CHARACTERIZATION OF NITRIC OXIDE-STIMULATED ADP-RIBOSYLATION OF VARIOUS PROTEINS FROM THE MOUSE MACROPHAGE CELL-LINE ANA-1 USING SODIUM-NITROPRUSSIDE AND THE NOVEL NITRIC-OXIDE DONATING COMPOUND DIETHYLAMINE DINITRIC OXIDE

We examined the ability of nitric oxide (NO) to stimulate the ADP-ribo sylation of proteins from the mouse macrophage cell line ANA-1. To dem onstrate a specific effect of NO, we used a novel compound named dieth ylamine dinitric oxide (DEA/NO; 1,1-diethyl-2-hydroxy-2-nitrosohydrazi ne, sodium salt; [Et(2)NN(O)NO]Na), which releases NO in aqueous solut ion at neutral pH. DEA/NO stimulated the ADP-ribosylation of at least three cytosolic proteins (M(r) = 28,000, 33,000, and 39,000) from ANA- 1 macrophages. The effect of DEA/NO on the ADP-ribosylation of the pre dominant target p39 was dose dependent(EC(50) = 80 mu M). Moreover, th e effect of DEA/NO was attributed specifically to released NO rather t han diethylamine or nitrite. Sodium nitroprusside (SNP) also stimulate d the ADP-ribosylation of cytosolic proteins from ANA-1 mouse macropha ges. However, SNP exhibited different time- and dose-dependent effects on the modification of p39. NO synthesized via the act activity of in terferon-gamma plus lipopolysaccharide-induced NO synthase also enhanc ed the ADP-ribosylation of p39, confirming that the effects of DEA/NO and SNP could be attributed to NO or reactive nitrogen oxide species. Neither pertussis toxin nor cholera toxin stimulated the ADP-ribosylat ion of p39; however, cholera toxin stimulated the ADP-ribosylation of proteins with approximate molecular weights of 28,000 and 33,000. Thes e data suggest that the induced expression of NO synthase in tumoricid al macrophages may be associated with autocrine and paracrine effects of NO that include the ADP-ribosylation of various proteins. Moreover, these results indicate that DEA/NO and related compounds may be usefu l as pharmacologic tools for investigating the-effects of NO and react ive nitrogen oxide species on macrophages.