Molecular Pharmacology, Vol 15, 78-85, Copyright © 1979 by the American Society for Pharmacology and Experimental Therapeutics

Induction of Tyrosine Hydroxylase and Dopamine -Hydroxylase in Cultured Mouse Neuroblastoma by 8Br-cAMP

Involvement of RNA and Protein Synthesis

¹ Department of Psychobiology, School of Biological Sciences, University of California, Irvine, California 92717

8Br-cAMP elevated tyrosine hydroxylase (E.C. 1.14.16.2, L-tyrosine, tetrahydropteridine: oxygen oxidoreductase [3-hydroxylating]) and dopamine beta-hydroxylase (E.C. 1.14.17.1, 3,4-dihydroxyphenylethylamine, ascorbate:oxygen oxidoreductase [-hydroxylating]) activities in neuroblastoma clone NBD-2 from the mouse C-1300 tumor. The increase in activities was associated with a 5-10 fold increase in the V_max of both enzymes for both substrate and cofactor. Immunoprecipitation of tyrosine hydroxylase in control and treated cells demonstrated that more immunoprecipitable enzyme was present in the treated samples. Maximal elevation of both tyrosine hydroxylase and dopamine beta-hydroxylase occurred after 48 hr of treatment with 1.0 mM 8Br-cAMP. Actinomycin D (0.1 µg/ml), cycloheximide (2.0 µ/ml) and lysine-deficient medium prevented the 8Br-cAMP elevation of enzyme activity. However, removal of cycloheximide or subsequent addition of lysine (even with simultaneous addition of actinomycin D) allowed the subsequent elevation of enzyme activity. The rate of increase in enzyme activity did not exhibit the lag period typically observed in cells treated only with 8Br-cAMP. These data suggest that cAMP elevates the level of tyrosine hydroxylase and dopamine beta-hydroxylase protein and that an increased production of messenger RNA may be one of the steps necessary for these effects.