Cyclic Adenosine Diphosphate Ribose Activates Ryanodine Receptors, whereas NAADP Activates Two-pore Domain Channels

Abstract

The mechanism by which cyclic adenosine diphosphate ribose (cADPR) and nicotinic acid adenine dinucleotide phosphate (NAADP) mobilize intracellular Ca 2؉ Intracellular Ca 2ϩ signals are initiated by Ca 2ϩ release from intracellular stores, and traditionally, the sarco/endoplasmic reticulum (S/ER) 2 has been considered to be the major releasable store. From this store, Ca 2ϩ may be released through the opening of inositol 1,4,5-trisphosphate receptors (IP 3 Rs) and/or ryanodine receptors (RyRs), the two groups of intracellular Ca 2ϩ release channels located on S/ER membranes. It is generally accepted that of the recognized Ca 2ϩ -mobilizing second messengers, inositol 1,4,5-trisphosphate facilitates this process by activating IP 3 Rs (1).By contrast, the mechanism by which the pyridine nucleotides cyclic adenosine diphosphate ribose (cADPR) and nicotinic acid adenine dinucleotide phosphate (NAADP) (2) mobilize intracellular Ca 2ϩ stores remains controversial. Although a wealth of evidence across a variety of cell types (3-9) supports the original proposal that cADPR activates RyRs (10), studies on reconstituted RyRs in lipid bilayers have failed to conclusively demonstrate direct regulation of these channels by cADPR (11), and it has been suggested that cADPR may initiate Ca 2ϩ signals via RyRs and IP 3 Rs by promoting Ca 2ϩ uptake into the S/ER by S/ER Ca 2ϩ ATPases (SERCA) (12)(13)(14). This proposal has not been effectively countered by studies on ventricular myocytes, which exclusively express RyR2. This is due to the fact that the principal regulatory effect of cADPR with respect to RyR2 is to increase the sensitivity of this RyR subtype to Ca 2ϩ -induced Ca 2ϩ release (8, 9), and in light of the fact that the sensitivity of RyRs to Ca 2ϩ -induced Ca 2ϩ release may be augmented by an increase in Ca 2ϩ concentration within the cytoplasm and/or S/ER lumen (15).The mechanism by which NAADP triggers intracellular Ca 2ϩ release has also been hotly debated. We recently identified a family of two-pore domain channels (TPC1-3, TPCN1-3 for gene name) as endolysosome-targeted, NAADP-gated Ca 2ϩ release channels (16, 17), and our findings have since been confirmed by others (18,19

Organelle-specific Subunit Interactions of the Vertebrate Two-pore Channel Family

“…Changes in intracellular Ca 2ϩ were reported by the fluorescence ratio (F 340 /F 380 ) of the Ca 2ϩ indicator Fura-2. As reported previously (1,18), intracellular dialysis of 10 nM NAADP failed to evoke a significant Ca 2ϩ transient in wild-type HEK293 cells (Fig. 4A, panel i), which express very low levels of TPC1 and TPC2 and do not express TPC3 (1,19).…”

Photoaffinity Labeling of Nicotinic Acid Adenine Dinucleotide Phosphate (NAADP) Targets in Mammalian Cells*

“…Next, we proceeded to apply the PAL approach in two other cell types, HEK293 cells and mouse pancreas. Both systems have been used as models for studying NAADP-evoked Ca 2ϩ signals (5,6,9,10,14,29,30). PAL samples from HEK WCL displayed a broadly similar labeling pattern to those observed from SKBR3 cells (Fig.…”

“…Supporting evidence derives from gain-and loss-of-function approaches, electrophysiological analyses, and radioligand binding. For example, overexpression of either of the two human TPC isoforms (TPC1 and TPC2) enhanced NAADPevoked Ca 2ϩ responses when assessed by Ca 2ϩ imaging in multiple cell lines (5)(6)(7)(8)(9)(10). Reciprocally, knockdown of endogenous TPC1 ablated NAADP responsiveness (7), and NAADP-activated Ca 2ϩ currents were abolished in pancreatic ␤-cells isolated from a TPC2 knock-out mouse (5).…”

Roles and mechanisms of the CD38/cyclic adenosine diphosphate ribose/Ca2+signaling pathway

“…In HEK293 cells transfected with an islet type RyR, which is a splice variant of the RyR2 gene by alternative splicing of exons 4 and 75, Ca 2+ release was enhanced in the presence of 100 μmol/L cADPR, and the effect could be reversed by preincubating with a cADPR antagonist, 8-bromo-cADPR (8-Br-cADPR) [13] . Similarly, cADPR triggered a marked Ca 2+ transient in HEK293 cells that stably expressed RyR1 and RyR3, and this Ca 2+ transient was abolished by dantrolene, an RyR antagonist [14] . In summary, all these results suggested that RyRs might serve as cADPR receptors (Figure 1).…”