Kindman L A, Meyer T
Department of Cell Biology, Stanford University Medical Center, California 94305.
Biochemistry. 1993 Feb 9;32(5):1270-7. doi: 10.1021/bi00056a011.
Quantal Ca2+ release is a novel motif for the mediation of signal transduction in which the amplitude of a biological response following multiple stepwise increases in agonist concentration is retained. The release of Ca2+ from permeabilized cells in response to the second messenger inositol 1,4,5-trisphosphate (InsP3) proceeds in this fashion. The mechanisms leading to quantal Ca2+ release are unknown. InsP3 releases 50-90% of the Ca2+ sequestered within the intracellular stores of mammalian cells permeabilized with saponin. However, preparation of microsomes results in the loss of this sensitivity. In this report, functionally intact intracellular Ca2+ stores were isolated from rat basophilic leukemia (RBL) cells by osmotic lysis followed by differential and sucrose density gradient centrifugation. From this preparation, 64% of the stored Ca2+ is released by InsP3. We demonstrate that quantal Ca2+ release is retained by isolated Ca2+ stores and is identical to that observed in permeabilized cells. Addition of a subsaturating (28 nM) concentration of InsP3 to permeabilized cells at 37 degrees C results in the release of only a small fraction of the sequestered Ca2+. When the cells are cooled to 11 degrees C, the remaining Ca2+ is rapidly released. Hence, the mechanism leading to the quantal nature of Ca2+ release is reversible and is thus not likely to be the result of a covalent modification of the channel protein or of the Ca2+ store.(ABSTRACT TRUNCATED AT 250 WORDS)
量子化Ca2+释放是信号转导介导中的一种新基序，在这种基序中，激动剂浓度多次逐步增加后生物反应的幅度得以保留。响应第二信使肌醇1,4,5-三磷酸（InsP3），Ca2+从透化细胞中释放就是以这种方式进行的。导致量子化Ca2+释放的机制尚不清楚。InsP3可释放用皂素透化的哺乳动物细胞胞内储存中50%至90%的Ca2+。然而，制备微粒体会导致这种敏感性丧失。在本报告中，通过渗透裂解，然后进行差速离心和蔗糖密度梯度离心，从大鼠嗜碱性白血病（RBL）细胞中分离出功能完整的胞内Ca2+储存。从该制备物中，64%的储存Ca2+可被InsP3释放。我们证明，分离出的Ca2+储存保留了量子化Ca2+释放，且与在透化细胞中观察到的相同。在37℃下向透化细胞中添加亚饱和（28 nM）浓度的InsP3，只会导致一小部分螯合的Ca2+释放。当细胞冷却至11℃时，剩余的Ca2+会迅速释放。因此，导致Ca2+释放量子化性质的机制是可逆的，因此不太可能是通道蛋白或Ca2+储存共价修饰的结果。（摘要截短于250字）
