REGULATION OF CYCLIC ADP RIBOSE SIGNALING SYSTEM

拟议研究的总体目标是深入了解 环状ADP-核糖(CADPR)动员钙离子的机制 细胞内储存，并阐明cADPR水平 在哺乳动物细胞中受到调控。Cadpr是一种自然产生的 β-烟酰胺腺嘌呤二核苷酸的代谢物) 已被发现在多种细胞中从细胞内存储中动员钙 单元类型。具体目标是：(1)鉴定和表征cadpr 哺乳动物系统中的结合蛋白，(2)研究 哺乳动物系统中cADPR的内源性水平，以及(3)表征 CADPR代谢酶的调控机制。 CADPR结合蛋白的研究将通过常规结合以及 通过光亲和标记技术。3-deaza-cADPR，一种有效的，非 CADPR的水解性类似物，是最近开发的，将是一种 CADPR与之相互作用的蛋白质检测中的重要探针。 内源性cADPR水平的调节将在几个方面进行研究 用放射免疫分析法检测cADPR的哺乳动物细胞系 实验室。将特别关注已知的特工 通过钙离子改变细胞内钙离子浓度 内流或通过动员体内的钙储存。机制(S) CADPR水平的调节将通过以下方式进行检查 负责合成的酶(ADP-核糖环化酶)和 降解(cADPR水解酶)从显示有变化的系统 CADPR水平。细胞内钙离子浓度的调节 在所有细胞中都是一个关键过程。钙在人体内起着重要的作用 过程的数量，包括神经递质释放、细胞骨架 变化、肌肉收缩、基因表达等。积累证据 提示cADPR可能是钙诱导的内源性调节因子。 兰尼定受体类钙释放(CICR) 频道。CADPR似乎增加了CICR对钙的敏感性 离子的作用方式与咖啡因非常相似。咖啡因有许多生理功能 行为，是世界上社会滥用最严重的毒品之一。这个 拟议的研究应提供关于cadpr所起作用的有用信息。 在调节钙稳态方面也可能提供有价值的 深入了解咖啡因的一些生物学作用。

The overall objectives of the proposed research are to gain insight into the mechanism by which cyclic ADP-ribose (cADPR) mobilizes calcium from intracellular stores and to elucidate the mechanisms by which cADPR levels are regulated in mammalian cells. cADPR is a naturally occurring metabolite of beta-nicotinamide adenine dinucleotide )beta-NAD) that has been found to mobilize calcium from intracellular stores in a variety of cell types. The specific aims are: (1) to identify and characterize cADPR binding proteins in mammalian systems, (2) to examine the regulation of endogenous levels of cADPR in mammalian systems, and (3) to characterize the mechanisms involved in the regulation of cADPR metabolic enzymes. cADPR binding proteins will be studied by conventional binding as well as by photoaffinity labeling techniques. 3-deaza-cADPR, a potent, non- hydrolyzable analog of cADPR, has recently been developed, and will be an important probe in the examination of proteins to which cADPR interacts. The regulation of endogenous levels of cADPR will be examined in several mammalian cell lines using a radioimmunoassay for cADPR developed in this laboratory. Particular attention will be focused on agents that are known to alter intracellular calcium concentrations, either through calcium influx or by mobilization of interal stores of calcium. The mechanism(s) by which cADPR levels are regulated will be examined by characterizing the enzymes responsible for the synthesis (ADP-ribosyl cyclase) and degradation (cADPR hydrolase) from systems shown to have alterations in cADPR levels. The regulation of the intracellular concentration of calcium is a critical process in all cells. Calcium plays an important role in a number of processes, including neurotransmitter release, cytoskeleton changes, muscle contraction, gene expression, etc. Accumulating evidence suggests that cADPR may be the endogenous regulator of calcium induced- calcium release (CICR) through ryanodine receptor type calcium release channels. cADPR appears to increase the sensitivity of CICR to calcium ions in a manner very similar to caffeine. Caffeine has many physiological actions and is one of the most socially abused drugs in the world. The proposed studies should provide useful information on the role cADPR plays in the regulation of calcium homeostasis and may also provide valuable insight into some of caffeine's biological actions.