title: Extracellular adenosine nucleotides stimulate protein kinase C activity and human neutrophil activation.
abstract: We studied the effect of adenosine nucleotides on several aspects of the functional activation of human peripheral blood polymorphonuclear leukocytes (PMN). Radiolabeled ATP bound to PMN in a manner suggesting the existence of specific binding sites because: 1) binding was reversed (92 +/- 6%) by 100-fold excess concentrations of unlabeled ATP but minimally by either ADP (43 +/- 12%) or GTP (37 +/- 8%); and 2) binding saturation was achieved (i.e., specific binding did not increase) above 250 microM ATP. Binding studies revealed that significant ATP hydrolysis occurred, even at low temperatures and in the presence of phosphatase inhibitors. Adenosine nucleotides activated signal transduction mechanisms in PMN because: 1) 1 to 100 microM ATP and 5'-adenylylimidodiphosphate (AMP-PNP) stimulated increased production of 1,2-diacylglycerols; 2) ATP (0.5 to 500 microM) and ADP (0.1 to 10 mM) induced increased insoluble protein kinase (PKC) activity in a dose-dependent manner when used at concentrations greater than 50 microM; 3) ATP (greater than or equal to 50 microM) induced a shift in the solubility of phorbol receptors from mostly soluble (89% in untreated cells) to mostly insoluble (68%), whereas ADP, GTP, and GDP were effective at higher concentrations; and 4) greater than or equal to 50 microM ATP stimulated increased phosphorylation of endogenous PMN proteins. AMP-PNP induced PKC activity and phosphoprotein changes that were qualitatively similar to those observed when PMN were treated with ATP, suggesting that extracellular ATP hydrolysis is not required for signal transduction to activate PKC. Functionally, ATP stimulated the secretion of specific (but not azurophil) granules because vitamin B12-binding protein and low levels of lysozyme, but not beta-glucuronidase, were released; qualitatively similar results were obtained by using AMP-PNP. These results suggest that certain adenosine nucleotides employed at physiologically relevant concentrations stimulate increased 1,2-diacylglycerol production, PKC activity, granule secretion, and endogenous phosphoprotein formation in a manner that is independent of extracellular ATP hydrolysis.
authors: Department of Pediatrics, University of Michigan Medical School, Ann Arbor 48109.
authors: K J
authors: KJ
authors: Balazovich
authors: L A
authors: LA
authors: Boxer
mesh_terms: D000227:Adenine Nucleotides / Q000494:pharmacology*; D002118:Calcium / Q000502:physiology; D002451:Cell Compartmentation / Q000187:drug effects; D004075:Diglycerides / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D005110:Extracellular Space; D006801:Humans; D066298:In Vitro Techniques; D008970:Molecular Weight; D009504:Neutrophils / Q000201:enzymology / Q000502:physiology*; D010750:Phosphoproteins / Q000502:physiology; D011493:Protein Kinase C / Q000378:metabolism*; D011956:Receptors, Cell Surface / Q000502:physiology*; D015398:Signal Transduction
chemical_list: D000227:Adenine Nucleotides; D004075:Diglycerides; D010750:Phosphoproteins; D011956:Receptors, Cell Surface; D011493:Protein Kinase C; D002118:Calcium