Dinucleotide polyphosphates contribute to purinergic signalling via inhibition of adenylate kinase activity

作者：Yegutkin Gennady G; Jankowski Joachim; Jalkanen Sirpa; Guenthner Thomas; Zidek Walter; Jankowski Vera^*

来源：Bioscience Reports, 2008, 28(4): 189-194.

DOI：10.1042/BSR20080052

• adenylate kinase
• dinucleotide polyphosphate
• purinergic signalling
• receptor antagonist
• vasoregulation

Dinucleoside polyphosphates are well described as direct vasoconstrictors and as mediators with strong proliferative properties, however, less is known about their effects on nucleotide-converting pathways. Therefore, the present study investigates the effects of ANA (diadenosine tetra phosphate), UNA (uridine adenosine tetraphosphate) and Ap(5)A (diadenosine pentaphosphate) and the non-selective P2 antagonist suramin on human serum and endothelial nucleotide-converting enzymes. Human serum and HUVECs (human umbilical vein endothelial cells) were pretreated with various concentrations of dinucleotide polyphosphates and suramin. Adenylate kinase and NDP kinase activities were then quantified radiochemically by TLC analysis of the ATP-induced conversion of [H-3]AMP and [H-3]ADP into [H-3)ADP/ATP and [H-3]ATP respectively. Endothelial NTPDase (nucleoside triphosphate diphosphohydrolase) activity was additionally determined using [H-3]ADP and [H-3]ATP as preferred substrates. Dinucleoside polyphosphates and suramin have an inhibitory effect on the serum adenylate kinase [plC(50) values (-log IC50): APA 4.67 +/- 0.03; UNA, 3.70 +/- 0.10; Ap(5)A, 6.31 +/- 0.03; suramin, 3.74 +/- 0.07], as well as on endothelial adenylate kinase (PIC50 values: APA 4.17 +/- 0.07; UPA 2.94 +/- 0.02; Ap(5)A, 5.97 +/- 0.04; suramin, 4.23 +/- 0.07), but no significant effects on serum NDP kinase, emphasizing the selectivity of these inhibitors. Furthermore, APA UNA, AP(5)A and suramin progressively inhibited the rates of [H-3]ADP (plC(50) values: AP(4)A 3.38 +/- 0.09; UPA 2.78 +/- 0.06; ANA, 4.42 +/- 0.11; suramin, 4.10 +/- 0.07) and [H-3]ATP (plC(50) values: ANA, 3.06 0.06; Ap(5)A, 3.05 +/- 0.12; suramin, 4.14 + 0.05) hydrolyses by cultured HLIVECs. UNA has no significant effect on the enclothelial NTPDase activity. Although the half-lives for Ap(4)A, UNA and Ap(5)A in serum are comparable with the incubation times of the assays used in the present study, secondary effects of the dinucleotide metabolites are not prominent for these inhibitory effects, since the concentration of metabolites formed are relatively insignificant compared with the 800 pmol/l ATP added as a phosphate donor in the adenylate kinase and NDP kinase assays. This comparative competitive study suggests that ANA and Ap(5)A contribute to the purinergic responses via inhibition of adenylate-kinase-mediated conversion of enclogenous ADP whereas UNA most likely mediates its vasoregulatory effects via direct binding-mediated mechanisms.

• 出版日期2008-8