title: Cyclic GMP-dependent and -independent effects on the synthesis of the calcium messengers cyclic ADP-ribose and nicotinic acid adenine dinucleotide phosphate.
abstract: Cyclic ADP-ribose (cADPR) and nicotinic acid adenine dinucleotide phosphate (NAADP) have been shown to mobilize intracellular Ca2+ stores by totally independent mechanisms, which are pharmacologically distinct from that activated by inositol trisphosphate. Although cADPR and NAADP are structurally and functionally different, they can be synthesized by a single enzyme having ADP-ribosyl cyclase activity. In this study, three different assays were used to measure the metabolism of cADPR in sea urchin egg homogenates including a radioimmunoassay, a Ca2+ release assay, and a thin layer chromatographic assay. Soluble and membrane-bound ADP-ribosyl cyclases were identified and both cyclized NAD to produce cADPR. The soluble cyclase was half-maximally stimulated by 5.3 microM cGMP, but not by cAMP, while the membrane-bound form was independent of cGMP. The two forms of the cyclase were also different in the pH dependence of utilizing nicotinamide guanine dinucleotide (NGD), a guanine analog of NAD, as substrate, indicating they are two separate enzymes. The stimulatory effect of cGMP required ATP or ATPgammaS (adenosine 5'-O-(3-thiotriphosphate)) and a cGMP-dependent kinase activity was shown to be present in the soluble fraction. The degradation of cADPR to ADP-ribose was catalyzed by cADPR hydrolase, which was found to be predominantly associated with membranes. Similar to the membrane-bound cyclase, the cADPR hydrolase activity was also independent of cGMP. Both the soluble and membrane fractions also catalyzed the synthesis of NAADP through exchanging the nicotinamide group of NADP with nicotinic acid (NA). The base-exchange activity was independent of cGMP and the half-maximal concentrations of NADP and NA needed were about 0.2 mM and 10 mM, respectively. The exchange reaction showed a preference for acidic pH, contrasting with the neutral pH optimum of the cyclase activities. The complex metabolic pathways characterized in this study indicate that there may be a multitude of regulatory mechanisms for controlling the endogenous concentrations of cADPR and NAADP.
authors.affiliation: Department of Physiology, University of Minnesota, Minneapolis, Minnesota 55455, USA.
authors.forename: R M
authors.initials: RM
authors.lastname: Graeff
authors.forename: L
authors.initials: L
authors.lastname: Franco
authors.forename: A
authors.initials: A
authors.lastname: De Flora
authors.forename: H C
authors.initials: HC
authors.lastname: Lee
mesh_terms: D036541:ADP-ribosyl Cyclase; D051997:ADP-ribosyl Cyclase 1; D000246:Adenosine Diphosphate Ribose / Q000031:analogs & derivatives* / Q000096:biosynthesis / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D015703:Antigens, CD; D000943:Antigens, Differentiation / Q000378:metabolism; D002118:Calcium / Q000378:metabolism*; D036563:Cyclic ADP-Ribose; D006152:Cyclic GMP / Q000378:metabolism*; D007700:Kinetics; D009244:NAD+ Nucleosidase / Q000378:metabolism; D009249:NADP / Q000031:analogs & derivatives* / Q000096:biosynthesis / Q000378:metabolism; D010063:Ovum / Q000378:metabolism; D012617:Sea Urchins
chemical_list: D015703:Antigens, CD; D000943:Antigens, Differentiation; D036563:Cyclic ADP-Ribose; D000246:Adenosine Diphosphate Ribose; D009249:NADP; C024376:NAADP; D000255:Adenosine Triphosphate; D036541:ADP-ribosyl Cyclase; D009244:NAD+ Nucleosidase; D051997:ADP-ribosyl Cyclase 1; D006152:Cyclic GMP; D002118:Calcium