CYCLIC ADP/RIBOSE-DEPENDENT CALCIUM RELEASE PATHWAY

循环 ADP/核糖依赖性钙释放途径

• 批准号: 2204936
• 负责人: HONCHEUNG LEE
• 金额: $ 15.14万
• 依托单位: UNIVERSITY OF MINNESOTA
• 依托单位国家: 美国
• 财政年份: 1994
• 资助国家: 美国
• 起止时间: 1994-07-15 至 1998-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2204936
• 关键词: ADP ribosylation; CD antigens; affinity labeling; animal tissue; calcium channel; calcium flux; chickens; cyclic GMP; egg /ovum; enzyme activity; enzyme mechanism; high performance liquid chromatography; hydrolase; laboratory rat; microsomes; nicotinamide adenine dinucleotide; nitric oxide; nucleotide metabolism; phosphorylation; protein kinase; protein purification; protein sequence; second messengers; western blottings

Mobilization of Ca+2 from intracellular stores is an important signaling mechanism in cells and is mediated by two major mechanisms, the inositol trisphosphate (IP3) pathway and the Ca+2-induced Ca+2 release (CICR) process. We have identified a Ca+2 mobilization system in sea urchin eggs which is totally independent of the IP3 pathway. This system is activated by a metabolite of NAD+ we called cyclic ADP-ribose (cADPR). In addition to sea urchin eggs, several mammalian cell types have been shown to be responsive to cADPR, indicating the generality of the mechanism. Increasing evidence suggests that cADPR may be an endogenous regulator of CICR in cells. Three recent advances indicate that the cADPR mechanism is tightly regulated. First, the synthetic pathway of cADPR has been shown to be stimulated by a cGMP-dependent mechanism. Second, a lymphocyte protein called CD38 has been shown to be a bifunctional enzyme that can catalyze both the synthesis and hydrolysis of cADPR and third, a soluble protein factor has been shown to be required for conferring the cADPR-sensitivity to microsomes. The proposed research will examine the regulation mechanisms of the cADPR pathway. (l) The soluble protein factors from brain and sea urchin eggs that confer the cADPR sensitivity to egg microsomes will be purified and characterized. We will investigate the possibilities that the soluble factor functions as a sensitizer of the cADPR-receptor to cADPR and/or the Ca+2 release mechanism to Ca+2. (2) The cGMP-dependent regulation mechanism of ADP-ribosyl cyclase and CD38 will be elucidated. We will use direct photoaffinity labeling and protein sequencing to identify the components involved in mediating the stimulatory effect of cGMP on the synthetic pathway of cADPR. (3) The enzymatic mechanisms of ADP-ribosyl cyclase and CD38 will be investigated. We will examine the formation of the ADP-ribosylated enzyme intermediates and identify the catalytic sites of the enzymes by photoaffinity labeling. (4) Finally, we will extend the results obtained from egg microsomes to intact eggs and to mammalian brain microsomes.

Ca+2从细胞内储存的动员是一个重要的信号