泡与质膜融合所必需的，但它的作用位点和机制尚不清楚。我们分析了在可渗透的PC12细胞中重建的钙离子触发的胞吐作用的动力学。CAPS-1通过作用于限速的、依赖钙离子的预融合步骤来提高钙离子触发的囊泡胞吐作用的初始速率。CAPS-1的活性依赖于先前的ATP依赖性引发，在此期间会发生磷脂酰肌醇-4,5-二磷酸（PIP2）的合成。CAPS-1的活性和与质膜的结合依赖于PIP2。在没有CAPS-1的情况下，钙离子在触发囊泡融合方面无效，反而会促进由于质膜PIP2减少导致的对CAPS-1的脱敏。我们得出结论，CAPS-1在ATP依赖性引发后作为一种PIP2结合蛋白发挥作用，以增强钙离子依赖性致密核心囊泡的胞吐作用。致密核心囊泡胞吐作用中必不可少的预融合步骤包括PIP2的顺序ATP依赖性合成以及随后CAPS-1的PIP2依赖性作用。PIP2水平和CAPS-1活性的调节将控制神经肽和单胺能递质的分泌。
