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The Effect of Actinomycin D and 5-Azacytidine on Macromolecular Synthesis in Murine Myeloma Tumor Cells

Department of Microbiology, School of Medicine, University of Washington, Seattle, Washington 98105

Abstract

A tissue culture-adapted cell line of the murine myeloma tumor MOPC 21 was employed to study the relationship between RNA and protein synthesis in myeloma tumor cells. This cell line constitutively synthesizes and secretes a large proportion of its total cellular protein as a homogeneous G immunoglobulin. Two inhibitors which function at the level of transcription, actinomycin D and 5-azacytidine, were used to obtain an estimate for the half-life of the mRNA coding for myeloma globulin. While a high concentration of actinomycin D (5.0 µg/ml) inhibited RNA synthesis by greater than 95% after one hr of incubation, a high concentration of 5-azacytidine (100 µg/ml) allowed considerable RNA synthesis for 4 hr. The half-life of the myeloma mRNA, as estimated by measuring the capacity to synthesize and secrete myeloma globulin while transcription of functional mRNA was inhibited, appeared to be of the order of 2 to 3 hr. The average half-life of the mRNA coding for total intracellular protein appeared to be similar to that of the globulin mRNA by these methods. The protein secreted in the presence of both actinomycin D and 5-azacytidine was in the form of a reduced amount of intact G globulin. Neither inhibitor showed a nonspecific effect on incorporation in a cell-free protein-synthesizing system. The implications of a relatively short-lived mRNA coding for myeloma globulin are discussed.

Footnotes

¹ Present address: Veterans Administration Hospital and Stanford University School of Medicine, Department of Medicine, 3801 Miranda Avenue, Palo Alto, California 94304. Requests for reprints should be addressed to this author.

² Present address: Department of Microbiology and Immunology, Mayo Clinic, Rochester, Minnesota.