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Abstract. suppresses the labeling of messenger RNA in HeLa cells. The drug has no effect on either the labeling of nuclear heterogeneous RNA or on the transport of messenger RNA into the cytoplasm. The results suggest that messenger RNA and nuclear heterogeneous RNA are synthesized separately, and that the transcription of messenger RNA is inhibited by the drug.The nuclei of metazoan cells contain giant heterogeneous RNA. This species constitutes a major fraction of the RNA labeled during a pulse and turns over rapidly.5 Its function remains to be defined.Some properties of the heterogeneous nuclear RNA (HnRNA) resemble those expected of cytoplasmic messenger RNA (mRNA). It is heterogeneous in size, its G + C content resembles that of cellular DNA, and it apparently hybridizes to a considerable fraction of the genome (J. E. Darnell, personal communication). However, a closer examination shows that HnRNA is actually quite different from mRNA found on polyribosomes. HnRNA ranges in sedimentation values from 10 S to greater than 200 S and is much larger than the mRNA found on cytoplasmic polyribosomes (8 S-30 S).6 While mRNA is found on polyribosomes in the cytoplasm, HnRNA is confined to the nucleus where no proteinsynthesizing structures have been found, at least after cell fractionation. The rate of labeling of HnRNA is much greater than that of cytoplasmic mRNA.One hypothesis that attempts to reconcile the disparate properties of these RNA species postulates that HnRNA is a precursor to cytoplasmic mRNA and that a small region of the large molecule is cleaved and exported to the cytoplasm.7 However, the kinetics of labeling of HnRNA and mRNA are very different and do not seem to support a simple precursor-product relation6 although conclusions drawn from such measurements are far from compelling.The experiments reported here also suggest that mRNA is not derived by cleavage from HnRNA, but rather is synthesized separately. Cordycepin, 3'-deoxyadenosine, a drug that appears to act primarily by interfering with transcription, selectively suppresses the labeling of cytoplasmic mRNA while it does not affect that of HnRNA.Materials and Methods. Cell culture: HeLa cells were grown in suspension culture in Eagle's medium as described.8 The cell density was 4 X 105 cells/ml during growth and 2 X 106 cells/ml during labeling.1878

The effect of protein synthesis inhibition by cycloheximide on nucleolar RNA synthesis and processing has been studied in HeLa cells . Synthesis of 45S RNA precursor falls rapidly after administration of the drug . However, the nucleolar content of 45S RNA remains relatively constant for at least I hr because the time required for cleavage of the precursor molecule into its products is lengthened after treatment with cycloheximide. The efficiency of transformation of 45S RNA to 32S RNA remains constant with approximately one molecule of the 32S RNA produced for each cleavage of a molecule of 45S RNA . However, shortly after the cessation of protein synthesis the formation of 18S RNA becomes abortive . The amount of 32S RNA present in the nucleolus remains relatively constant . After long periods of protein synthesis inhibition the 28S RNA continues to be synthesized and exported to the cytoplasm but at a greatly reduced rate . When the protein synthesis inhibitor is removed, a prompt, although partial, recovery in the synthesis rate of 45S RNA occurs . The various aspects of RNA synthesis regulation and processing are discussed .

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