Modulator and messenger functions of cyclic ADP-ribose in calcium signaling

Cyclic ADP-ribose (cADPR), a Ca+2 mobilizing cyclic nucleotide derived from NAD+, is emerging as an endogenous modulator of the Ca(+2)-induced Ca+2 release (CICR) mechanism in cells. cADPR was discovered because of the prominent delay in the initiation of Ca+2 release by NAD+ in sea urchin egg homogenates, which was due to enzymatic conversion to cADPR. In addition to the egg, an invertebrate cell, amphibian neurons, a variety of mammalian cells and plant vacuoles are found to be responsive to cADPR, indicating its generality. The cyclic structure of cADPR has been determined by X-ray crystallography. A series of analogs has been synthesized, which includes cyclic GDP-ribose, a fluorescent analog, a series of specific antagonists, a photoaffinity label and caged cADPR. The use of these analogs of cADPR has provided definitive evidence for the authenticity of its Ca+2 mobilizing activity and insights for understanding its mechanism and biological functions. Results show that its action requires a soluble protein which is identified as calmodulin. The effect of calmodulin is synergistic with cADPR and both act to sensitize CICR to Ca+2. Together, the Ca+2 sensitivity of CICR can be increased by several orders of magnitude. In addition to being a modulator of CICR. cADPR can also function as a messenger. Activation of its synthetic enzyme can lead to large increases in cellular concentrations of cADPR, which would sensitize CICR to such an extent that even basal levels of cellular Ca+2 are sufficient to trigger further release. This is operationally equivalent to being a Ca+2 messenger. Three types of enzymes are involved in the metabolism of cADPR, a soluble ADP-ribosyl cyclase; a bifunctional ecto-enzyme, CD38, which is also a lymphocyte antigen; and an intracellular enzyme activable by a cGMP-dependent process. The importance of two cysteine residues in the bifunctionality of CD38 has been shown by site-directed mutagenesis. Both ADP-ribosyl cyclase and CD38 can catalyze the exchange of the nicotinamide group in NADP+ with nicotinic acid, leading to the formation of another Ca+2 mobilizing metabolite, nicotinic acid dinucleotide phosphate (NAADP). Pharmacological and desensitization studies show that the NAADP-mechanism is totally independent of the cADPR- and inositol trisphosphate-mechanisms and the Ca+2 stores responsive to NAADP are separable from those sensitive to the other two Ca+2 agonists. Microinjection studies show that all three mechanisms are present and functional in cells. The emerging picture of multiplicity in Ca+2 signaling mechanisms underscores the versatility of Ca+2 in regulating diverse cellular functions.