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Regulation of Cyclic Adenosine 3':5'-Monophosphate-binding Protein in N-18 Mouse Neuroblastoma Cells

Department of Pharmacology, Harvard Medical School, Boston, Massachusetts 02115 [A. Y.-C. L., W. W. F.], and Department of Chemistry, Wright Chemistry Laboratory, Rutgers University, Piscataway, New Jersey 08854 [K. Y. C.]

The regulation of cyclic adenosine 3':5'-monophosphate (cAMP)-binding protein in N-18 neuroblastoma cells in tissue culture was studied by the covalent incorporation of 8-azidocyclic adenosine 3':5'-[³²P]monophosphate, together with the techniques of sodium dodecyl sulfate:polyacrylamide gel electrophoresis and autoradiography. Greater than 95% of the total cAMP binding activity of N-18 neuroblastoma cells was identified as being regulatory subunits of the type I (R_I) and type II (R_II) species, with R_I being the predominant form of the two (R_I:R_II = 3:1). The specific activity of R_I but not of R_II increased 3-fold when cells were grown in medium containing 1% rather than 10% fetal calf serum. Under the same conditions, the specific activity of acetylcholinesterase increased 3- to 5-fold. The increase in R_I was inversely related to the serum concentration in the medium and was specific for cells at the stationary phase of growth. An increase in intracellular cAMP, concomitant with the increase in R_I, was also observed. Morphological examination of stationary-phase neuroblastoma cells maintained in medium containing 1% fetal calf serum suggested the presence of a high proportion of highly-differentiated cells. It is proposed that the regulatory control of R_I cAMP-binding protein by serum may involve modulation of intracellular cAMP and that the expression of R_I may be used as a biochemical index of differentiation in mouse neuroblastoma cells.

¹ Recipient of Grant AM 20274 from the NIH. To whom requests for reprints should be addressed.

² Recipient of Grant CA 24479 from the NIH.

Received 6/16/80. Accepted 8/ 4/80.