TY - JOUR
T1 - Extracellular NAD+ is an agonist of the human P2Y11 purinergic receptor in human granulocytes
AU - Moreschi, Iliana
AU - Bruzzone, Santina
AU - Nicholas, Robert A.
AU - Fruscione, Floriana
AU - Sturla, Laura
AU - Benvenuto, Federica
AU - Usai, Cesare
AU - Meis, Sabine
AU - Kassack, Matthias U.
AU - Zocchi, Elena
AU - De Flora, Antonio
PY - 2006/10/20
Y1 - 2006/10/20
N2 - Micromolar concentrations of extracellular β-NAD+ (NAD e
+) activate human granulocytes (superoxide and NO generation and chemotaxis) by triggering: (i) overproduction of cAMP, (ii) activation of protein kinase A, (iii) stimulation of ADP-ribosyl cyclase and overproduction of cyclic ADP-ribose (cADPR), a universal Ca2+ mobilizer, and (iv) influx of extracellular Ca2+. Here we demonstrate that exposure of granulocytes to millimolar rather than to micromolar NAD e
+ generates both inositol 1,4,5-trisphosphate (IP 3) and cAMP, with a two-step elevation of intracellular calcium levels ([Ca2+]i): a rapid, IP3-mediated Ca 2+ release, followed by a sustained influx of extracellular Ca 2+ mediated by cADPR. Suramin, an inhibitor of P2Y receptors, abrogated NADe
+-induced intracellular increases of IP 3, cAMP, cADPR, and [Ca2+]i, suggesting a role for a P2Y receptor coupled to both phospholipase C and adenylyl cyclase. The P2Y11 receptor is the only known member of the P2Y receptor subfamily coupled to both phospholipase C and adenylyl cyclase. Therefore, we performed experiments on hP2Y11-transfected 1321N1 astrocytoma cells: micromolar NADe
+ promoted a two-step elevation of the [Ca2+]i due to the enhanced intracellular production of IP3, cAMP, and cADPR in 1321N1-hP2Y11 but not in untransfected 1321N1 cells. In human granulocytes NF157, a selective and potent inhibitor of P2Y11, and the down-regulation of P2Y11 expression by short interference RNA prevented NADe +-induced intracellular increases of [Ca2+]i and chemotaxis. These results demonstrate that β-NADe + is an agonist of the P2Y11 purinoceptor and that P2Y11 is the endogenous receptor in granulocytes mediating the sustained [Ca2+]i increase responsible for their functional activation.
AB - Micromolar concentrations of extracellular β-NAD+ (NAD e
+) activate human granulocytes (superoxide and NO generation and chemotaxis) by triggering: (i) overproduction of cAMP, (ii) activation of protein kinase A, (iii) stimulation of ADP-ribosyl cyclase and overproduction of cyclic ADP-ribose (cADPR), a universal Ca2+ mobilizer, and (iv) influx of extracellular Ca2+. Here we demonstrate that exposure of granulocytes to millimolar rather than to micromolar NAD e
+ generates both inositol 1,4,5-trisphosphate (IP 3) and cAMP, with a two-step elevation of intracellular calcium levels ([Ca2+]i): a rapid, IP3-mediated Ca 2+ release, followed by a sustained influx of extracellular Ca 2+ mediated by cADPR. Suramin, an inhibitor of P2Y receptors, abrogated NADe
+-induced intracellular increases of IP 3, cAMP, cADPR, and [Ca2+]i, suggesting a role for a P2Y receptor coupled to both phospholipase C and adenylyl cyclase. The P2Y11 receptor is the only known member of the P2Y receptor subfamily coupled to both phospholipase C and adenylyl cyclase. Therefore, we performed experiments on hP2Y11-transfected 1321N1 astrocytoma cells: micromolar NADe
+ promoted a two-step elevation of the [Ca2+]i due to the enhanced intracellular production of IP3, cAMP, and cADPR in 1321N1-hP2Y11 but not in untransfected 1321N1 cells. In human granulocytes NF157, a selective and potent inhibitor of P2Y11, and the down-regulation of P2Y11 expression by short interference RNA prevented NADe +-induced intracellular increases of [Ca2+]i and chemotaxis. These results demonstrate that β-NADe + is an agonist of the P2Y11 purinoceptor and that P2Y11 is the endogenous receptor in granulocytes mediating the sustained [Ca2+]i increase responsible for their functional activation.
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U2 - 10.1074/jbc.M606625200
DO - 10.1074/jbc.M606625200
M3 - Article
C2 - 16926152
AN - SCOPUS:33845998565
VL - 281
SP - 31419
EP - 31429
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
SN - 0021-9258
IS - 42
ER -