A Derivative of NADP Mobilizes Calcium Stores Insensitive to Inositol Trisphosphate and Cyclic ADP-ribose

doi:10.1074/jbc.270.5.2152

A Derivative of NADP Mobilizes Calcium Stores Insensitive to Inositol Trisphosphate and Cyclic ADP-ribose (*)

Hon Cheung Lee(§) and
Robert Aarhus

From the Department of Physiology, University of Minnesota, Minneapolis, Minnesota 55455

^§ To whom correspondence should be addressed:
6-182 Lyon Lab., Dept. of Physiology, University of Minnesota, Minneapolis, MN 55455.
Tel.: 612-625-7120; Fax: 612-625-0991.

Abstract

We have previously shown that alkaline treatment of NADP generates a derivative which can mobilize Ca from sea urchin egg homogenates (Clapper, D. L., Walseth, T. F., Dargie, P. J., and Lee, H. C.(1987) J. Biol. Chem. 262, 9561-9568). In this study, the active derivative was purified and shown by high pressure liquid chromatography to be distinct from NADP and NADPH. However, its proton NMR spectrum was virtually identical to that of NADP. The mass of its molecular ion was measured by high resolution mass spectrometry to be 743.0510, one mass unit larger than the corresponding ion of NADP. These results are consistent with the active derivative being nicotinic acid adenine dinucleotide phosphate (NAADP). Ca release induced by NAADP was saturable with a half-maximal concentration of about 30 nM. The release was specific since NADP and nicotinic acid adenine dinucleotide were ineffective even at 10-40-fold higher concentrations. The NAADP-dependent Ca release showed desensitization and was insensitive to heparin and a specific antagonist of cyclic ADP-ribose (cADPR), 8-amino-cADPR. The release mechanism did not require calmodulin. This is similar to the inositol trisphosphate-sensitive release but distinct from that of cADPR. That the NAADP-sensitive Ca stores were different from those sensitive to inositol trisphosphate- or cADPR was further indicated by their differences in distribution on Percoll density gradients. Microinjection of NAADP into live sea urchin eggs induced transient elevation of intracellular Ca and triggered the cortical reaction, indicating the NAADP-dependent mechanism is operative in intact cells.

Footnotes

↵* This work was supported by National Institutes of Health Grants HD17484 and HD32040 (to H. C. L.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore by hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵¹ The abbreviations used are:

IP₃

inositol 1,4,5-trisphosphate

cADPR

cyclic ADP-ribose

NAADP

nicotinic acid adenine dinucleotide phosphate

A-NADP

alkaline-activated NADP

NAAD

nicotinic acid adenine dinucleotide

HPLC

high pressure liquid chromatography.
- Received October 18, 1994.
- Revision received November 22, 1994.