Ca2+/calmodulin-dependent nitric oxide synthase activity in the human cervix carcinoma cell line ME-180.
We show here that the human cervix carcinoma cell line ME-180 expresses a constitutive nitric oxide (NO) synthase, as demonstrated by formation of [3H]citrulline and nitrite. The enzyme is dependent on tetrahydrobiopterin, NADPH, flavins and Ca2+/calmodulin. Enzyme activity is located in the cytosol rather than in the membrane fraction and can be inhibited by NG-monomethyl-L-arginine (NMMA). An antiserum to NO synthase purified from porcine cerebellum inhibited the enzyme activity. ME-180 cells released NO, as was shown by stimulation of guanylate cyclase (EC 4.6.1.2) in RFL-6 detector cells; this release was stimulated 8-fold by the Ca2+ ionophore A23187 and 2-fold by increasing the intracellular tetrahydrobiopterin levels with cytokines. This is the first characterization of a Ca2+/calmodulin-dependent NO synthase activity in human epithelial-type tumour cells.
Werner-Felmayer
G
G
Institute for Medical Chemistry and Biochemistry, University of Innsbruck, Austria.
Werner
E R
ER
Fuchs
D
D
Hausen
A
A
Mayer
B
B
Reibnegger
G
G
Weiss
G
G
Wachter
H
H
D000594:Amino Acid Oxidoreductases / Q000378:metabolism*; D001120:Arginine / Q000031:analogs & derivatives / Q000494:pharmacology; D002118:Calcium / Q000494:pharmacology*; D002147:Calmodulin / Q000494:pharmacology*; D006152:Cyclic GMP / Q000378:metabolism; D005260:Female; D006801:Humans; D007371:Interferon-gamma / Q000494:pharmacology; D007700:Kinetics; D008070:Lipopolysaccharides / Q000494:pharmacology; D019001:Nitric Oxide Synthase; D011621:Pteridines / Q000494:pharmacology; D011994:Recombinant Proteins; D014407:Tumor Cells, Cultured; D002583:Uterine Cervical Neoplasms / Q000201:enzymology*; D019323:omega-N-Methylarginine
D002147:Calmodulin; D008070:Lipopolysaccharides; D011621:Pteridines; D011994:Recombinant Proteins; D019323:omega-N-Methylarginine; D007371:Interferon-gamma; D001120:Arginine; D019001:Nitric Oxide Synthase; D000594:Amino Acid Oxidoreductases; D006152:Cyclic GMP; D002118:Calcium