title: Synthesis of NAADP and cADPR in mitochondria.
abstract: Here we investigated whether cADPR and NAADP are synthesized in mitochondria. We found that ADPR-cyclase activity is present in mitochondria. In addition, we describe for the first time synthesis of NAADP in this intracellular organelle. ADPR-cyclase activities (V(MAX)) and NAADP synthesis in mitochondria were about 4-fold lower than that in plasma membranes. Otherwise, ADPR-cyclases in mitochondria and in plasma membranes have similar catalytic properties in terms of apparent K(m) for the substrate NGD and K(i) values for inhibition by dithiotreitol, beta-NAD, and nicotinamide. ADPR-cyclase in plasma membranes and to a lesser degree mitochondrial enzyme, was inhibited by Zn(2+) and Cu(2+); ADPR-cyclase from mitochondria was more stable upon thermal inactivation. CD38 antigen, determined by Western blot, was well-expressed in plasma membranes but was far less so (17-fold less) in mitochondria. The major difference between ADPR-cyclase activity in mitochondria and plasma membranes is that mitochondrial cyclase activity was increased by incubation with nonionic detergents. Conversely, the incubation with phosphatidylinositol-specific phosphodiesterase C (PI-PLC) released ADPR-cyclase activity from plasma membranes, but not from mitochondria. We conclude that ADPR-cyclase in mitochondria and in plasma membranes are both multifunctional enzymes with similar catalytic properties; however, the two ADPR-cyclases differ in the mode of anchoring to the membrane: by glycosylphosphoinositol anchor in plasma membranes and by hydrophobic interactions in mitochondria. In addition, synthesis of NAADP can also be found in intracellular organelles via mitochondria. We propose that independent mitochondrial cADPR and NAADP systems may have an intracrine signaling function that is not dependent on direct input by extracellular hormonal stimuli, but rather responds to changes of intermediary cellular metabolism.
authors.affiliation: Department of Physiology and Biophysics, Mayo Clinic and Mayo Medical School, 921B Guggenheim Building, Rochester, Minnesota 55905, USA.
authors.forename: M
authors.initials: M
authors.lastname: Liang
authors.forename: E N
authors.initials: EN
authors.lastname: Chini
authors.forename: J
authors.initials: J
authors.lastname: Cheng
authors.forename: T P
authors.initials: TP
authors.lastname: Dousa
mesh_terms: D036541:ADP-ribosyl Cyclase; D051997:ADP-ribosyl Cyclase 1; D000246:Adenosine Diphosphate Ribose / Q000031:analogs & derivatives* / Q000037:antagonists & inhibitors / Q000096:biosynthesis; D000818:Animals; D015703:Antigens, CD; D000943:Antigens, Differentiation / Q000378:metabolism*; D002451:Cell Compartmentation; D002462:Cell Membrane / Q000201:enzymology*; D036563:Cyclic ADP-Ribose; D004229:Dithiothreitol / Q000494:pharmacology; D017261:Glycosylphosphatidylinositols; D006150:Guanine Nucleotides / Q000378:metabolism; D007527:Isoenzymes / Q000378:metabolism; D008562:Membrane Glycoproteins; D008930:Mitochondria, Liver / Q000201:enzymology*; D009097:Multienzyme Complexes / Q000378:metabolism; D009243:NAD / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology; D009244:NAD+ Nucleosidase / Q000378:metabolism*; D009249:NADP / Q000031:analogs & derivatives* / Q000096:biosynthesis; D009536:Niacinamide / Q000494:pharmacology; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D054801:Phosphoinositide Phospholipase C; D051381:Rats; D017207:Rats, Sprague-Dawley; D010738:Type C Phospholipases / Q000378:metabolism
chemical_list: D015703:Antigens, CD; D000943:Antigens, Differentiation; D017261:Glycosylphosphatidylinositols; D006150:Guanine Nucleotides; D007527:Isoenzymes; D008562:Membrane Glycoproteins; D009097:Multienzyme Complexes; D009243:NAD; D036563:Cyclic ADP-Ribose; D000246:Adenosine Diphosphate Ribose; D009536:Niacinamide; D009249:NADP; C024376:NAADP; C102728:nicotinamide guanine dinucleotide; D010738:Type C Phospholipases; D054801:Phosphoinositide Phospholipase C; D036541:ADP-ribosyl Cyclase; C498156:Cd38 protein, rat; D009244:NAD+ Nucleosidase; D051997:ADP-ribosyl Cyclase 1; D043265:Phosphatidylinositol Diacylglycerol-Lyase; D004229:Dithiothreitol